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Changes in Acetyl CoA Levels during the Early Embryonic Development of Xenopus laevis
Coenzyme A (CoA) is a ubiquitous and fundamental intracellular cofactor. CoA acts as a carrier of metabolically important carboxylic acids in the form of CoA thioesters and is an obligatory component of a multitude of catabolic and anabolic reactions. Acetyl CoA is a CoA thioester derived from catabolism of all major carbon fuels. This metabolite is at a metabolic crossroads, either being further metabolised as an energy source or used as a building block for biosynthesis of lipids and cholesterol. In addition, acetyl CoA serves as the acetyl donor in protein acetylation reactions, linking metabolism to protein post-translational modifications. Recent studies in yeast and cultured mammalian cells have suggested that the intracellular level of acetyl CoA may play a role in the regulation of cell growth, proliferation and apoptosis, by affecting protein acetylation reactions. Yet, how the levels of this metabolite change in vivo during the development of a vertebrate is not known. We measured levels of acetyl CoA, free CoA and total short chain CoA esters during the early embryonic development of Xenopus laevis using HPLC. Acetyl CoA and total short chain CoA esters start to increase around midblastula transition (MBT) and continue to increase through stages of gastrulation, neurulation and early organogenesis. Pre-MBT embryos contain more free CoA relative to acetyl CoA but there is a shift in the ratio of acetyl CoA to CoA after MBT, suggesting a metabolic transition that results in net accumulation of acetyl CoA. At the whole-embryo level, there is an apparent correlation between the levels of acetyl CoA and levels of acetylation of a number of proteins including histones H3 and H2B. This suggests the level of acetyl CoA may be a factor, which determines the degree of acetylation of these proteins, hence may play a role in the regulation of embryogenesis.
Tsuchiya, Yugo; Pham, Uyen; Hu, Wanzhou; Ohnuma, Shin-ichi; Gout, Ivan
Soraphen A, an inhibitor of acetyl CoA carboxylase activity, interferes with fatty acid elongation.
Acetyl CoA carboxylase (ACC1 and ACC2) generates malonyl CoA, a substrate for de novo lipogenesis (DNL) and an inhibitor of mitochondrial fatty acid ?-oxidation (FAO). Malonyl CoA is also a substrate for microsomal fatty acid elongation, an important pathway for saturated (SFA), mono- (MUFA) and polyunsaturated fatty acid (PUFA) synthesis. Despite the interest in ACC as a target for obesity and cancer therapy, little attention has been given to the role ACC plays in long chain fatty acid synthesis. This report examines the effect of pharmacological inhibition of ACC on DNL and palmitate (16:0) and linoleate (18:2, n-6) metabolism in HepG2 and LnCap cells. The ACC inhibitor, soraphen A, lowers cellular malonyl CoA, attenuates DNL and the formation of fatty acid elongation products derived from exogenous fatty acids, i.e., 16:0 and 18:2, n-6; IC(50)?5nM. Elevated expression of fatty acid elongases (Elovl5, Elovl6) or desaturases (FADS1, FADS2) failed to override the soraphen A effect on SFA, MUFA or PUFA synthesis. Inhibition of fatty acid elongation leads to the accumulation of 16- and 18-carbon unsaturated fatty acids derived from 16:0 and 18:2, n-6, respectively. Pharmacological inhibition of ACC activity will not only attenuate DNL and induce FAO, but will also attenuate the synthesis of very long chain saturated, mono- and polyunsaturated fatty acids. PMID:21184748
Jump, Donald B; Torres-Gonzalez, Moises; Olson, L Karl
Soraphen A, an inhibitor of acetyl CoA carboxylase activity, interferes with fatty acid elongation
Acetyl CoA carboxylase (ACC1 & ACC2) generates malonyl CoA, a substrate for de novo lipogenesis (DNL) and an inhibitor of mitochondrial fatty acid ?-oxidation (FAO). Malonyl CoA is also a substrate for microsomal fatty acid elongation, an important pathway for saturated (SFA), mono- (MUFA) and polyunsaturated fatty acid (PUFA) synthesis. Despite the interest in ACC as a target for obesity and cancer therapy, little attention has been given to the role ACC plays in long chain fatty acid synthesis. This report examines the effect of pharmacological inhibition of ACC on DNL & palmitate (16:0) and linoleate (18:2,n-6) metabolism in HepG2 and LnCap cells. The ACC inhibitor, soraphen A, lowers cellular malonyl CoA, attenuates DNL and the formation of fatty acid elongation products derived from exogenous fatty acids, i.e., 16:0 & 18:2,n-6; IC50 ~ 5 nM. Elevated expression of fatty acid elongases (Elovl5, Elovl6) or desaturases (FADS1, FADS2) failed to override the soraphen A effect on SFA, MUFA or PUFA synthesis. Inhibition of fatty acid elongation leads to the accumulation of 16- and 18-carbon unsaturated fatty acids derived from 16:0 and 18:2,n-6, respectively. Pharmacological inhibition of ACC activity will not only attenuate DNL and induce FAO, but will also attenuate the synthesis of very long chain saturated, mono- and polyunsaturated fatty acids.
Jump, Donald B.; Torres-Gonzalez, Moises; Olson, L. Karl
Materials and methods for the alteration of enzyme and acetyl CoA levels in plants
The present invention provides nucleic acid and amino acid sequences of acetyl CoA synthetase (ACS), plastidic pyruvate dehydrogenase (pPDH), ATP citrate lyase (ACL), Arabidopsis pyruvate decarboxylase (PDC), and Arabidopsis aldehyde dehydrogenase (ALDH), specifically ALDH-2 and ALDH-4. The present invention also provides a recombinant vector comprising a nucleic acid sequence encoding one of the aforementioned enzymes, an antisense sequence thereto or a ribozyme therefor, a cell transformed with such a vector, antibodies to the enzymes, a plant cell, a plant tissue, a plant organ or a plant in which the level of an enzyme has been altered, and a method of producing such a plant cell, plant tissue, plant organ or plant. Desirably, alteration of the level of enzyme results in an alteration of the level of acetyl CoA in the plant cell, plant tissue, plant organ or plant. In addition, the present invention provides a recombinant vector comprising an antisense sequence of a nucleic acid sequence encoding pyruvate decarboxylase (PDC), the E1.alpha. subunit of pPDH, the E1.beta. subunit of pPDH, the E2 subunit of pPDH, mitochondrial pyruvate dehydrogenase (mtPDH) or aldehyde dehydrogenase (ALDH) or a ribozyme that can cleave an RNA molecule encoding PDC, E1.alpha. pPDH, E1.beta. pPDH, E2 pPDH, mtPDH or ALDH.
Nikolau, Basil J. (Ames, IA); Wurtele, Eve S. (Ames, IA); Oliver, David J. (Ames, IA); Schnable, Patrick S. (Ames, IA); Wen, Tsui-Jung (Ames, IA)
Acetyl CoA Carboxylase: Isolation and Characterization of Native Biotin Carboxyl Carrier Protein
A large form of biotin carboxyl carrier protein (BCCPL) has been isolated from extracts of Escherichia coli. It has a minimal molecular weight of 20,000, according to its behavior on sodium dodecylsulfate-polyacrylamide gel electrophoresis, and contains approximately 1 mol of biotin per 22,000 g of protein. BCCPL exhibits Km values, in the biotin carboxylase and transcarboxylase half-reactions of acetyl CoA carboxylase, of 2 � 10-7 M and 4 � 10-7 M, respectively; these values are 50-100 times lower than those obtained with smaller forms of BCCP previously isolated. Electrophoresis of crude extracts of E. coli indicates that the major biotin-containing protein migrates at the same rate as BCCPL, which suggests that BCCPL is the native form of BCCP in E. coli. Images
Fall, R. Ray; Nervi, A. M.; Alberts, Alfred W.; Vagelos, P. Roy
Apicoplast acetyl Co-A carboxylase of the human malaria parasite is not targeted by cyclohexanedione herbicides.
Malaria parasites retain a relict plastid (apicoplast) from a photosynthetic ancestor. The apicoplast is a useful drug target but the specificity of compounds believed to target apicoplast fatty acid biosynthesis has become uncertain, as this pathway is not essential in blood stages of the parasite. Herbicides that inhibit the plastid acetyl Coenzyme A (Co-A) carboxylase of plants also kill Plasmodium falciparum in vitro, but their mode of action remains undefined. We characterised the gene for acetyl Co-A carboxylase in P. falciparum. The P. falciparum acetyl-CoA carboxylase gene product is expressed in blood stage parasites and accumulates in the apicoplast. Ablation of the gene did not render parasites insensitive to herbicides, suggesting that these compounds are acting off-target in blood stages of P. falciparum. PMID:24583112
Goodman, Christopher D; Mollard, Vanessa; Louie, Theola; Holloway, Georgina A; Watson, Keith G; McFadden, Geoffrey I
Correlation of ATP Citrate Lyase and Acetyl CoA Levels with Trichothecene Production in Fusarium graminearum
Thecorrelation of ATP citrate lyase (ACL) and acetyl CoA levels with trichothecene production in Fusarium graminearum was investigated using an inhibitor (precocene II) and an enhancer (cobalt chloride) of trichothecene production by changing carbon sources in liquid medium. When precocene II (30 �M) was added to inhibit trichothecene production in a trichothecene high-production medium containing sucrose, ACL expression was reduced and ACL mRNA level as well as acetyl CoA amount in the fungal cells were reduced to the levels observed in a trichothecene trace-production medium containing glucose or fructose. The ACL mRNA level was greatly increased by addition of cobalt chloride in the trichothecene high-production medium, but not in the trichothecene trace-production medium. Levels were reduced to those level in the trichothecene trace-production medium by addition of precocene II (300 �M) together with cobalt chloride. These results suggest that ACL expression is activated in the presence of sucrose and that acetyl CoA produced by the increased ALC level may be used for trichothecene production in the fungus. These findings also suggest that sucrose is important for the action of cobalt chloride in activating trichothecene production and that precocene II may affect a step down-stream of the target of cobalt chloride.
Sakamoto, Naoko; Tsuyuki, Rie; Yoshinari, Tomoya; Usuma, Jermnak; Furukawa, Tomohiro; Nagasawa, Hiromichi; Sakuda, Shohei
Correlation of ATP citrate lyase and acetyl CoA levels with trichothecene production in Fusarium graminearum.
The correlation of ATP citrate lyase (ACL) and acetyl CoA levels with trichothecene production in Fusarium graminearum was investigated using an inhibitor (precocene II) and an enhancer (cobalt chloride) of trichothecene production by changing carbon sources in liquid medium. When precocene II (30 �M) was added to inhibit trichothecene production in a trichothecene high-production medium containing sucrose, ACL expression was reduced and ACL mRNA level as well as acetyl CoA amount in the fungal cells were reduced to the levels observed in a trichothecene trace-production medium containing glucose or fructose. The ACL mRNA level was greatly increased by addition of cobalt chloride in the trichothecene high-production medium, but not in the trichothecene trace-production medium. Levels were reduced to those level in the trichothecene trace-production medium by addition of precocene II (300 �M) together with cobalt chloride. These results suggest that ACL expression is activated in the presence of sucrose and that acetyl CoA produced by the increased ALC level may be used for trichothecene production in the fungus. These findings also suggest that sucrose is important for the action of cobalt chloride in activating trichothecene production and that precocene II may affect a step down-stream of the target of cobalt chloride. PMID:24284828
Sakamoto, Naoko; Tsuyuki, Rie; Yoshinari, Tomoya; Usuma, Jermnak; Furukawa, Tomohiro; Nagasawa, Hiromichi; Sakuda, Shohei
Hydrolysis and transpeptidation of peptide substrates by acetyl-pepsin.
Treatment of swine pepsin with acetylimidazole to acetylate approximately five of its 16 tyrosyl residues causes a significant enhancement of catalytic efficiency (kcat/Km) toward substrates such as dansyl-glycyl-glycyl-L-phenylalanyl-L-phenylalanine 3-(4-pyridyl)propyl ester and benzyloxy-carbonyl-(glycyl)n-p-nitroLphenylalnyl-Lphenylalanyl-L-tyrosine (where n = 0, 1,2). Stopped-flow kinetic studies, under conditions of enzyme excess, with the dansyl peptide have shown that, as with untreated pepsin, the rate-limiting step in the over-all catalytic process is associated with the decomposition of the first detectable enzyme-substrate complex, whose dissociation constant is approximately equal to the Km found in steady-state kinetic experiments. With substrates of the type benzoyl-(glycyl)n-nitro-L-phenylalanyl-L-tyrosine, an increase in the chain length of the peptide leads to an increase in the value of kcat/Km, supporting the view that secondary enzyme-substrate interactions may produce at the extended active site conformational changes that are reflected in higher catalytic efficiency. This effect is more marked with acetyl-pepsin than with untreated pepsin, and suggests that the conformational mobility of the active site is increased by partial acetylation. Acetyl-pepsin is less effective than untreated pepsin in catalyzing transpeptidation reactions in which acetyl-L-phenylalanyl-L-tyrosine and benzyloxycarbonyl-(glycyl)n-p-nitro-L-phenylalanine are the reactants; this finding is consistent with the more rapid hydrolysis of the product of transpeptidation.
Richman, P G; Fruton, J S
Enzymatic Carboxylation of Biotin: Molecular and Catalytic Properties of a Component Enzyme of Acetyl CoA Carboxylase*
The biotin carboxylase component of acetyl CoA carboxylase has been purified approximately 2000 times from Escherichia coli. This protein, which catalyzes the carboxylation of free d-biotin, is free of the biotin-containing carboxyl carrier protein, is homogeneous by polyacrylamide gel electrophoresis and analytical ultracentrifugation, and has been crystallized. Biotin carboxylase, with a molecular weight of approximately 100,000, is composed of two 50,000-dalton subunits. The catalytic capacity of biotin carboxylase is markedly enhanced by ethanol (11 times at 15% v/v), and certain other organic solvents; this may mimic an effector-mediated response. The kinetic effect is exclusively on the maximal velocity of the reaction. Activation by ethanol is reversible and not accompanied by aggregation or disaggregation of the enzyme. Images
Dimroth, Peter; Guchhait, Ras B.; Stoll, Erwin; Lane, M. Daniel
Determination of the quantity of acetyl CoA carboxylase by (/sup 14/C)methyl avidin binding
Conditions are described under which monomeric (/sup 14/C)methyl avidin binds to SDS-denatured biotin enzymes and remains bound through polyacrylamide gel electrophoresis. The location of radioactive proteins on the dried gel was determined by fluorography and their identity was established by subunit molecular weight. The relative quantity of bound radioactive avidin, stoichiometrically equivalent to the molar quantity of biotin protein, can be determined by scanning the fluorograph with a soft laser densitometer. To determine the absolute quantity of biotin protein, the radioactive areas of the dried gel were cut out, resolubilized, and assayed for radioactivity. Since the specific radioactivity of the (/sup 14/C)methyl avidin was known, the quantity of avidin bound and therefore the quantity of biotin enzyme could be calculated. The method is illustrated by the analysis of purified acetyl CoA carboxylase and is applied to the analysis of biotin enzymes in isolated rat liver mitochondria.
Roman-Lopez, C.R.; Goodson, J.; Allred, J.B.
Kinetics of acetyl CoA: Arylamine N?acetyltransferase from rapid and slow acetylator shrimp livers
N?acetyltransferase (NAT) activity was determined in 200 shrimps (Penaeus chinensis) livers using 2?aminofluorene (AF) and p?aminobenzoic acid (PABA) as substrates. Overall, the liver NAT activity of the 100 females was higher than the liver NAT activity of the 100 males. The activities (mean � S.D.) of NAT from liver of males was 1.27 � 0.52 nmol\\/min\\/mg protein for the acetylation
Jing G. Chung; Jau H. Lee; Heng C. Ho; Chin C. Ho; Jem M. Lai; Shih H. Chang; Chi F. Hung
Aberrant mitosis in fission yeast mutants defective in fatty acid synthetase and acetyl CoA carboxylase
Two fission yeast temperature-sensitive mutants, cut6 and lsd1, show a defect in nuclear division. The daughter nuclei differ dramatically in size (the phenotype designated lsd, large and small daughter). Fluorescence in situ hybridization (FISH) revealed that sister chromatids were separated in the lsd cells, but appeared highly compact in one of the two daughter nuclei. EM showed asymmetric nuclear elongation followed by unequal separation of nonchromosomal nuclear structures in these mutant nuclei. The small nuclei lacked electron- dense nuclear materials and contained highly compacted chromatin. The cut6+ and lsd1+ genes are essential for viability and encode, respectively, acetyl CoA carboxylase and fatty acid synthetase, the key enzymes for fatty acid synthesis. Gene disruption of lsd1+ led to the lsd phenotype. Palmitate in medium fully suppressed the phenotypes of lsd1. Cerulenin, an inhibitor for fatty acid synthesis, produced the lsd phenotype in wild type. The drug caused cell inviability during mitosis but not during the G2-arrest induced by the cdc25 mutation. A reduced level of fatty acid thus led to impaired separation of non- chromosomal nuclear components. We propose that fatty acid is directly or indirectly required for separating the mother nucleus into two equal daughters.
Dinuclear nickel complexes modeling the structure and function of the acetyl CoA synthase active site
A dinuclear nickel complex with methyl and thiolate ligands, Ni(dadtEt)Ni(Me)(SDmp) (2), has been synthesized as a dinuclear Nid�Nip-site model of acetyl-CoA synthase (ACS) (dadtEt is N,N?-diethyl-3,7-diazanonane-1,9-dithiolate; Dmp is 2,6-dimesitylphenyl). Complex 2 was prepared via 2 methods: (i) ligand substitution of a dinuclear Ni(II)�Ni(II) cation complex [Ni(dadtEt) Ni(tmtu)2] (OTf)2(1) with MeMgBr and KSDmp (tmtu is tetramethylthiourea), (ii) methyl transfer from methylcobaloxime Co(dmgBF2)2(Me)(Py) (5) to a Ni(II)�Ni(0) complex such as [Ni(dadtEt)Ni(cod)] (3), generated in situ from Ni(dadtEt) and Ni(cod)2, followed by addition of KSDmp (cod is 1,5-cyclooctadiene; dmgBF2 is difluoroboryl-dimethylglyoximate). Method ii models the formation of Nip�Me species proposed as a plausible intermediate in ACS catalysis. The reaction of 2 with excess CO affords the acetylthioester CH3C(O)SDmp (8) with concomitant formation of Ni(dadtEt)Ni(CO)2 (9) and Ni(CO)4 plus Ni(dadtEt). When complex 2 is treated with 1 equiv of CO in the presence of excess 1,5-cyclooctadiene, the formation of 9 and Ni(CO)4 is considerably suppressed, and instead the dinuclear Ni(II)�Ni(0) complex is generated in situ, which further affords 2 upon successive treatment with Co(dmgBF2)2(Me)(Py) (5) and KSDmp. These results suggest that (i) ACS catalysis could include the Nid(II)�Nip(0) state as the active species, (ii) The Nid(II)�Nip(0) species could first react with methylcobalamin to afford Nid(II)�Nip(II)�Me, and (iii) CO insertion into the Nip�Me bond and the successive reductive elimination of acetyl-CoA occurs immediately when CoA is coordinated to the Nip site to form the active Nid(II)�Nip(0) species.
Ito, Mikinao; Kotera, Mai; Matsumoto, Tsuyoshi; Tatsumi, Kazuyuki
Pig Liver Esterase Catalyzed Hydrolysis of Methyl 2,3-Di-O-Acetyl-5-Deoxy-?- and ?-D-Arabinofuranosides
The regioselectivity of a pig liver esterase (PLE) catalyzed hydrolysis of methyl 2,3-di-O-acetyl-5-deoxy-?-D-arabinofuranoside (1) and methyl 2,3-di-O-acetyl-5-deoxy-?-D-arabinofuranoside (2) was established by GLC. Diacetate 1 gave exclusively methyl 3-O-acetyl-5-deoxy-?-D-arabinofuranoside while diacetate 2 produced both methyl 2-O-acetyl-5-deoxy-?-D-arabinofuranoside and methyl 3-O-acetyl-5-deoxy-?-D-arabinofuranoside which were resistant to subsequent hydrolysis. The Michaelis constants and maximal velocities were determined for 1 and 2. The first-order rate constants
Jitka Moravcov�; Ivan Hamern�k; Gabriela Funkov�; Jindra ?apkov�; Karel Kefurt
Chromium downregulates the expression of Acetyl CoA Carboxylase 1 gene in lipogenic tissues of domestic goats: a potential strategy for meat quality improvement.
Acetyl CoA Carboxylase 1 (ACC1) is a biotin-dependent enzyme that catalyzes the carboxylation of Acetyl CoA to form Malonyl CoA, the key intermediate metabolite in fatty acid synthesis. In this study, the mRNA expression of the ACC1 gene was evaluated in four different tissues (liver, visceral fat, subcutaneous fat, and longissimus muscle) of the domestic goat (Capra hircus) kids feeding on four different levels of trivalent chromium (0, 0.5, 1, and 1.5mg/day) as food supplementation. RT-qPCR technique was used for expression analyses and heat shock protein 90 gene (HSP-90) was considered as reference gene for data normalization. Our results revealed that 1.5mg/day chromium significantly reduced the expression of the ACC1 gene in liver, visceral fat, and subcutaneous fat tissues, but not in longissimus muscles (P<0.05). We measured some phenotypic traits of kid's carcasses to detect their probable correlations with chromium-mediated downregulation of ACC1 expression. Interestingly, changes in ACC1 expression were accompanied with decreased accumulation of fats in adipose tissues such that the subcutaneous fat thickness and heart fat percentage decreased in kids feeding on chromium. By contrast, chromium supplemented kids showed higher percentage of muscles despite the fact that their total body weight did not differ from that of non-supplemented kids. Our study suggests that trivalent chromium alters the direction of energy accumulation towards muscles rather than fats and provides insights into application of chromium supplementation as a useful strategy for improvement of meat quality in domestic animals. PMID:24704275
Najafpanah, Mohammad Javad; Sadeghi, Mostafa; Zali, Abolfazl; Moradi-Shahrebabak, Hossein; Mousapour, Hojatollah
Simple rapid hydrolysis of acetyl protecting groups in the FDG synthesis using cation exchange resins
A new solid phase method for hydrolysis of acetyl groups from the intermediate 2-[18F]fluoro-2-deoxy-glucose tetraacetate ([18F]FDG-Ac4) was developed. Fast cleavage occurs with Dowex 50 sulfonic acid resin (H+ form) at ~100 �C in the absence of bulk water. [18F]FDG-Ac4 reaction mixtures were efficiently converted to neutral aqueous solutions of pure FDG in 10�15 min. The method avoids problems associated with
G. Keith Mulholland
Kinetics of Acid Hydrolysis of Water-Soluble Spruce O-Acetyl Galactoglucomannans
Water-soluble O-acetyl galactoglucomannan (GGM) is a softwood-derived polysaccharide, which can be extracted on an industrial scale from wood or mechanical pulping waters and now is available in kilogram scale for research and development of value-added products. To develop applications of GGM, information is needed on its stability in acidic conditions. The kinetics of acid hydrolysis of GGM was studied at
Chunlin Xu; Andrey Pranovich; L. Vahasalo; Jarl Hemming; Bjarne Holmbom; Henk A. Schols; S. Willfor
The role of acetyl xylan esterase in the solubilization of xylan and enzymatic hydrolysis of wheat straw and giant reed
Background Due to the complexity of lignocellulosic materials, a complete enzymatic hydrolysis into fermentable sugars requires a variety of cellulolytic and xylanolytic enzymes. Addition of xylanases has been shown to significantly improve the performance of cellulases and to increase cellulose hydrolysis by solubilizing xylans in lignocellulosic materials. The goal of this work was to investigate the effect of acetyl xylan esterase (AXE) originating from Trichoderma reesei on xylan solubilization and enzymatic hydrolysis of cellulose. Results The solubilization of xylan in pretreated wheat straw and giant reed (Arundo donax) by xylanolytic enzymes and the impact of the sequential or simultaneous solubilization of xylan on the hydrolysis of cellulose by purified enzymes were investigated. The results showed that the removal of acetyl groups in xylan by AXE increased the accessibility of xylan to xylanase and improved the hydrolysis of xylan in pretreated wheat straw and giant reed. Solubilization of xylan led to an increased accessibility of cellulose to cellulases and thereby increased the hydrolysis extent of cellulose. A clear synergistic effect between cellulases and xylanolytic enzymes was observed. The highest hydrolysis yield of cellulose was obtained with a simultaneous use of cellulases, xylanase and AXE, indicating the presence of acetylated xylan within the cellulose matrix. Acetylated xylobiose and acetylated xylotriose were produced from xylan without AXE, as confirmed by atmospheric pressure matrix-assisted laser desorption/ionization ion trap mass spectrometry. Conclusions The results in this paper demonstrate that supplementation of xylanase with AXE enhances the solubilization of xylan to some extent and, consequently, increases the subsequent hydrolysis of cellulose. The highest hydrolysis yield was, however, obtained by simultaneous hydrolysis of xylan and cellulose, indicating a layered structure of cellulose and xylan chains in the cell wall substrate. AXE has an important role in the hydrolysis of lignocellulosic materials containing acetylated xylan.
Kinetics of acid hydrolysis of water-soluble spruce O-acetyl galactoglucomannans.
Water-soluble O-acetyl galactoglucomannan (GGM) is a softwood-derived polysaccharide, which can be extracted on an industrial scale from wood or mechanical pulping waters and now is available in kilogram scale for research and development of value-added products. To develop applications of GGM, information is needed on its stability in acidic conditions. The kinetics of acid hydrolysis of GGM was studied at temperatures up to 90 degrees C in the pH range of 1-3. Molar mass and molar mass distribution were determined using size exclusion chromatography with multiangle laser light scattering and refractive index detection. The molar mass of GGM decreased considerably with treatment time at temperatures above 70 degrees C and pH below 2. The molar mass distribution broadened with hydrolysis time. A first-order kinetic model was found to match the acid hydrolysis. The reaction rate constants at various pH values and temperatures were calculated on the basis of the first-order kinetic model. Furthermore, the activation energy, E, was obtained from the Arrhenius plot. The activation energy E was 150 kJ mol (-1) for acid hydrolysis of spruce GGM. The apparent rate constant during acid hydrolysis increased by a factor of 10 with a decrease in pH by 1 unit, regardless of temperature. In addition, gas chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry were applied to study the released GGM monomers and oligomers. PMID:18333617
Xu, Chunlin; Pranovich, Andrey; V�h�salo, Lari; Hemming, Jarl; Holmbom, Bjarne; Schols, Henk A; Willf�r, Stefan
2-Acetylthiamin pyrophosphate (acetyl-TPP) pH-rate profile for hydrolysis of acetyl-TPP and isolation of acetyl-TPP as a transient species in pyruvate dehydrogenase catalyzed reactions
Rate constants for the hydrolysis of acetyl-TPP were measured pH values of 2.5 and 7.5 and plotted as log k{sub obs} versus pH. The pH-rate profile defined two legs, each with a slope of +1 but separated by a region of decreased slope between pH 4 and pH 6. The rates were insensitive to buffer concentrations. Each leg of the profile reflected specific-base-catalyzed hydrolysis of acetyl-TPP, analogous to the hydrolysis of 2-acetyl-3,4-dimethylthiazolium ion. The separation of the two legs of this profile has been shown to be caused by the ionization of a group exhibiting a pK{sub a} of 4.73 within acetyl-TPP that is remote from the acetyl group, the aminopyrimidine ring, which is promoted below pH 4.73. The protonation level of this ring has been shown to control the equilibrium partitioning of acetyl-TPP among its carbinolamine, keto, and hydrate forms. The differential partitioning of these species is a major factor causing the separation between the two legs of the pH-rate profile. The characteristic pH-rate profile and the availability of synthetic acetyl-TPP have facilitated the isolation and identification of (1-{sup 14}C)acetyl-TPP from acid-quenched enymatic reaction mixtures at steady states. (1-{sup 14}C)Acetyl-TPP was identified as a transient species in reactions catalyzed by the PDH complex or the pyruvate dehydrogenase component of the complex (E{sub 1}). The pH-rate profile for hydrolysis of (1-{sup 14}C)-acetyl-TPP, isolated from enzymatic reactions was found to be indistinguishable from that for authentic acetyl-TPP, which constituted positive identification of the {sup 14}C-labeled enzymic species.
Gruys, K.J.; Datta, A.; Frey, P.A. (Univ. of Wisconsin, Madison (USA))
Simvastatin Induced Neurite Outgrowth Unveils Role of Cell Surface Cholesterol and Acetyl CoA Carboxylase in SH-SY5Y Cells
Statins are known to modulate cell surface cholesterol (CSC) and AMP-activated protein kinase (AMPK) in non-neural cells; however no study demonstrates whether CSC and AMPK may regulate simvastatin induced neuritogenesis (SIN). We found that simvastatin (SIM) maintains CSC as shown by Fillipin III staining, Flotillin-2 protein expression / localization and phosphorylation of various receptor tyrosine kinases (RTKs) in the plasma membrane. Modulation of CSC revealed that SIN is critically dependent on this CSC. Simultaneously, phospho array for mitogen activated protein kinases (MAPKs) revealed PI3K / Akt as intracellular pathway which modulates lipid pathway by inhibiting AMPK activation. Though, SIM led to a transient increase in AMPK phosphorylation followed by a sudden decline; the effect was independent of PI3K. Strikingly, AMPK phosphorylation was regulated by protein phosphatase 2A (PP2A) activity which was enhanced upon SIM treatment as evidenced by increase in threonine phosphorylation. Moreover, it was observed that addition of AMP analogue and PP2A inhibitor inhibited SIN. Bio-composition of neurites shows that lipids form a major part of neurites and AMPK is known to regulate lipid metabolism majorly through acetyl CoA carboxylase (ACC). AMPK activity is negative regulator of ACC activity and we found that phosphorylation of ACC started to decrease after 6 hrs which becomes more pronounced at 12 hrs. Addition of ACC inhibitor showed that SIN is dependent on ACC activity. Simultaneously, addition of Fatty acid synthase (FAS) inhibitor confirmed that endogenous lipid pathway is important for SIN. We further investigated SREBP-1 pathway activation which controls ACC and FAS at transcriptional level. However, SIM did not affect SREBP-1 processing and transcription of its target genes likes ACC1 and FAS. In conclusion, this study highlights a distinct role of CSC and ACC in SIN which might have implication in process of neuronal differentiation induced by other agents.
Raina, Varshiesh; Gupta, Sarika; Yadav, Saurabh; Surolia, Avadhesha
Determination of the degree of substitution of acetylated starch by hydrolysis, 1H NMR and TGA\\/IR
The suitability of TGA\\/IR method for the analysis of the degree of substitution (DS) of acetylated starch (SA) was assessed. Ten standard samples were analysed for their DS by hydrolysis method and, those of the standards which could be properly dissolved, also by 1H NMR. NMR analysis shows as well that the standards had other structural differences than DS. By
Matti Elomaa; Tomas Asplund; Pasi Soininen; Reino Laatikainen; Soili Peltonen; Sari Hyv�rinen; Arto Urtti
From Arylamine N-Acetyltransferase to Folate-Dependent Acetyl CoA Hydrolase: Impact of Folic Acid on the Activity of (HUMAN)NAT1 and Its Homologue (MOUSE)NAT2
Acetyl Coenzyme A-dependent N-, O- and N,O-acetylation of aromatic amines and hydrazines by arylamine N-acetyltransferases is well characterised. Here, we describe experiments demonstrating that human arylamine N-acetyltransferase Type 1 and its murine homologue (Type 2) can also catalyse the direct hydrolysis of acetyl Coenzyme A in the presence of folate. This folate-dependent activity is exclusive to these two isoforms; no acetyl Coenzyme A hydrolysis was found when murine arylamine N-acetyltransferase Type 1 or recombinant bacterial arylamine N-acetyltransferases were incubated with folate. Proton nuclear magnetic resonance spectroscopy allowed chemical modifications occurring during the catalytic reaction to be analysed in real time, revealing that the disappearance of acetyl CH3 from acetyl Coenzyme A occurred concomitantly with the appearance of a CH3 peak corresponding to that of free acetate and suggesting that folate is not acetylated during the reaction. We propose that folate is a cofactor for this reaction and suggest it as an endogenous function of this widespread enzyme. Furthermore, in silico docking of folate within the active site of human arylamine N-acetyltransferase Type 1 suggests that folate may bind at the enzyme�s active site, and facilitate acetyl Coenzyme A hydrolysis. The evidence presented in this paper adds to our growing understanding of the endogenous roles of human arylamine N-acetyltransferase Type 1 and its mouse homologue and expands the catalytic repertoire of these enzymes, demonstrating that they are by no means just xenobiotic metabolising enzymes but probably also play an important role in cellular metabolism. These data, together with the characterisation of a naphthoquinone inhibitor of folate-dependent acetyl Coenzyme A hydrolysis by human arylamine N-acetyltransferase Type 1/murine arylamine N-acetyltransferase Type 2, open up a range of future avenues of exploration, both for elucidating the developmental role of these enzymes and for improving chemotherapeutic approaches to pathological conditions including estrogen receptor-positive breast cancer.
Laurieri, Nicola; Dairou, Julien; Egleton, James E.; Stanley, Lesley A.; Russell, Angela J.; Dupret, Jean-Marie; Sim, Edith; Rodrigues-Lima, Fernando
An acetylglucomannan esterase of Aspergillus oryzae; purification, characterization and role in the hydrolysis of O-acetyl-galactoglucomannan.
An acetyl glucomannan esterase (AGME) was purified to electrophoretic homogeneity from the culture supernatant of Aspergillus oryzae. This new enzyme had a molecular mass of 36 kDa and an isoelectric point of 4.6. It was most active in the pH range 5.0-5.5 and was stable for 24 h at 40 degrees C at pH 5.0-6.0. The purified esterase liberated acetic acid from O-acetyl-galactoglucomannan, O-acetyl-4-O- methylglucuronoxylan and alpha-naphtyl acetate. The specific activity was 10-times higher for acetylated mannan than for acetylated xylan. The enzyme was able to act on polymeric substrate but activity was clearly enhanced by addition of mannanase from Trichoderma reesei and alpha-galactosidase from guar seeds. Presence of mannanase also increased the liberation of acetic acid in long-term hydrolysis (24 h), while the addition of alpha-galactosidase had no effect. No significant synergism between these two glycanases and the previously characterized esterase of A. oryzae (FE), which is also able to deacetylate galactoglucomannan, was observed. Even though the AGME had 8-times higher specific galactomannan deacetylating activity than the FE, the maximum amount of acetic acid liberated from the polymeric galactoglucomannan by AGME was only 80% of that of FE. Both esterases clearly enhanced the action of mannanase and alpha-galactosidase in the degradation of O-acetyl-galactoglucomannan isolated from Norway spruce. PMID:7576539
Tenkanen, M; Thornton, J; Viikari, L
The hydrolysis of the O-glycosidic linkages (depolymerization) and the N-acetyl linkage (de-N-acetylation) of partially N-acetylated chitosans were studied in dilute and concentrated HCl. The rate of hydrolysis of the glycosidic linkages was found to be equal to the rate of de-N-acetylation in dilute acid, while the glycosidic linkages was hydrolysed more than 10 times faster than the N-acetyl linkage in
K. M. V�rum; M. H. Ott�y; O. Smidsr�d
Presence of Acetyl Coenzyme A (CoA) Carboxylase and Propionyl-CoA Carboxylase in Autotrophic Crenarchaeota and Indication for Operation of a 3-Hydroxypropionate Cycle in Autotrophic Carbon Fixation
The pathway of autotrophic CO2 fixation was studied in the phototrophic bacterium Chloroflexus aurantiacus and in the aerobic thermoacidophilic archaeon Metallosphaera sedula. In both organisms, none of the key enzymes of the reductive pentose phosphate cycle, the reductive citric acid cycle, and the reductive acetyl coenzyme A (acetyl-CoA) pathway were detectable. However, cells contained the biotin-dependent acetyl-CoA carboxylase and propionyl-CoA
Effect of the alpha-methyl substituent on chemoselectivity in esterase-catalyzed hydrolysis of S-acetyl sulfanylalkanoates.
The isomeric compounds 1 and 3, which differ only in the position of a methyl substituent, give opposite chemoselectivities in an esterase-catalyzed hydrolysis reaction. The esterase was chemoselective for the oxoester in 1, but for the thiol ester group in 3. A high enantioselectivity was observed for both 1 and 3. PMID:10905865
Kumar, I; Jolly, R S
An acetylglucomannan esterase of Aspergillus oryzae; purification, characterization and role in the hydrolysis of O-acetyl-galactoglucomannan
An acetyl glucomannan esterase (AGME) was purified to eiectrophoretic homogeneity from the culture supernatant of Aspergillus oryzae. This new enzyme had a molecular mass of 36 kDa and an isoelectric point of 4.6. It was most active in the pH range 5.0�5.5 and was stable for 24 h at 40 �C at pH 5.0�6.0. The purified esterase liberated acetic acid
Maija Tenkanen; Jeff Thornton; Liisa Viikari
Synthesis and Kinetics of Hydrolysis of 3,5-Dimethyl-N-acetyl-p-benzoquinone Imine: An Undergraduate Laboratory
NASA Astrophysics Data System (ADS)
The synthesis of the title compound by a three-step procedure is described. The hydrolysis kinetics, which involve two consecutive psuedo-first-order processes, are also described. The synthesis and kinetics experiments described here are proposed for incorporation into undergraduate laboratory courses under a variety of formats. The compound described here is related to a toxic metabolite of the common analgesics acetaminophen and phenacetin.
Buccigross, Jeanne M.; Metz, Christa; Elliot, Lori; Becker, Pamela; Earley, Angela S.; Hayes, Jerry W.; Novak, Michael; Underwood, Gayl A.
Physicochemical characterisation of enzymatically hydrolysed derivatives of acetylated starch
Potato starch modified to different degrees by substitution with acetyl groups was the subject of this study undertaken to determine the influence of conditions of enzymatic hydrolysis on the surface-active properties of hydrolysates of acetylated starch. The effect of acetylation of starch preparation on its susceptibility to enzymatic hydrolysis in the membrane reactor was also considered. All hydrolysates of acetylated
Emilia Konowa?; Gra?yna Lewandowicz; Joanna Le Thanh-Blicharz; Krystyna Prochaska
Nucleosome acetylation sequencing to study the establishment of chromatin acetylation.
The establishment of posttranslational chromatin modifications is a major mechanism for regulating how genomic DNA is utilized. However, current in vitro chromatin assays do not monitor histone modifications at individual nucleosomes. Here we describe a strategy, nucleosome acetylation sequencing, that allows us to read the amount of modification at each nucleosome. In this approach, a bead-bound trinucleosome substrate is enzymatically acetylated with radiolabeled acetyl CoA by the SAGA complex from Saccharomyces cerevisae. The product is digested by restriction enzymes that cut at unique sites between the nucleosomes and then counted to quantify the extent of acetylation at each nucleosomal site. We find that we can sensitively, specifically, and reproducibly follow enzyme-mediated nucleosome acetylation. Applying this strategy, when acetylation proceeds extensively, its distribution across nucleosomes is relatively uniform. However, when substrates are used that contain nucleosomes mutated at the major sites of SAGA-mediated acetylation, or that are studied under initial rate conditions, changes in the acetylation distribution can be observed. Nucleosome acetylation sequencing should be applicable to analyzing a wide range of modifications. Additionally, because our trinucleosomes synthesis strategy is highly modular and efficient, it can be used to generate nucleosomal systems in which nucleosome composition differs across the array. PMID:24769374
Mittal, Chitvan; Blacketer, Melissa J; Shogren-Knaak, Michael A
Hydrolysis of low concentrations of the acetylthiocholine analogs acetyl(homo)thiocholine and acetyl(nor)thiocholine by acetylcholinesterase may be limited by selective gating at the enzyme peripheral site
Hydrolysis of acetylcholine by acetylcholinesterase (AChE) is extremely rapid, with a second-order hydrolysis rate constant kE (often denoted kcat/KM) that approaches 108 M?1s?1. AChE contains a deep active site gorge with two sites of ligand binding, an acylation site (or A-site) containing the catalytic triad at the base of the gorge and a peripheral site (or P-site) near the gorge entrance. The P-site is known to contribute to catalytic efficiency with acetylthiocholine (AcSCh) by transiently trapping the substrate in a low affinity complex on its way to the A-site, where a short-lived acyl enzyme intermediate is produced. Here we ask whether the P-site does more than simply trap the substrate but in fact selectively gates entry to the A-site to provide specificity for AcSCh (and acetylcholine) relative to the close structural analogs acetylhomothiocholine (Ac-hSCh, which adds one additional methylene group to thiocholine) and acetylnorthiocholine (Ac-nSCh, which deletes one methylene group from thiocholine). We synthesized Ac-hSCh and Ac-nSCh and overcame technical difficulties associated with instability of the northiocholine hydrolysis product. We then compared the catalytic parameters of these substrates with AChE to those of AcSCh. Values of kE for Ac-hSCh and Ac-nSCh were about 2% of that for AcSCh. The kE for AcSCh is close to the theoretical diffusion-controlled limit for the substrate association rate constant, but kE values for Ac-hSCh or Ac-nSCh are too low to be limited by diffusion control. However, analyses of kinetic solvent isotope effects and inhibition patterns for P-site inhibitors indicate that these two analogs also do not equilibrate with the A-site prior to the initial acylation step of catalysis. We propose that kE for these substrates is partially rate-limited by a gating step that involves the movement of bound substrate from the P-site to the A-site.
Beri, Veena; Auletta, Jeffrey T.; Maharvi, Ghulam M.; Wood, Juanita F.; Fauq, Abdul H.; Rosenberry, Terrone L.
A novel neutral xylanase with high SDS resistance from Volvariella volvacea: characterization and its synergistic hydrolysis of wheat bran with acetyl xylan esterase.
A neutral xylanase (XynII) from Volvariella volvacea was identified and characterized. Unlike other modular xylanases, it consists of only a single GH10 catalytic domain with a unique C-terminal sequence (W-R-W-F) and a phenylalanine and proline-rich motif (T-P-F-P-P-F) at N-terminus, indicating that it is a novel GH10 xylanase. XynII exhibited optimal activity at pH 7 and 60 �C and stability over a broad range of pH 4.0-10.0. XynII displayed extreme highly SDS resistance retaining 101.98, 92.99, and 69.84 % activity at the presence of 300 mM SDS on birchwood, soluble oat spelt, and beechwood xylan, respectively. It remained largely intact after 24 h of incubation with proteinase K at a protease to protein ratio of 1:50 at 37 �C. The kinetic constants K(m) value towards beechwood xylan was 0.548 mg ml?�, and the k(cat)/K(m) ratio, reflecting the catalytic efficiency of the enzyme, was 126.42 ml mg?� s?� at 60 �C. XynII was a true endo-acting xylanase lacking cellulase activity. It has weak activity towards xylotriose but efficiently hydrolyzed xylans and xylooligosaccharides larger than xylotriose mainly to xylobiose. Synergistic action with acetyl xylan esterase (AXEI) from V. volvacea was observed for de-starched wheat bran. The highest degree of synergy (DS 1.42) was obtained in sequential reactions with AXEI digestion preceding XynII. The high SDS resistance and intrinsic stability suggested XynII may have potential applications in various industrial processes especially for the detergent and textile industries and animal feed industries. PMID:23903903
Zheng, Fei; Huang, Jingxuan; Yin, Yuhao; Ding, Shaojun
Antagonism of P2Y1-induced vasorelaxation by acyl CoA: a critical role for palmitate and 3?-phosphate
Background and Purpose Acyl derivatives of CoA have been shown to act as antagonists at human platelet and recombinant P2Y1 receptors, but little is known about their effects in the cardiovascular system. This study evaluated the effect of these endogenous nucleotide derivatives at P2Y1 receptors natively expressed in rat and porcine blood vessels. Experimental Approach Isometric tension recordings were used to evaluate the effects of CoA, acetyl CoA, palmitoyl CoA (PaCoA) and 3?-dephospho-palmitoyl-CoA on concentration relaxation�response curves to ADP and uridine triphosphate (UTP). A FlexStation monitored ADP- and UTP-evoked calcium responses in HEK293 cells. Key Results Acetyl CoA and PaCoA, but not CoA, inhibited endothelium-dependent relaxations to ADP with apparent selectivity for P2Y1 receptors (over P2Y2/4 receptors) in rat thoracic aorta; PaCoA was more potent than acetyl CoA (331-fold vs. fivefold shift of ADP response curve evoked by 10 ?M PaCoA and acetyl CoA, respectively); the apparent pA2 value for PaCoA was 6.44. 3?-dephospho-palmitoyl-CoA (10 ?M) was significantly less potent than PaCoA (20-fold shift). In porcine mesenteric arteries, PaCoA and the P2Y1 receptor antagonist MRS2500 blocked ADP-mediated endothelium-dependent relaxations; in contrast, they were ineffective against ADP-mediated endothelium-independent relaxation in porcine coronary arteries (which does not involve P2Y1 receptors). Calcium responses evoked by ADP activation of endogenous P2Y1 receptors in HEK293 cells were inhibited in the presence of PaCoA, which failed to alter responses to UTP (acting at endogenous P2Y2/4 receptors). Conclusions and Implications Acyl derivatives of CoA can act as endogenous selective antagonists of P2Y1 receptors in blood vessels, and this inhibitory effect critically depends on the palmitate and 3?-ribose phosphate substituents on CoA.
Alefishat, E; Alexander, SPH; Ralevic, V
Mapping sugar beet pectin acetylation pattern.
Homogalacturonan-derived partly methylated and/or acetylated oligogalacturonates were recovered after enzymatic hydrolysis (endo-polygalacturonase+pectin methyl esterase+side-chain degrading enzymes) of sugar beet pectin followed by anion-exchange and size exclusion chromatography. Around 90% of the GalA and 75% of the acetyl groups present in the initial sugar beet pectin were recovered as homogalacturonan-derived oligogalacturonates, the remaining GalA and acetyl belonging to rhamnogalacturonic regions. Around 50% of the acetyl groups present in sugar beet homogalacturonans were recovered as partly methylated and/or acetylated oligogalacturonates of degree of polymerisation 5 whose structures were determined by electrospray ionization ion trap mass spectrometry (ESI-IT-MSn). 2-O-acetyl- and 3-O-acetyl-GalA were detected in roughly similar amounts but 2,3-di-O-acetylation was absent. Methyl-esterified GalA residues occurred mainly upstream 2-O-acetyl GalA. Oligogalacturonates containing GalA residues that are at once methyl- and acetyl-esterified were recovered in very limited amounts. A tentative mapping of the distribution of acetyl and methyl esters within sugar beet homogalacturonans is proposed. Unsubstituted GalA residues are likely to be present in limited amounts (approximately 10% of total GalA residues), due to the fact that methyl and acetyl groups are assumed to be most often not carried by the same residues. PMID:16024056
Ralet, Marie-Christine; Cabrera, Juan Carlos; Bonnin, Estelle; Qu�m�ner, Bernard; Hell�n, Pilar; Thibault, Jean-Fran�ois
Differences among Adult COAs and Adult Non-COAs on Levels of Self-Esteem, Depression, and Anxiety.
ERIC Educational Resources Information Center
Examined self-esteem, depression, and anxiety among 60 adult children of alcoholics (COAs) and 143 adult non-COAs. Subjects completed Children of Alcoholics Screening Test, demographic questionnaire, Beck Depression Inventory, State-Trait Anxiety Inventory, and Coopersmith Self-Esteem Inventory. Found no significant differences between COAs and�
Dodd, David T.; Roberts, Richard L.
Acetylation of woody lignocellulose: significance and regulation
Non-cellulosic cell wall polysaccharides constitute approximately one quarter of usable biomass for human exploitation. In contrast to cellulose, these components are usually substituted by O-acetyl groups, which affect their properties and interactions with other polymers, thus affecting their solubility and extractability. However, details of these interactions are still largely obscure. Moreover, polysaccharide hydrolysis to constituent monosaccharides is hampered by the presence of O-acetyl groups, necessitating either enzymatic (esterase) or chemical de-acetylation, increasing the costs and chemical consumption. Reduction of polysaccharide acetyl content in planta is a way to modify lignocellulose toward improved saccharification. In this review we: (1) summarize literature on lignocellulose acetylation in different tree species, (2) present data and current hypotheses concerning the role of O-acetylation in determining woody lignocellulose properties, (3) describe plant proteins involved in lignocellulose O-acetylation, (4) give examples of microbial enzymes capable to de-acetylate lignocellulose, and (5) discuss prospects for exploiting these enzymes in planta to modify xylan acetylation.
Pawar, Prashant Mohan-Anupama; Koutaniemi, Sanna; Tenkanen, Maija; Mellerowicz, Ewa J.
Mechanism of action and biological profile of HMG CoA reductase inhibitors. A new therapeutic alternative.
Lovastatin (MK-803, mevinolin) and simvastatin (MK-733, synvinolin), 2 highly potent 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitors, have been heralded as breakthrough therapy for the treatment of atherosclerotic disease. This paper discusses the biochemical attributes of these HMG CoA reductase inhibitors, their structures and inhibitory properties in a variety of biological systems and presents the rationale for their therapeutic use. Not only do lovastatin and simvastatin potently inhibit cholesterol biosynthesis; they also can result in the induction of hepatic low density lipoprotein (LDL) receptors, thus increasing the catabolism of LDL-cholesterol. Lovastatin and simvastatin are the first HMG CoA reductase inhibitors to receive regulatory agency approval for marketed use. Their safety profiles are reviewed and 2 aspects of this evaluation are stressed. First, the objective in the clinical use of these inhibitors is to normalise plasma cholesterol levels in hypercholesterolaemic individuals. This contrasts with the profound reductions in cholesterol obtained when normocholesterolaemic animals are treated by the high doses of these drugs required for toxicological assessment. Second, both lovastatin and simvastatin are administered as prodrugs in their lactone forms. As lactones, they readily undergo first-pass metabolism, hepatic sequestration and hydrolysis to the active form. Consequently, lovastatin and simvastatin achieve lower plasma drug levels than do other HMG CoA reductase inhibitors in clinical development. Low plasma levels have been established as an important determinant of safety in the use of HMG CoA reductase inhibitors in both animal and human studies. PMID:3076125
Slater, E E; MacDonald, J S
A Nuclear Pyruvate Dehydrogenase Complex Is Important for the Generation of Acetyl-CoA and Histone Acetylation.
DNA transcription, replication, and repair are regulated by histone acetylation, a process that requires the generation of acetyl-coenzyme A (CoA). Here, we show that all the subunits of the mitochondrial pyruvate dehydrogenase complex (PDC) are also present and functional in the nucleus of mammalian cells. We found that knockdown of nuclear PDC in isolated functional nuclei decreased the de novo synthesis of acetyl-CoA and acetylation of core histones. Nuclear PDC levels increased in a cell-cycle-dependent manner and in response to serum, epidermal growth factor, or mitochondrial stress; this was accompanied by a corresponding decrease in mitochondrial PDC levels, suggesting a translocation from the mitochondria to the nucleus. Inhibition of nuclear PDC decreased acetylation of specific lysine residues on histones important for G1-S phase progression and expression of S phase markers. Dynamic translocation of mitochondrial PDC to the nucleus provides a pathway for nuclear acetyl-CoA synthesis required for histone acetylation and epigenetic regulation. PMID:24995980
Sutendra, Gopinath; Kinnaird, Adam; Dromparis, Peter; Paulin, Roxane; Stenson, Trevor H; Haromy, Alois; Hashimoto, Kyoko; Zhang, Nancy; Flaim, Eric; Michelakis, Evangelos D
Decreased Muscle Acetyl-Coenzyme A Carboxylase 2 mRNA and Insulin Resistance in Formerly Obese Subjects
Objective:A relationship between free fatty acids, intramuscular triglycerides (TGMs), and insulin resistance is widely accepted. The intracellular level of malonyl-coenzyme A (CoA) was suggested to be the possible link. Acetyl-CoA carboxylase (ACC) is a key enzyme in fatty acid metabolism, catalyzing the synthesis of malonyl-CoA, a fatty acid acyl-chain elongation unit, from acetyl-CoA. We assessed ACC2 mRNA expression variations in
Giuseppina Rosa; Melania Manco; Natalie Vega; Aldo V. Greco; Marco Castagneto; Hubert Vidal; Geltrude Mingrone
Captan (N-(trichloromethylthio)-4-cyclohexene-1,2-dicarboximide) undergoes hydrolysis readily in water with a maximum half-life of 710 min. Over the pH range 2-6, the reaction is pH independent and the pseudo-first-order rate constant is (1.8 + or - 0.1) x 10 to the -5th power/s....
Inhibition of Neutral Lipase from Castor Bean Lipid Bodies by Coenzyme A (CoA) and Oleoyl-CoA 1
The neutral lipase (EC 22.214.171.124) in lipid body membranes isolated from the endosperm of 4 day old castor (Ricinus communis L.) seedlings catalyzes the hydrolysis of [14C]trioleoylglycerol, releasing [14C]oleic acid for up to 4 hours. However, the addition of Mg-ATP and coenzyme A (CoA), which are present in the cytoplasm of plant cells, caused a progressive inhibition of the neutral lipase such that after 15 minutes, release of [14C]oleic acid was almost undetectable. A fatty acyl CoA synthetase was found in the lipid body membrane which converts [14C]oleic acid produced from the lipase reaction to [14C]oleoyl-CoA under these conditions. The concentration of free oleoyl-CoA in the reaction mixture when the lipase was inhibited by 50% was calculated to be about 21 micromolar. It was found that a mixture of exogenously added oleoyl-CoA and CoA was most effective in causing lipase inhibition. Little inhibition of lipase was detected in the presence of CoA alone. It is possible that this effect is important In vivo in coordinating lipase activity with fatty acid oxidation.
Hills, Matthew J.; Murphy, Denis J.; Beevers, Harry
Process for producing N-acetyl-D-glucosamine
US Patent & Trademark Office Database
This invention pertains to a novel process for directly producing N-acetyl-D-glucosamine from chitin. More particularly, this invention pertains to a novel process for producing N-acetyl-D-glucosamine utilizing an ensemble of the chitinase family of enzymes to hydrolyze chitin of crustacea shells. The invention includes a process for producing N-acetyl-D-glucosamine by enzymatically hydrolyzing chitin with an ensemble of chitinolytic enzymes, including chitinase and chitobiase. In particular, using a two-stage chitin-hydrolysis reactor.
AMP-forming acetylCoA synthetase from the extremely halophilic archaeon Haloarcula marismortui : purification, identification and expression of the encoding gene, and phylogenetic affiliation
Halophilic archaea activate acetate via an (acetate)-inducible AMP-forming acetyl-CoA synthetase (ACS), (Acetate + ATP + CoA ? Acetyl-CoA + AMP + PPi). The enzyme from Haloarcula marismortui was purified to homogeneity. It constitutes a 72-kDa monomer and exhibited a temperature optimum of 41�C and a pH optimum of 7.5. For optimal activity, concentrations between 1�M and 1.5�M�KCl were required, whereas
Christopher Br�sen; Peter Sch�nheit
The effect of acetyl-coenzyme A on phosphate-activated glutaminase from pig kidney and brain
Phosphate-activated glutaminase (EC 220.127.116.11; l-glutamine amidohydrolase) purified from pig kidney and brain is activated by CoA and short-chain acyl-CoA derivatives. Acetyl-CoA is the most powerful activator (KA about 0.2mm). Acetyl-CoA is maximally effective in the absence of other activating anions such as phosphate and citrate, and at low glutamine concentrations. The negative co-operative substrate activation observed at pH7 becomes more pronounced in the presence of acetyl-CoA. Similarly to phosphate, acetyl-CoA produces at high protein concentrations a different type of activation, which is time-dependent, depends on protein concentration and is accompanied by an increase in the sedimentation coefficient. Acetyl-CoA, phosphate and citrate appear to have binding sites in common. No significant difference was observed between kidney and brain phosphate-activated glutaminase.
Kvamme, Elling; Torgner, Ingeborg Aasland
Inherited disorders of 3-methylcrotonyl CoA carboxylation.
The clinical course of 4 patients who had reduced activities of 3-methylcrotonyl CoA carboxylase (also called 3-methylcrotonylglycinuria) is described. Two children presented with a metabolic acidosis, one in the neonatal period and the other with episodes of acidosis that started in the second year of life. In the other 2 children neurological symptoms were prominent, one having infantile spasms and the other developmental regression with a skin rash and alopecia. Three of the children responded well to oral biotin and dietary protein restriction but the fourth, despite a biochemical response to biotin, has a severe neurological handicap. The clinical presentation of inborn errors of 3-methylcrotonyl CoA carboxylase is variable. Metabolic acidosis may not be conspicuous and instead neurological features may predominate.
Leonard, J V; Seakins, J W; Bartlett, K; Hyde, J; Wilson, J; Clayton, B
Pyrazole-based HMG CoA reductase inhibitors
US Patent & Trademark Office Database
Novel compounds and pharmaceutical compositions useful as hypocholesterolemic and hypolipidemic agents are described. More specifically, potent inhibitors of the enzyme 3-hydroxy-3-methylglutaryl-coenzyme A reductase ("HMG CoA reductase") are described. Methods of using such compounds and compositions to treat subjects, including humans, suffering from hyperlipidemia, hypercholesterolemia, hypertriglyceridemia, atherosclerosis, Alzheimer's Disease, benign prostatic hypertrophy (BPH), diabetes and osteoporosis are also described.
Reverse genetic characterization of two paralogous acetoacetyl CoA thiolase genes in Arabidopsis reveals their importance in plant growth and development.
Acetoacetyl CoA thiolase (AACT, EC 22.214.171.124) catalyzes the condensation of two acetyl CoA molecules to form acetoacetyl CoA. Two AACT-encoding genes, At5g47720 (AACT1) and At5g48230 (AACT2), were functionally identified in the Arabidopsis genome by direct enzymological assays and functional expression in yeast. Promoter::GUS fusion experiments indicated that AACT1 is primarily expressed in the vascular system and AACT2 is highly expressed in root tips, young leaves, top stems and anthers. Characterization of T-DNA insertion mutant alleles at each AACT locus established that AACT2 function is required for embryogenesis and for normal male gamete transmission. In contrast, plants lacking AACT1 function are completely viable and show no apparent growth phenotypes, indicating that AACT1 is functionally redundant with respect to AACT2 function. RNAi lines that express reduced levels of AACT2 show pleiotropic phenotypes, including reduced apical dominance, elongated life span and flowering duration, sterility, dwarfing, reduced seed yield and shorter root length. Microscopic analysis reveals that the reduced stature is caused by a reduction in cell size and fewer cells, and male sterility is caused by loss of the pollen coat and premature degeneration of the tapetal cells. Biochemical analyses established that the roots of AACT2 RNAi plants show quantitative and qualitative alterations in phytosterol profiles. These phenotypes and biochemical alterations are reversed when AACT2 RNAi plants are grown in the presence of mevalonate, which is consistent with the role of AACT2 in generating the bulk of the acetoacetyl CoA precursor required for the cytosol-localized, mevalonate-derived isoprenoid biosynthetic pathway. PMID:22332816
Jin, Huanan; Song, Zhihong; Nikolau, Basil J
Synthesis of sepiapterin-C via hydrolysis of 6-ethynylpteridine.
Acid hydrolysis of 6-ethynylpteridine catalyzed by mercury oxide gives 6-acetyl-2-amino-3,4-dihydropteridin-4-one in good yield. Partial reduction of the product with dissolved Al in NH3 solution afforded sepiapterin-C. PMID:24660457
Nxumalo, Winston; Dinsmore, Andrew
Characterization of O-Acetylation of N-Acetylglucosamine
Peptidoglycan (PG) N-acetyl muramic acid (MurNAc) O-acetylation is widely spread in Gram-positive bacteria and is generally associated with resistance against lysozyme and endogenous autolysins. We report here the presence of O-acetylation on N-acetylglucosamine (GlcNAc) in Lactobacillus plantarum PG. This modification of glycan strands was never described in bacteria. Fine structural characterization of acetylated muropeptides released from L. plantarum PG demonstrated that both MurNAc and GlcNAc are O-acetylated in this species. These two PG post-modifications rely on two dedicated O-acetyltransferase encoding genes, named oatA and oatB, respectively. By analyzing the resistance to cell wall hydrolysis of mutant strains, we showed that GlcNAc O-acetylation inhibits N-acetylglucosaminidase Acm2, the major L. plantarum autolysin. In this bacterial species, inactivation of oatA, encoding MurNAc O-acetyltransferase, resulted in marked sensitivity to lysozyme. Moreover, MurNAc over-O-acetylation was shown to activate autolysis through the putative N-acetylmuramoyl-l-alanine amidase LytH enzyme. Our data indicate that in L. plantarum, two different O-acetyltransferases play original and antagonistic roles in the modulation of the activity of endogenous autolysins.
Bernard, Elvis; Rolain, Thomas; Courtin, Pascal; Guillot, Alain; Langella, Philippe; Hols, Pascal; Chapot-Chartier, Marie-Pierre
Replacement of the Saccharomyces cerevisiae acetyl-CoA synthetases by alternative pathways for cytosolic acetyl-CoA synthesis.
Cytosolic acetyl-coenzyme A is a precursor for many biotechnologically relevant compounds produced by Saccharomyces cerevisiae. In this yeast, cytosolic acetyl-CoA synthesis and growth strictly depend on expression of either the Acs1 or Acs2 isoenzyme of acetyl-CoA synthetase (ACS). Since hydrolysis of ATP to AMP and pyrophosphate in the ACS reaction constrains maximum yields of acetyl-CoA-derived products, this study explores replacement of ACS by two ATP-independent pathways for acetyl-CoA synthesis. After evaluating expression of different bacterial genes encoding acetylating acetaldehyde dehydrogenase (A-ALD) and pyruvate-formate lyase (PFL), acs1? acs2? S. cerevisiae strains were constructed in which A-ALD or PFL successfully replaced ACS. In A-ALD-dependent strains, aerobic growth rates of up to 0.27 h(-1) were observed, while anaerobic growth rates of PFL-dependent S. cerevisiae (0.20 h(-1)) were stoichiometrically coupled to formate production. In glucose-limited chemostat cultures, intracellular metabolite analysis did not reveal major differences between A-ALD-dependent and reference strains. However, biomass yields on glucose of A-ALD- and PFL-dependent strains were lower than those of the reference strain. Transcriptome analysis suggested that reduced biomass yields were caused by acetaldehyde and formate in A-ALD- and PFL-dependent strains, respectively. Transcript profiles also indicated that a previously proposed role of Acs2 in histone acetylation is probably linked to cytosolic acetyl-CoA levels rather than to direct involvement of Acs2 in histone acetylation. While demonstrating that yeast ACS can be fully replaced, this study demonstrates that further modifications are needed to achieve optimal in vivo performance of the alternative reactions for supply of cytosolic acetyl-CoA as a product precursor. PMID:24269999
Kozak, Barbara U; van Rossum, Harmen M; Benjamin, Kirsten R; Wu, Liang; Daran, Jean-Marc G; Pronk, Jack T; van Maris, Antonius J A
SIRT3 Deacetylates Mitochondrial 3-Hydroxy-3-Methylglutaryl CoA Synthase 2 and Regulates Ketone Body Production
SUMMARY The mitochondrial sirtuin SIRT3 regulates metabolic homeostasis during fasting and calorie restriction. We identified mitochondrial 3-hydroxy-3-methylglutaryl CoA synthase 2 (HMGCS2) as an acetylated protein and a possible target of SIRT3 in a proteomics survey in hepatic mitochondria from Sirt3?/? (SIRT3KO) mice. HMGCS2 is the rate-limiting step in ?-hydroxybutyrate synthesis and is hyperacetylated at lysines 310, 447, and 473 in the absence of SIRT3. HMGCS2 is deacetylated by SIRT3 in response to fasting in wild-type mice, but not in SIRT3KO mice. HMGCS2 is deacetylated in vitro when incubated with SIRT3 and in vivo by overexpression of SIRT3. Deacetylation of HMGCS2 lysines 310, 447, and 473 by incubation with wild-type SIRT3 or by mutation to arginine enhances its enzymatic activity. Molecular dynamics simulations show that in silico deacetylation of these three lysines causes conformational changes of HMGCS2 near the active site. Mice lacking SIRT3 show decreased ?-hydroxybutyrate levels during fasting. Our findings show SIRT3 regulates ketone body production during fasting and provide molecular insight into how protein acetylation can regulate enzymatic activity.
Shimazu, Tadahiro; Hirschey, Matthew D.; Hua, Lan; Dittenhafer-Reed, Kristin E.; Schwer, Bjoern; Lombard, David B.; Li, Yu; Bunkenborg, Jakob; Alt, Frederick W.; Denu, John M.; Jacobson, Matthew P.; Verdin, Eric
Effect of hydroxypropyl ?-cyclodextrin on physical properties and transition parameters of amylose�lipid complexes of native and acetylated starches
The effect of hydroxpropyl ?-cyclodextrin (HP?-CD) on physical properties and digestibility of wheat, potato, waxy maize and high-amylose maize starches before and after acetylation was studied. Effect of HP?-CD on amylose�lipid complexes in native and acetylated potato starches synthesized using ?-lysophosphatidylcholine was also studied. Acetylation increased swelling factor, amylose leaching, peak viscosity and susceptibility to ?-amylase hydrolysis, but decreased gelatinization
Anil Gunaratne; Harold Corke
The Natural Mentors of Adolescent Children of Alcoholics (COAs): Implications for Preventive Practices.
ERIC Educational Resources Information Center
Late adolescent children of alcoholics (COAs) were interviewed about their relationship with a natural mentor. Results showed that a typical mentor was a same-sex relative who had been responsible for initiating the mentor-like relationship. Differences in the reported adjustment of COAs with and without natural mentors are considered in light of�
Cavell, Timothy A.; Meehan, Barbara T.; Heffer, Robert W.; Holladay, Janice J.
Genetics Home Reference: Succinyl-CoA:3-ketoacid CoA transferase deficiency
... literature OMIM Genetic disorder catalog Conditions > Succinyl-CoA:3-ketoacid CoA transferase deficiency On this page: Description ... definitions Reviewed December 2011 What is succinyl-CoA:3-ketoacid CoA transferase deficiency? Succinyl-CoA:3-ketoacid ...
Characterization and mode of action of two acetyl xylan esterases from Chrysosporium lucknowense C1 active towards acetylated xylans.
Two novel acetyl xylan esterases, Axe2 and Axe3, from Chrysosporium lucknowense (C1), belonging to the carbohydrate esterase families 5 and 1, respectively, were purified and biochemically characterized. Axe2 and Axe3 are able to hydrolyze acetyl groups both from simple acetylated xylo-oligosaccharides and complex non-soluble acetylglucuronoxylan. Both enzymes performed optimally at pH 7.0 and 40 �C. Axe2 has a clear preference for acetylated xylo-oligosaccharides (AcXOS) with a high degree of substitution and Axe3 does not show such preference. Axe3 has a preference for large AcXOS (DP 9-12) when compared to smaller AcXOS (especially DP 4-7) while for Axe2 the size of the oligomer is irrelevant. Even though there is difference in substrate affinity towards acetylated xylooligosaccharides from Eucalyptus wood, the final hydrolysis products are the same for Axe2 and Axe3: xylo-oligosaccharides containing one acetyl group located at the non-reducing xylose residue remain as examined using MALDI-TOF MS, CE-LIF and the application of an endo-xylanase (GH 10). PMID:22112517
Pouvreau, L; Jonathan, M C; Kabel, M A; Hinz, S W A; Gruppen, H; Schols, H A
The diversity of acetylated proteins
Acetylation of proteins, either on various amino-terminal residues or on the ?-amino group of lysine residues, is catalyzed by a wide range of acetyltransferases. Amino-terminal acetylation occurs on the bulk of eukaryotic proteins and on regulatory peptides, whereas lysine acetylation occurs at different positions on a variety of proteins, including histones, transcription factors, nuclear import factors, and ?-tubulin.
Bogdan Polevoda; Fred Sherman
N-Acetylaspartate reductions in brain injury: impact on post-injury neuroenergetics, lipid synthesis, and protein acetylation.
N-Acetylaspartate (NAA) is employed as a non-invasive marker for neuronal health using proton magnetic resonance spectroscopy (MRS). This utility is afforded by the fact that NAA is one of the most concentrated brain metabolites and that it produces the largest peak in MRS scans of the healthy human brain. NAA levels in the brain are reduced proportionately to the degree of tissue damage after traumatic brain injury (TBI) and the reductions parallel the reductions in ATP levels. Because NAA is the most concentrated acetylated metabolite in the brain, we have hypothesized that NAA acts in part as an extensive reservoir of acetate for acetyl coenzyme A synthesis. Therefore, the loss of NAA after TBI impairs acetyl coenzyme A dependent functions including energy derivation, lipid synthesis, and protein acetylation reactions in distinct ways in different cell populations. The enzymes involved in synthesizing and metabolizing NAA are predominantly expressed in neurons and oligodendrocytes, respectively, and therefore some proportion of NAA must be transferred between cell types before the acetate can be liberated, converted to acetyl coenzyme A and utilized. Studies have indicated that glucose metabolism in neurons is reduced, but that acetate metabolism in astrocytes is increased following TBI, possibly reflecting an increased role for non-glucose energy sources in response to injury. NAA can provide additional acetate for intercellular metabolite trafficking to maintain acetyl CoA levels after injury. Here we explore changes in NAA, acetate, and acetyl coenzyme A metabolism in response to brain injury. PMID:24421768
Moffett, John R; Arun, Peethambaran; Ariyannur, Prasanth S; Namboodiri, Aryan M A
N-Acetyl-4-aminophenol (paracetamol), N-acetyl-2-aminophenol and acetanilide in urine samples from the general population, individuals exposed to aniline and paracetamol users.
Epidemiological studies suggest associations between the use of N-acetyl-4-aminophenol (paracetamol) during pregnancy and increased risks of reproductive disorders in the male offspring. Previously we have reported a ubiquitous urinary excretion of N-acetyl-4-aminophenol in the general population. Possible sources are (1) direct intake of paracetamol through medication, (2) paracetamol residues in the food chain and (3) environmental exposure to aniline or related substances that are metabolized into N-acetyl-4-aminophenol. In order to elucidate the origins of the excretion of N-acetyl-4-aminophenol in urine and to contribute to the understanding of paracetamol and aniline metabolism in humans we developed a rapid, turbulent-flow HPLC-MS/MS method with isotope dilution for the simultaneous quantification of N-acetyl-4-aminophenol and two other aniline related metabolites, N-acetyl-2-aminophenol and acetanilide. We applied this method to three sets of urine samples: (1) individuals with no known exposure to aniline and also no recent paracetamol medication; (2) individuals after occupational exposure to aniline but no paracetamol medication and (3) paracetamol users. We confirmed the omnipresent excretion of N-acetyl-4-aminophenol. Additionally we revealed an omnipresent excretion of N-acetyl-2-aminophenol. In contrast, acetanilide was only found after occupational exposure to aniline, not in the general population or after paracetamol use. The results lead to four preliminary conclusions: (1) other sources than aniline seem to be responsible for the major part of urinary N-acetyl-4-aminophenol in the general population; (2) acetanilide is a metabolite of aniline in man and a valuable biomarker for aniline in occupational settings; (3) aniline baseline levels in the general population measured after chemical hydrolysis do not seem to originate from acetanilide and hence not from a direct exposure to aniline itself and (4) N-acetyl-2-aminophenol does not seem to be related to aniline nor to N-acetyl-4-aminophenol in man. PMID:24370547
Dierkes, Georg; Weiss, Tobias; Modick, Hendrik; K�fferlein, Heiko Udo; Br�ning, Thomas; Koch, Holger M
Dehydration and Oxidation of Cellulose Hydrolysis Products in Acidic Solution
The conversion of cellulose in homogenous solution to hydrolysis, dehydration,and oxidation products was investigated. A reaction sequence involving acetylation and degradation by aqueous acids led to humic acid, hydroxymethylfurfural, and levulinic acid as the isolated end-products. The influence of various reaction parameters upon the yields was studied.In the presence of ferric chloride some furfural was formed in addition to the
Klaus Garves
Determination of hepatic 3-hydroxy-3-methylglutaryl CoA reductase activity in man
Summary Procedures were developed for the determination of the activity of the microsomal enzyme 3-hydroxy-3-methylgluta- ryl CoA reductase (HMG CoA reductase, EC 220.127.116.11) in human liver. The enzyme assay could be carried out with as lit- tle as 20 mg of fresh liver tissue, thus making the method appli- cable to specimens obtained by percutaneous liver biopsy. Ex- periments were
G. Nicolau; S. Shefer; G. Salen; E. H. Mosbach
Simultaneous high-performance liquid chromatography determination of coenzyme A, dephospho-coenzyme A, and acetyl-coenzyme A in normal and pantothenic acid-deficient rats.
We describe here a simultaneous high-performance liquid chromatography method for practical and rapid determination of coenzyme A (CoA), dephospho-CoA, and acetyl-CoA in tissues. These coenzymes are biosynthesized from the vitamin pantothenic acid (PaA), which is involved in the metabolism of fatty acids, amino acid catabolism, and several other nutrients. The method employed a Tosoh TSK-GEL ODS-100 V column (250�4.6mm i.d., particle size 5?m) eluted with 100mmol/L NaH(2)PO(4) and 75mmol/L CH(3)COONa (pH was adjusted to 4.6 by the addition of concentrated H(3)PO(4))-acetonitrile (94:6, v/v) at a flow rate of 1.0ml/min. The ultraviolet detector was set at 259nm. The limits of detection for CoA, dephospho-CoA, and acetyl-CoA all were 10pmol. The method was applied to the analysis of several tissues of rats fed normal and PaA-free diets. The results clearly showed that the method was suitable for the simultaneous determination of CoA, dephospho-CoA, and acetyl-CoA in the liver, heart, kidney, spleen, testis, large colon, and muscle, but not for the small intestine, of rats. PMID:22922385
Shibata, Katsumi; Nakai, Takumi; Fukuwatari, Tsutomu
Electrospray tandem mass spectrometry of underivatised acetylated xylo-oligosaccharides.
Acetylated neutral (Xyl(n)Ac(m)) and acidic xylo-oligosaccharides (Xyl(n)Ac(m)MeGlcA, and Xyl(n)Ac(m)MeGlcAHex) obtained by partial acid hydrolysis of Eucalyptus globulus wood glucuronoxylans and fractionated by preparative ligand exchange/size-exclusion chromatography were identified by electrospray ionisation mass spectrometry (ESI-MS). Low molecular weight acetylated xylo-oligosaccharides were studied by ESI-tandem mass spectrometry (MS/MS). All the acetylated xylo-oligosaccharides showed an abundant ion due to the neutral loss of 60 Da (CH(3)CO(2)H) in the MS/MS spectra. The presence of diacetylated xylo-oligosaccharides was confirmed by the ions formed by loss of two molecules of acetic acid. Furthermore, characteristic [Xyl(res)Ac(2)+Na](+) and [XylAc(2)+Na](+) ions, and ions due to loss of XylAc(2), indicate that both acetyl groups are located in the same Xyl residue. On the other hand, losses of Xyl(res)Ac and XylAc are also observed as well as [Xyl(res)Ac+Na](+) and [XylAc+Na](+) , indicating the location of both acetyl groups in different Xyl residues, in some cases even in adjacent xyloses. The MS/MS spectra of triacetylated xylo-oligosaccharides were complex due to the presence of different isobaric xylo-oligosaccharides containing the acetyl groups at different locations in the xylo-oligosaccharide backbone. In the MS/MS spectra of acidic xylo-oligosaccharides, the ion at m/z 387, [Xyl(res)AcMeGlcA+Na](+), indicates that the acetyl groups are preferentially linked to Xyl substituted with MeGlcA. However, acidic xylo-oligosaccharides with the acetyl and 4-O-methylglucuronic acid groups in different Xyl residues were also identified. In neutral and in acidic xylo-oligosaccharides several possible locations of the acetyl groups were identified, namely at terminal positions. In summary, ESI-MS/MS is shown to be a powerful tool for the characterisation of acetylated patterns in complex mixtures of oligosaccharides. PMID:16276485
Reis, Ana; Pinto, Paula; Evtuguin, D V; Neto, Carlos P; Domingues, P; Ferrer-Correia, A J; Domingues, M Ros�rio M
Surface acetylation of bacterial cellulose
Bacterial cellulose was partially acetylated by the fibrous acetylationmethod to modify its physical properties, while preserving the microfibrillarmorphology. The overall degree of substitution was varied from 0.04 to 2.77 bychanging the amount of acetic anhydride added. X-ray diffraction of thepartially acetylated samples showed the crystalline pattern of unmodified celluloseI up to moderate degrees of acetylation, and the change in peak
Dae-Young Kim; Yoshiharu Nishiyama; Shigenori Kuga
Acetylation of endogenous STAT proteins.
Acetylation of signal transducer and activator of transcription (STAT) proteins has been recognized as a significant mechanism for the regulation of their cellular functions. Site-specific antibodies are available only for a minority of STATs. The detection of acetylated STATs by immunoprecipitation (IP) followed by western blot (WB) will be described in the following chapter. Defined conditions for cell lysis and IP will be elucidated on the basis of STAT1 acetylation. PMID:23296729
Ginter, Torsten; Heinzel, Thorsten; Kr�mer, Oliver H
The cytokine-inducible transcription factors signal transducer and activator of transcription 5A and 5B (STAT5A and STAT5B) are important for the proper development of multicellular eukaryotes. Disturbed signaling cascades evoking uncontrolled expression of STAT5 target genes are associated with cancer and immunological failure. Here, we summarize how STAT5 acetylation is integrated into posttranslational modification networks within cells. Moreover, we focus on how inhibitors of deacetylases and tyrosine kinases can correct leukemogenic signaling nodes involving STAT5. Such small molecules can be exploited in the fight against neoplastic diseases and immunological disorders.
Kosan, Christian; Ginter, Torsten; Heinzel, Thorsten; Kramer, Oliver H
Structure of the p300 histone acetyltransferase bound to acetyl-coenzyme A and its analogues.
The p300 and CBP transcriptional coactivator paralogs (p300/CBP) regulate a variety of different cellular pathways, in part, by acetylating histones and more than 70 non-histone protein substrates. Mutation, chromosomal translocation, or other aberrant activities of p300/CBP are linked to many different diseases, including cancer. Because of its pleiotropic biological roles and connection to disease, it is important to understand the mechanism of acetyl transfer by p300/CBP, in part so that inhibitors can be more rationally developed. Toward this goal, a structure of p300 bound to a Lys-CoA bisubstrate HAT inhibitor has been previously elucidated, and the enzyme's catalytic mechanism has been investigated. Nonetheless, many questions underlying p300/CBP structure and mechanism remain. Here, we report a structural characterization of different reaction states in the p300 activity cycle. We present the structures of p300 in complex with an acetyl-CoA substrate, a CoA product, and an acetonyl-CoA inhibitor. A comparison of these structures with the previously reported p300/Lys-CoA complex demonstrates that the conformation of the enzyme active site depends on the interaction of the enzyme with the cofactor, and is not apparently influenced by protein substrate lysine binding. The p300/CoA crystals also contain two poly(ethylene glycol) moieties bound proximal to the cofactor binding site, implicating the path of protein substrate association. The structure of the p300/acetonyl-CoA complex explains the inhibitory and tight binding properties of the acetonyl-CoA toward p300. Together, these studies provide new insights into the molecular basis of acetylation by p300 and have implications for the rational development of new small molecule p300 inhibitors. PMID:24819397
Maksimoska, Jasna; Segura-Pe�a, Dario; Cole, Philip A; Marmorstein, Ronen
Polymorphically acetylated aminoglutethimide in humans.
The urinary excretion during 24 h of aminoglutethimide (AG) its major metabolite (N-acetylAG) and two minor metabolites (N-formylAG and nitroG) were measured in 10 volunteers given AG who had been typed for acetylator phenotype using sulphadimidine. The slow acetylators of sulphadimidine excreted more AG (mean 28% of the administered dose) than did the fast acetylators (12%), but the latter excreted more of the dose as N-acetylAG (8.8%) than did the former (3.9%). NitroG and N-formylAG were minor urinary metabolites of AG in humans. The former was more abundant in the urine of slow acetylators (0.10% of the dose) than in that of fast acetylators (0.047%), whereas the respective proportions of doses excreted as the N-formyl derivative (0.475 and 0.465%) were not significantly different for the two acetylator phenotypes. These results show that AG is among those drugs that are polymorphically acetylated in humans.
Coombes, R. C.; Foster, A. B.; Harland, S. J.; Jarman, M.; Nice, E. C.
Enzymatic hydrolysis of molasses
Kinetic studies of the enzymatic hydrolysis of molasses were conducted using glucoamylase. Central Sugar Refinery SDN BHD contains 13�20% glucose. The molasses was diluted and the kinetic experiments were conducted at 67 �C with 100�1000 mg\\/l of glucoamylase. The glucose contents of the molasses were enhanced after hydrolysis of molasses solution with 1000 mg\\/l glucoamylase. A Lineweaver�Burk plot was obtained
Ghasem D. Najafpour; Cheong Poi Shan
Exome Sequence Reveals Mutations in CoA Synthase as a Cause of Neurodegeneration with Brain Iron Accumulation
Neurodegeneration with brain iron accumulation (NBIA) comprises a clinically and genetically heterogeneous group of disorders with progressive extrapyramidal signs and neurological deterioration, characterized by iron accumulation in the basal ganglia. Exome sequencing revealed the presence of recessive missense mutations in COASY, encoding coenzyme A (CoA) synthase in one NBIA-affected subject. A second unrelated individual carrying mutations in COASY was identified by Sanger sequence analysis. CoA synthase is a bifunctional enzyme catalyzing the final steps of CoA biosynthesis by coupling phosphopantetheine with ATP to form dephospho-CoA and its subsequent phosphorylation to generate CoA. We demonstrate alterations in RNA and protein expression levels of CoA synthase, as well as CoA amount, in fibroblasts derived from the two clinical cases and in yeast. This is the second inborn error of coenzyme A biosynthesis to be implicated in NBIA.
Dusi, Sabrina; Valletta, Lorella; Haack, Tobias B.; Tsuchiya, Yugo; Venco, Paola; Pasqualato, Sebastiano; Goffrini, Paola; Tigano, Marco; Demchenko, Nikita; Wieland, Thomas; Schwarzmayr, Thomas; Strom, Tim M.; Invernizzi, Federica; Garavaglia, Barbara; Gregory, Allison; Sanford, Lynn; Hamada, Jeffrey; Bettencourt, Conceicao; Houlden, Henry; Chiapparini, Luisa; Zorzi, Giovanna; Kurian, Manju A.; Nardocci, Nardo; Prokisch, Holger; Hayflick, Susan; Gout, Ivan; Tiranti, Valeria
Succinyl CoA: 3-oxoacid CoA transferase (SCOT): Human cDNA cloning, human chromosomal mapping to 5p13, and mutation detection in a SCOT-deficient patient
Succinyl CoA: 3-oxoacid CoA transferase (SCOT; E.C.126.96.36.199) mediates the rate-determining step of ketolysis in extrahepatic tissues, the esterification of acetoacetate to CoA for use in energy production. Hereditary SCOT deficiency in humans causes episodes of severe ketoacidosis. We obtained human-heart SCOT cDNA clones spanning the entire 1,560-nt coding sequence. Sequence alignment of the human SCOT peptides with other known CoA
S. Kassovska-Bratinova; M. F. Robert; G. A. Mitchell
Binding of acyl CoA by fatty acid binding protein and the effect on fatty acid activation
The ability of purified rat liver and heart fatty acid binding proteins (FABPs) to bind oleoyl CoA and modulate acyl CoA synthesis by microsomal membranes was investigated. Using binding assays employing either Lipidex 1000 or multilamellar liposomes to sequester unbound ligand, rat liver but not rat heart FABP was shown to bind radiolabeled acyl CoA. Binding studies suggest that liver
R. E. Burrier; C. R. Manson; P. Brecher
Distinct roles of carbohydrate esterase family CE16 acetyl esterases and polymer-acting acetyl xylan esterases in xylan deacetylation.
Mass spectrometric analysis was used to compare the roles of two acetyl esterases (AE, carbohydrate esterase family CE16) and three acetyl xylan esterases (AXE, families CE1 and CE5) in deacetylation of natural substrates, neutral (linear) and 4-O-methyl glucuronic acid (MeGlcA) substituted xylooligosaccharides (XOS). AEs were similarly restricted in their action and apparently removed in most cases only one acetyl group from the non-reducing end of XOS, acting as exo-deacetylases. In contrast, AXEs completely deacetylated longer neutral XOS but had difficulties with the shorter ones. Complete deacetylation of neutral XOS was obtained after the combined action of AEs and AXEs. MeGlcA substituents partially restricted the action of both types of esterases and the remaining acidic XOS were mainly substituted with one MeGlcA and one acetyl group, supposedly on the same xylopyranosyl residue. These resisting structures were degraded to great extent only after inclusion of ?-glucuronidase, which acted with the esterases in a synergistic manner. When used together with xylan backbone degrading endoxylanase and ?-xylosidase, both AE and AXE enhanced the hydrolysis of complex XOS equally. PMID:24140638
Koutaniemi, S; van Gool, M P; Juvonen, M; Jokela, J; Hinz, S W; Schols, H A; Tenkanen, M
Progressing batch hydrolysis process
A progressive batch hydrolysis process for producing sugar from a lignocellulosic feedstock, comprising passing a stream of dilute acid serially through a plurality of percolation hydrolysis reactors charged with said feedstock, at a flow rate, temperature and pressure sufficient to substantially convert all the cellulose component of the feedstock to glucose; cooling said dilute acid stream containing glucose, after exiting the last percolation hydrolysis reactor, then feeding said dilute acid stream serially through a plurality of prehydrolysis percolation reactors, charged with said feedstock, at a flow rate, temperature and pressure sufficient to substantially convert all the hemicellulose component of said feedstock to glucose; and cooling the dilute acid stream containing glucose after it exits the last prehydrolysis reactor.
Wright, John D. (Denver, CO)
Purification and characterization of a cytoplasmic enzyme component of the Na+-activated malonate decarboxylase system of Malonomonas rubra: acetyl-S-acyl carrier protein: malonate acyl carrier protein-SH transferase.
Malonate decarboxylation by crude extracts of Malonomonas rubra was specifically activated by Na+ and less efficiently by Li+ ions. The extracts contained an enzyme catalyzing CoA transfer from malonyl-CoA to acetate, yielding acetyl-CoA and malonate. After about a 26-fold purification of the malonyl-CoA:acetate CoA transferase, an almost pure enzyme was obtained, indicating that about 4% of the cellular protein consisted of the CoA transferase. This abundance of the transferase is in accord with its proposed role as an enzyme component of the malonate decarboxylase system, the key enzyme of energy metabolism in this organism. The apparent molecular weight of the polypeptide was 67,000 as revealed from SDS-polyacrylamide gel electrophoresis. A similar molecular weight was estimated for the native transferase by gel chromatography, indicating that the enzyme exists as a monomer. Kinetic analyses of the CoA transferase yielded the following: pH-optimum at pH 5.5, an apparent Km for malonyl-CoA of 1.9mM, for acetate of 54mM, for acetyl-CoA of 6.9mM, and for malonate of 0.5mM. Malonate or citrate inhibited the enzyme with an apparent Ki of 0.4mM and 3.0mM, respectively. The isolated CoA transferase increased the activity of malonate decarboxylase of a crude enzyme system, in which part of the endogenous CoA transferase was inactivated by borohydride, about three-fold. These results indicate that the CoA transferase functions physiologically as a component of the malonate decarboxylase system, in which it catalyzes the transfer of acyl carrier protein from acetyl acyl carrier protein and malonate to yield malonyl acyl carrier protein and acetate. Malonate is thus activated on the enzyme by exchange for the catalytically important enzymebound acetyl thioester residues noted previously. This type of substrate activation resembles the catalytic mechanism of citrate lyase and citramalate lyase. PMID:18251085
Hilbi, H; Dimroth, P
Stabilization of Cox1p intermediates by the Cox14p-Coa3p complex
Cox14p and Coa3p have been shown to regulate translation of the mitochondrial COX1 mRNA and to be required for assembly of cytochrome oxidase. We present evidence that Cox14p and Coa3p stabilize previously identified Cox1p intermediates and that in the absence of either protein, Cox1p aggregates with itself and other mitochondrial gene products, including cytochrome b, Var1p and Cox2p. Our evidence suggests that Cox1p assembly intermediates are in close proximity to other mitochondrially translated proteins and that an important function of Cox14p and Coa3p is to prevent Cox1 from entering into unproductive aggregation pathways.
McStay, Gavin P.; Su, Chen Hsien; Tzagoloff, Alexander
Histone Acetylation in Drug Addiction
Regulation of chromatin structure through post-translational modifications of histones (e.g. acetylation) has emerged as an important mechanism to translate a variety of environmental stimuli, including drugs of abuse, into specific changes in gene expression. Since alterations in gene expression are thought to contribute to the development and maintenance of the addicted state, recent efforts are aimed at identifying how drugs of abuse alter chromatin structure and the enzymes which regulate it. This review discusses how drugs of abuse alter histone acetylation in brain reward regions, through which enzymes this occurs, and ultimately what role histone acetylation plays in addiction-related behaviors.
Renthal, William; Nestler, Eric J.
Histone H3 tail acetylation modulates ATP-dependent remodeling through multiple mechanisms.
There is a close relationship between histone acetylation and ATP-dependent chromatin remodeling that is not fully understood. We show that acetylation of histone H3 tails affects SWI/SNF (mating type switching/ sucrose non fermenting) and RSC (remodels structure of chromatin) remodeling in several distinct ways. Acetylation of the histone H3 N-terminal tail facilitated recruitment and nucleosome mobilization by the ATP-dependent chromatin remodelers SWI/SNF and RSC. Tetra-acetylated H3, but not tetra-acetylated H4 tails, increased the affinity of RSC and SWI/SNF for nucleosomes while also changing the subunits of SWI/SNF that interact with the H3 tail. The enhanced recruitment of SWI/SNF due to H3 acetylation is bromodomain dependent, but is not further enhanced by additional bromodomains found in RSC. The combined effect of H3 acetylation and transcription activators is greater than either separately which suggests they act in parallel to recruit SWI/SNF. Besides enhancing recruitment, H3 acetylation increased nucleosome mobilization and H2A/H2B displacement by RSC and SWI/SNF in a bromodomain dependent manner and to a lesser extent enhanced ATP hydrolysis independent of bromodomains. H3 and H4 acetylation did not stimulate disassembly of adjacent nucleosomes in short arrays by SWI/SNF or RSC. These data illustrate how histone acetylation modulates RSC and SWI/SNF function, and provide a mechanistic insight into their collaborative efforts to remodel chromatin. PMID:21749977
Chatterjee, Nilanjana; Sinha, Divya; Lemma-Dechassa, Mekonnen; Tan, Song; Shogren-Knaak, Michael A; Bartholomew, Blaine
First principles study of structural, electronic and magnetic properties of Mn2CoAs
NASA Astrophysics Data System (ADS)
We have performed first-principle calculations of the structural, electronic and magnetic properties of Mn2CoAs Heusler alloy, using full-potential linearized augmented plane wave (FP-LAPW) scheme within the GGA. Features such as the lattice constant, the bulk modulus and its pressure derivative are reported. The electronic band structures and density of states of the Mn2CoAs compound show that the spin-up electrons are metallic, but the spin-down bands have a gap of 0.48 eV, resulting in stable half-metallic ferrimagnetic behavior with a magnetic moment of 4.00 ?B.
Berri, Saadi; Ibrir, M.; Maouche, D.; Bensalem, R.
[Acid hydrolysis of mandioca].
The influence of time of hydrolysis, pression of the process, ratio of mass of flour and volume and concentration of the acid solution was studied in the hydrolytic processes for Cassava flour. The aim was to obtain fermentable sugars, and the results were submitted to variance analysis. PMID:1228838
Colombo, A J; Schneiderman, B; Baruffaldi, R; Nacco, R
Role of oxidative enzymatic treatments on enzymatic hydrolysis of softwood.
The impact of oxidative modification and partial removal of lignin by laccase-mediator treatments on the enzymatic hydrolysis of steam-pretreated softwood (SPS) was evaluated. Two mediators, N-hydroxy-N-phenylacetamide (NHA) and its acetylated precursor, were oxidized by the laccase from Trametes hirsuta, and their effects on the activity of cellulolytic enzymes and on the hydrolysis yield of SPS were examined. Both simultaneous and sequential combinations of laccase-mediator treatments with commercial cellulases increased the sugar yield in the enzymatic hydrolysis of SPS. The maximal increase was 21% when a sequential treatment was applied. Laccase treatment alone was also shown to improve hydrolysis. NHA oxidized by laccase inhibited significantly the cellulases of Trichoderma reesei, but the presence of the solid substrate protected the activities against oxidative inactivation. Surface analysis of the lignocellulosic substrate before and after the laccase and cellulase treatments revealed an enrichment of lignin and an increase of carboxylic groups on the surface of the hydrolysis residue. PMID:15129438
Palonen, Hetti; Viikari, Liisa
Determinants within the C-Terminal Domain of Streptomyces lividans Acetyl-CoA Synthetase that Block Acetylation of Its Active Site Lysine In Vitro by the Protein Acetyltransferase (Pat) Enzyme
Reversible lysine acetylation (RLA) is a widespread regulatory mechanism that modulates the function of proteins involved in diverse cellular processes. A strong case has been made for RLA control exerted by homologues of the Salmonella enterica protein acetyltransferase (SePat) enzyme on the broadly distributed AMP-forming CoA ligase (a.k.a. acyl-CoA synthetases) family of metabolic enzymes, with acetyl-CoA synthetase (Acs) being the paradigm in the field. Here we investigate why the Acs homologue in Streptomyces lividans (SlAcs) is poorly acetylated in vitro by the S. lividans protein acetyltransferase (SlPat) enzyme. Chimeras of S. enterica Acs (SeAcs) and S. lividans Acs (SlAcs) constructed during the course of this work were acetylated by SlPatA in vitro, retained most of their activity, and were under RLA control in a heterologous host. We identified SeAcs residues N- and C-terminal to the target lysine that when introduced into SlAcs, rendered the latter under RLA control. These results lend further support to the idea that Pat enzymes interact with extensive surfaces of their substrates. Finally, we suggest that acetylation of SlAcs depends on factors or conditions other than those present in our in vitro system. We also discuss possible explanations why SlAcs is not controlled by RLA as defined in other bacterial species.
Tucker, Alex C.; Escalante-Semerena, Jorge C.
Acetylation of wood causes photobleaching.
This paper deals with the photobleaching of acetylated wood. The acetylated spruce wood was irradiated by artificial sunlight emitted from xenon lamp with covering several kinds of band-pass filter. The lightness (L(*)) of acetylated wood increased with integral irradiance. The chroma (?C(*)) decreased by light-irradiation with wavelength from 430nm to 500nm. However, the light-irradiation including ultraviolet ray region made it decrease after increase with integral irradiance. The visible light made hue angle (h�) increase, however, the ultraviolet ray made it decrease. The lignin degradation and the production of carbonyl groups were observed by light-irradiation including ultraviolet ray. However, no remarkable changes in IR spectra were observed by visible light-irradiation. Photobleaching of acetylated wood was caused by mainly visible light without modifying the IR spectra of lignin. PMID:20696591
Mitsui, Katsuya
Copper supplementation restores cytochrome c oxidase assembly defect in a mitochondrial disease model of COA6 deficiency.
Mitochondrial respiratory chain biogenesis is orchestrated by hundreds of assembly factors, many of which are yet to be discovered. Using an integrative approach based on clues from evolutionary history, protein localization and human genetics, we have identified a conserved mitochondrial protein, C1orf31/COA6, and shown its requirement for respiratory complex IV biogenesis in yeast, zebrafish and human cells. A recent next-generation sequencing study reported potential pathogenic mutations within the evolutionarily conserved Cx9CxnCx10C motif of COA6, implicating it in mitochondrial disease biology. Using yeast coa6? cells, we show that conserved residues in the motif, including the residue mutated in a patient with mitochondrial disease, are essential for COA6 function, thus confirming the pathogenicity of the patient mutation. Furthermore, we show that zebrafish embryos with zfcoa6 knockdown display reduced heart rate and cardiac developmental defects, recapitulating the observed pathology in the human mitochondrial disease patient who died of neonatal hypertrophic cardiomyopathy. The specific requirement of Coa6 for respiratory complex IV biogenesis, its intramitochondrial localization and the presence of the Cx9CxnCx10C motif suggested a role in mitochondrial copper metabolism. In support of this, we show that exogenous copper supplementation completely rescues respiratory and complex IV assembly defects in yeast coa6? cells. Taken together, our results establish an evolutionarily conserved role of Coa6 in complex IV assembly and support a causal role of the COA6 mutation in the human mitochondrial disease patient. PMID:24549041
Ghosh, Alok; Trivedi, Prachi P; Timbalia, Shrishiv A; Griffin, Aaron T; Rahn, Jennifer J; Chan, Sherine S L; Gohil, Vishal M
Molecular Structure of Acetyl Peroxide
NSDL National Science Digital Library
Acetyl peroxide is a colorless liquid with a pungent odor. It is generally stored as a 25% solution in dimethyl phthalate to prevent detonation. It may explode if heated, or in contact with combustible materials. As the pure material, acetyl peroxide is unstable and incompatible with organic materials. The compound is harmful by inhalation, ingestion and skin contact. It is used as an initiator and catalyst for resins, and it also promotes polymerization in the manufacture of certain plastics.
Coenzyme A esters of 2-aryloxyphenoxypropionate herbicides and 2-arylpropionate antiinflammatory drugs are potent and stereoselective inhibitors of rat liver acetyl-CoA carboxylase.
The CoA esters of diclofop, haloxyfop and fluazifop are up to 425-fold more potent than the corresponding unconjugated herbicides as inhibitors of rat liver acetyl-CoA carboxylase (EC 188.8.131.52); the most potent inhibitor is (R)-fluazifopyl-CoA2 (Ki = 0.03 microM). The binding site is stereoselective for (R)-diclofop, the herbicidally active enantiomer, and for (R)-diclofopyl-CoA. The CoA esters of the antiinflammatory drugs ibuprofen and fenoprofen also strongly inhibit this carboxylase. (S)-Ibuprofenyl-CoA (Ki = 0.7 microM), the CoA ester of the enantiomer with antiinflammatory activity, is 15-fold more potent as an inhibitor than (R)-ibuprofenyl-CoA. These results suggest that some of the biological effects of these herbicides and antiinflammatory drugs in animals may be due to the inhibition of acetyl-CoA carboxylase by their acyl-CoA derivatives. PMID:1347398
Kemal, C; Casida, J E
Isomerization of 1-O-indol-3-ylacetyl-beta-D-glucose. Enzymatic hydrolysis of 1-O, 4-O, and 6-O-indol-3-ylacetyl-beta-D-glucose and the enzymatic synthesis of indole-3-acetyl glycerol by a hormone metabolizing complex
NASA Technical Reports Server (NTRS)
The first compound in the series of reactions leading to the ester conjugates of indole-3-acetic acid (IAA) in kernels of Zea mays sweet corn is the acyl alkyl acetal, 1-O-indol-3-ylacetyl-beta-D-glucose (1-O-IAGlu). The enzyme catalyzing the synthesis of this compound is UDP-glucose:indol-3-ylacetate glucosyl-transferase (IAGlu synthase). The IAA moiety of the high energy compound 1-O-IAGlu may be enzymatically transferred to myo-inositol or to glycerol or the 1-O-IAGlu may be enzymatically hydrolyzed. Alternatively, nonenzymatic acyl migration may occur to yield the 2-O, 4-O, and 6-O esters of IAA and glucose. The 4-O and 6-O esters may then be enzymatically hydrolyzed to yield free IAA and glucose. This work reports new enzymatic activities, the transfer of IAA from 1-O-IAGlu to glycerol, and the enzyme-catalyzed hydrolysis of 4-O and 6-O-IAGlu. Data is also presented on the rate of non-enzymatic acyl migration of IAA from the 1-O to the 4-O and 6-O positions of glucose. We also report that enzymes catalyzing the synthesis of 1-O-IAGlu and the hydrolysis of 1-O, 4-O, and 6-O-IAGlu fractionate as a hormone metabolizing complex. The association of synthetic and hydrolytic capabilities in enzymes which cofractionate may have physiological significance.
Kowalczyk, S.; Bandurski, R. S.
Binding of acyl CoA by fatty acid binding protein and the effect on fatty acid activation
The ability of purified rat liver and heart fatty acid binding proteins (FABPs) to bind oleoyl CoA and modulate acyl CoA synthesis by microsomal membranes was investigated. Using binding assays employing either Lipidex 1000 or multilamellar liposomes to sequester unbound ligand, rat liver but not rat heart FABP was shown to bind radiolabeled acyl CoA. Binding studies suggest that liver FABP has a single binding site for acyl CoA which is separate from the two binding sites for fatty acids. Experiments were then performed to determine how binding may influence acyl CoA metabolism by liver microsomes or heart sarcoplasmic reticulum. Using liposomes as fatty acid donors, liver FABP stimulated acyl CoA production whereas heart FABP did not stimulate production over control values. /sup 14/C-Fatty acid-FABP complexes were prepared, incubated with membranes and acyl CoA synthetase activity was determined. Up to 70% of the fatty acid could be converted to acyl CoA in the presence of liver FABP but in the presence of heart FABP, only 45% of the fatty acid was converted. The amount of product formed was not changed by additional membrane, enzyme cofactor, or incubation time. Liver but not heart FABP bound the acyl CoA formed and removed it from the membranes. These studies suggest that liver FABP can increase the amount of acyl CoA by binding this ligand thereby removing it from the membrane and possibly aiding transport within the cell.
Burrier, R.E.; Manson, C.R.; Brecher, P.
Steady-state concentrations of coenzyme A, acetyl-coenzyme A and long-chain fatty acyl-coenzyme A in rat-liver mitochondria oxidizing palmitate
1. Fluorimetric assays are described for CoASH, acetyl-CoA and long-chain fatty acyl-CoA, and are sensitive to at least 50??moles of each. 2. Application of these assays to rat-liver mitochondria oxidizing palmitate in the absence and presence of carnitine indicated two pools of intramitochondrial CoA. One pool could be acylated by palmitate and ATP, and the other pool acylated by palmitate with ATP and carnitine, or by palmitoylcarnitine alone. 3. The intramitochondrial content of acetyl-CoA is increased by the oxidation of palmitate both in the absence and presence of l-malate. 4. The conversion of palmitoyl-CoA into acetyl-CoA by ?-oxidation takes place without detectable accumulation of acyl-CoA intermediates.
Garland, P. B.; Shepherd, D.; Yates, D. W.
Fatty infiltration in the liver in medium chain acyl CoA dehydrogenase deficiency.
Fatty infiltration of the liver at postmortem examination has been recommended as a criterion for selection of infants who have died suddenly and unexpectedly for further biochemical investigation for disorders of fatty acid oxidation. We describe a boy with medium chain acyl CoA dehydrogenase deficiency who died four months after diagnosis and in whom only minimal hepatic fatty infiltration was found.
Losty, H C; Lee, P; Alfaham, M; Gray, O P; Leonard, J V
Acetyltransferase and human hemoglobin acetylation
A minor component of human fetal hemoglobin (Hb F) is acetylated at the amino-terminus of the ..gamma..-globin chains. A similar minor component of Hb F is formed during translation of cord blood mRNA in the rabbit reticulocyte lysate system. The acetylation appeared to be enzymatic. This system contains an acetyltransferase capable of acetylating histones and hemoglobins. The enzyme, partially purified by histone-Sepharose affinity chromatography was capable of incorporating labeled acetyl- group from 1-(/sup 14/C-acetyl)-CoA into both human Hb F/sub 0/ and HB A/sub 0/, but at a lower rate than for histones. Characterization of the labeled products indicated that the ..cap alpha..-chains of both hemoglobins were being acetylated presumably at a lysyl-residue, but in the case of Hb F/sub 0/ the amino-terminus of the ..gamma..-globin chains was acetylated as well. While histone-Sepharose bound more than 95% of the enzyme, Sepharose linked Hb F/sub 0/, ..gamma..-globin chains, and Hb Bart's bound 14, 5, and 12% of the activity, respectively. Enzyme bound to these resins was not any more active on the hemoglobins than was the enzyme bound to the histone-Sepharose. The histone-Sepharose was also used to detect the enzyme in human cord blood red cells separated by dextran 40 density gradient centrifugation. Activity was found mostly in the young cells, and was directly related to the number of reticulocytes present in any one fraction.
Kasten-Jolly, J.; Qiu, C.C.; Abraham, E.C.
Progressing Batch Hydrolysis Reactor
In all dilute acid hydrolysis processes for glucose production, conditions severe enough to hydrolyze crystalline cellulose to glucose are also severe enough to degrade the glucose into undesirable compounds such as hydroxy-methylfurfural (HMF), levulinic acid, and formic acid. One way to minimize the sugar degradation is to remove the sugars from the reaction zone before substantial degradation occurs. Sugars are most efficiently removed by a reactor system that uses countercurrent flow of liquids and solids, which allows simultaneous achievement of high yields and high sugar concentrations. The progressing batch hydrolysis process, invented and now under development at SERI, uses several percolation reactors in series to simulate countercurrent flow of liquids and solids. In this way, the advantages of countercurrent flow are achieved, and the mechanical and operational simplicity of the percolation reactor is retained. This paper describes the theory and operation of the progressing batch hydrolysis reactor and presents the results of our mathematical modeling of the system. 25 refs., 7 figs.
Wright, J.D.; Bergeron, P.W.; Werdene, P.J.
Metabolic biology of 3-methylglutaconic acid-uria: a new perspective.
Over the past 25�years a growing number of distinct syndromes/mutations associated with compromised mitochondrial function have been identified that share a common feature: urinary excretion of 3-methylglutaconic acid (3MGA). In the leucine degradation pathway, carboxylation of 3-methylcrotonyl CoA leads to formation of 3-methylglutaconyl CoA while 3-methylglutaconyl CoA hydratase converts this metabolite to 3-hydroxy-3-methylglutaryl CoA (HMG CoA). In "primary" 3MGA-uria, mutations in the hydratase are directly responsible for the accumulation of 3MGA. On the other hand, in all "secondary" 3MGA-urias, no defect in leucine catabolism exists and the metabolic origin of 3MGA is unknown. Herein, a path to 3MGA from mitochondrial acetyl CoA is proposed. The pathway is initiated when syndrome-associated mutations/DNA deletions result in decreased Krebs cycle flux. When this occurs, acetoacetyl CoA thiolase condenses two acetyl CoA into acetoacetyl CoA plus CoASH. Subsequently, HMG CoA synthase 2 converts acetoacetyl CoA and acetyl CoA to HMG CoA. Under syndrome-specific metabolic conditions, 3-methylglutaconyl CoA hydratase converts HMG CoA into 3-methylglutaconyl CoA in a reverse reaction of the leucine degradation pathway. This metabolite fails to proceed further up the leucine degradation pathway owing to the kinetic properties of 3-methylcrotonyl CoA carboxylase. Instead, hydrolysis of the CoA moiety of 3-methylglutaconyl CoA generates 3MGA, which appears in urine. If experimentally confirmed, this pathway provides an explanation for the occurrence of 3MGA in multiple disorders associated with compromised mitochondrial function. PMID:24407466
Su, Betty; Ryan, Robert O
De novo CoA biosynthesis is required to maintain DNA integrity during development of the Drosophila nervous system.
In a forward genetic screen in Drosophila melanogaster, aimed to identify genes required for normal locomotor function, we isolated dPPCS (the second enzyme of the Coenzyme A biosynthesis pathway). The entire Drosophila CoA synthesis route was dissected, annotated and additional CoA mutants were obtained (dPANK/fumble) or generated (dPPAT-DPCK). Drosophila CoA mutants suffer from neurodegeneration, altered lipid homeostasis and the larval brains display increased apoptosis. Also, de novo CoA biosynthesis is required to maintain DNA integrity during the development of the central nervous system. In humans, mutations in the PANK2 gene, the first enzyme in the CoA synthesis route, are associated with pantothenate kinase-associated neurodegeneration. Currently, the pathogenesis of this neurodegenerative disease is poorly understood. We provide the first comprehensive analysis of the physiological implications of mutations in the entire CoA biosynthesis route in an animal model system. Surprisingly, our findings reveal a major role of this conserved pathway in maintaining DNA and cellular integrity, explaining how impaired CoA synthesis during CNS development can elicit a neurodegenerative phenotype. PMID:18407920
Bosveld, Floris; Rana, Anil; van der Wouden, Petra E; Lemstra, Willy; Ritsema, Martha; Kampinga, Harm H; Sibon, Ody C M
Crystallographic trapping of the glutamyl-CoA thioester intermediate of family I CoA transferases
Coenzyme A transferases are involved in a broad range of biochemical processes in both prokaryotes and eukaryotes, and exhibit a diverse range of substrate specificities. The YdiF protein from Escherichia coli O157:H7 is an acyl-CoA transferase of unknown physiological function, and belongs to a large sequence family of CoA transferases, present in bacteria to humans, which utilize oxoacids as acceptors. In vitro measurements showed that YdiF displays enzymatic activity with short-chain acyl-CoAs. The crystal structures of YdiF and its complex with CoA, the first co-crystal structure for any Family I CoA transferase, have been determined and refined at 1.9 and 2.0 Angstrom resolution, respectively. YdiF is organized into tetramers, with each monomer having an open {alpha}/{beta} structure characteristic of Family I CoA transferases. Co-crystallization of YdiF with a variety of CoA thioesters in the absence of acceptor carboxylic acid resulted in trapping a covalent {gamma}-glutamyl-CoA thioester intermediate. The CoA binds within a well defined pocket at the N- and C-terminal domain interface, but makes contact only with the C-terminal domain. The structure of the YdiF complex provides a basis for understanding the different catalytic steps in the reaction of Family I CoA transferases.
Rangarajan,E.; Li, Y.; Ajamian, E.; Iannuzzi, P.; Kernaghan, S.; Fraser, M.; Cygler, M.; Matte, A.
The emission of corrosive vapours by wood. Sweet-chestnut (Castanea stiva) and wych-elm (Ulmusglabrau) O-acetyl-4-O-methylglucuronoxylans extracted with dimethyl sulphoxide.
1. O-Acetylated 4-O-methylglucuronoxylans were isolated from sweet chestnut and wych elm, either green or incubated at 48 degrees and 100% relative humidity for 36 weeks. 2. The chlorine-ethanolamine method of delignification resulted in a 50% loss of O-acetyl groups from green wych elm compared with an 18% loss from green sweet chestnut. 3. The acid-chlorite method gave an acceptable loss of O-acetyl groups in three cases, but incubated sweet chestnut showed a 44.6% loss. However, it is believed that this is due to the loss of simple O-acetylated xylose sugars resulting from glycosidic hydrolysis, rather than removal of O-acetyl groups by direct hydrolysis. Assuming that this occurs in a random manner, it is unlikely to have much structural significance. 4. Dimethyl sulphoxide extraction of chestnut holocellulose and elm holocellulose, green and incubated, yielded O-acetyl glucuronoxylans containing 10.2, 3.8, 13.1 and 7.7% O-acetyl groups respectively. 5. The location of these O-acetyl groups was determined by Bouveng's method in which phenyl isocyanate is used as a blocking group. PMID:5808312
Cochrane, G C; Gray, J D; Arni, P C
Hydrolytic stability of water-soluble spruce O-acetyl galactoglucomannans
Water-soluble native O-acetyl galactoglucomannan (GGM) from spruce is a polysaccharide that can be produced in an industrial scale. To develop GGM applications, information is needed on its stability, particularly under acidic conditions. Therefore, acid hydrolysis of spruce GGM was investigated at various pH levels and temperatures. The results allow an estimation of the stability of GGM under food processing conditions
Chunlin Xu; Andrey Pranovich; J. Hemmimg; Bjarne Holmbom; Simone Albrecht; Henk A. Schols; S. Willfor
Acetylation at O-2 of arabinofuranose residues in feruloylated arabinoxylan from bamboo shoot cell-walls.
Hydrolysis of bamboo shoot cell-walls with Driselase (a fungal enzyme preparation) gave an arabinoxylan trisaccharide with ferulic and acetic acids as ester groups. The structure of this oligosaccharide was determined to be O-[2-O-acetyl-5-O-[E)-feruloyl)-alpha-L-arabinofuranosyl]-(1----3) -O-beta- D-xylopyranosyl-(1----4)-D-xylopyranose, on the basis of spectroscopy and methylation analysis. PMID:1367649
Ishii, T
Alkaline thermal sludge hydrolysis.
The waste activated sludge (WAS) treatment of wastewater produces excess sludge which needs further treatment prior to disposal or incineration. A reduction in the amount of excess sludge produced, and the increased dewaterability of the sludge are, therefore, subject of renewed attention and research. A lot of research covers the nature of the sludge solids and associated water. An improved dewaterability requires the disruption of the sludge cell structure. Previous investigations are reviewed in the paper. Thermal hydrolysis is recognized as having the best potential to meet the objectives and acid thermal hydrolysis is most frequently used, despite its serious drawbacks (corrosion, required post-neutralization, solubilization of heavy metals and phosphates, etc.). Alkaline thermal hydrolysis has been studied to a lesser extent, and is the subject of the detailed laboratory-scale research reported in this paper. After assessing the effect of monovalent/divalent cations (respectively, K(+)/Na(+) and Ca(2+)/Mg(2+)) on the sludge dewaterability, only the use of Ca(2+) appears to offer the best solution. The lesser effects of K(+), Na(+) and Mg(2+) confirm previous experimental findings. As a result of the experimental investigations, it can be concluded that alkaline thermal hydrolysis using Ca(OH)(2) is efficient in reducing the residual sludge amounts and in improving the dewaterability. The objectives are fully met at a temperature of 100 degrees C; at a pH approximately 10 and for a 60-min reaction time, where all pathogens are moreover killed. Under these optimum conditions, the rate of mechanical dewatering increases (the capillary suction time (CST) value is decreased from approximately 34s for the initial untreated sample to approximately 22s for the hydrolyzed sludge sample) and the amount of DS to be dewatered is reduced to approximately 60% of the initial untreated amount. The DS-content of the dewatered cake will be increased from 28 (untreated) to 46%.Finally, the mass and energy balances of a wastewater treatment plant with/without advanced sludge treatment (AST) are compared. The data clearly illustrate the benefits of using an alkaline AST-step in the system. PMID:12573845
Neyens, E; Baeyens, J; Creemers, C
Thermostable Enzymes in Lignocellulose Hydrolysis
Thermostable enzymes offer potential benefits in the hydrolysis of lignocellulosic substrates; higher\\u000a specific activity decreasing the amount of enzymes, enhanced stability allowing improved hydrolysis performance\\u000a and increased flexibility with respect to process configurations, all leading to improvement of the overall\\u000a economy of the process. New thermostable cellulase mixtures were composed of cloned fungal enzymes for\\u000a hydrolysis experiments. Three thermostable cellulases,
Liisa Viikari; Marika Alapuranen; Terhi Puranen; Jari Vehmaanper�; Matti Siika-aho
Acetyl esterase production by Termitomyces clypeatus
Production of acetyl esterase by Termitomyces clypeatus was stimulated by xylan, cellulose, arabinose and arabinose-containing polysaccharides in the growth medium. The culture filtrate was equally active with p-nitrophenyl acetate and acetyl xylan. Acetyl xylan was completely deacetylated by the enzyme. Activity was optimum at pH 6.5 and at 50�C. The Km values for p-nitrophenyl acetate and acetyl xylan were 0.83
A. Mukhopadhyay; P. P. Hazra; T. Sengupta; A. K. Ghosh; S. Sengupta
Peripheral Effects of FAAH Deficiency on Fuel and Energy Homeostasis: Role of Dysregulated Lysine Acetylation
Background FAAH (fatty acid amide hydrolase), primarily expressed in the liver, hydrolyzes the endocannabinoids fatty acid ethanolamides (FAA). Human FAAH gene mutations are associated with increased body weight and obesity. In our present study, using targeted metabolite and lipid profiling, and new global acetylome profiling methodologies, we examined the role of the liver on fuel and energy homeostasis in whole body FAAH?/? mice. Methodology/Principal Findings FAAH?/? mice exhibit altered energy homeostasis demonstrated by decreased oxygen consumption (Indirect calorimetry). FAAH?/? mice are hyperinsulinemic and have adipose, skeletal and hepatic insulin resistance as indicated by stable isotope phenotyping (SIPHEN). Fed state skeletal muscle and liver triglyceride levels was increased 2�3 fold, while glycogen was decreased 42% and 57% respectively. Hepatic cholesterol synthesis was decreased 22% in FAAH?/? mice. Dysregulated hepatic FAAH?/? lysine acetylation was consistent with their metabolite profiling. Fasted to fed increases in hepatic FAAH?/? acetyl-CoA (85%, p<0.01) corresponded to similar increases in citrate levels (45%). Altered FAAH?/? mitochondrial malate dehydrogenase (MDH2) acetylation, which can affect the malate aspartate shuttle, was consistent with our observation of a 25% decrease in fed malate and aspartate levels. Decreased fasted but not fed dihydroxyacetone-P and glycerol-3-P levels in FAAH?/? mice was consistent with a compensating contribution from decreased acetylation of fed FAAH?/? aldolase B. Fed FAAH?/? alcohol dehydrogenase (ADH) acetylation was also decreased. Conclusions/Significance Whole body FAAH deletion contributes to a pre-diabetic phenotype by mechanisms resulting in impairment of hepatic glucose and lipid metabolism. FAAH?/? mice had altered hepatic lysine acetylation, the pattern sharing similarities with acetylation changes reported with chronic alcohol treatment. Dysregulated hepatic lysine acetylation seen with impaired FAA hydrolysis could support the liver's role in fostering the pre-diabetic state, and may reflect part of the mechanism underlying the hepatic effects of endocannabinoids in alcoholic liver disease mouse models.
Vaitheesvaran, Bhavapriya; Yang, Li; Hartil, Kirsten; Glaser, Sherrye; Yazulla, Stephen; Bruce, James E.; Kurland, Irwin J.
Effects of leptin, troglitazone, and dietary fat on stearoyl CoA desaturase
Leptin, troglitazone, and high fat feeding profoundly influence the lipid content of various tissues. To determine if they affect the expression of stearoyl CoA desaturase (SCD)-1 and -2, their mRNA was measured in livers of normal, hyperleptinemic, troglitazone-treated, and fat-fed rats. Hyperleptinemia, which reduces tissue TG by downregulating lipogenic enzymes and upregulating fatty acid oxidation, lowered SCD-1 96% below controls
Tetsuya Kakuma; Young Lee; Roger H Unger
coa types and antimicrobial resistance profile of Staphylococcus aureus isolates from cases of bovine mastitis
The objective of the study was to determine the interrelationship between coa types of Staphylococcus aureus isolates from bovine mastitis cases and their susceptibility patterns to commonly used antimicrobial agents. In this study,\\u000a a total of 63 S. aureus isolates obtained from 387 clinical and subclinical bovine mastitis milk specimens were examined for antimicrobial susceptibility\\u000a against various classes of antibiotics
Habib Dastmalchi Saei
Upregulation of Endothelial Nitric Oxide Synthase by HMG CoA Reductase Inhibitors
Background�Oxidized low-density lipoprotein (ox-LDL) causes endothelial dysfunction in part by decreasing the availability of endothelial nitric oxide (NO). Although HMG CoA reductase inhibitors restore endothelial function by reducing serum cholesterol levels, it is not known whether they can also directly upregulate endothelial NO synthase (ecNOS) activity. Methods and Results�Human saphenous vein endothelial cells were treated with ox-LDL (50 mg\\/mL thiobarbituric
Ulrich Laufs; Vito La Fata; Jorge Plutzky; James K. Liao
A Route Optimization Via Recursive CoA Substitution for Nested Mobile Networks
\\u000a In nested mobile networks, the undesirable effects due to nonoptimal routing tends to get aggravated, leading to excessively\\u000a long packet sizes and transfer delays. In this paper, in order to resolve the non-optimal routing problem, also known as �pinball\\u000a routing problem� in the literature, we propose a new route optimization scheme where the care-of address (CoA) in each binding\\u000a update
Young Beom Kim; Kang-yoon Lee; Hyunchul Ku; Eui-nam Huh
Structural and Functional Similarities between Mitochondrial Malate Dehydrogenase and L-3-Hydroxyacyl CoA Dehydrogenase
Pig heart mitochondrial malate dehydrogenase (EC 188.8.131.52), which has been obtained free of electrophoretic subforms, has been shown to have a molecular weight of 67,000 and to be composed of two polypeptide chains. Comparison of these and other properties, such as amino-acid composition, isoelectric point, and keto-substrate inhibition, with those of L-3-hydroxyacyl CoA dehydrogenase (EC 22.214.171.124), another NAD+-dependent dehydrogenase of
Barbara E. Noyes; Beat E. Glatthaar; John S. Garavelli; Ralph A. Bradshaw
Identifying Related Documents For Research Paper Recommender By CPA and COA
This work-in-progress paper introduces two new approaches called Citation Proximity Analysis (CPA) and Citation Order Analysis (COA). They can be applied to identify related documents for the purpose of research paper recommender systems. CPA is a variant of co-citation analysis that additionally considers the proximity of citations to each other within an article's full-text. The underlying idea is that the
Bela Gipp; J�ran Beel
Kinetic studies on two isoforms of acetyl-CoA carboxylase from maize leaves.
The steady-state kinetics of two multifunctional isoforms of acetyl-CoA carboxylase (ACCase) from maize leaves (a major isoform, ACCase1 and a minor isoform, ACCase2) have been investigated with respect to reaction mechanism, inhibition by two graminicides of the aryloxyphenoxypropionate class (quizalofop and fluazifop) and some cellular metabolites. Substrate interaction and product inhibition patterns indicated that ADP and P(i) products from the first partial reaction were not released before acetyl-CoA bound to the enzymes. Product inhibition patterns did not match exactly those predicted for an ordered Ter Ter or a random Ter Ter mechanism, but were close to those postulated for an ordered mechanism. ACCase2 was about 1/2000 as sensitive as ACCase1 to quizalofop but only about 1/150 as sensitive to fluazifop. Fitting inhibition data to the Hill equation indicated that binding of quizalofop or fluazifop to ACCase1 was non-cooperative, as shown by the Hill constant (n(app)) values of 0.86 and 1.16 for quizalofop and fluazifop respectively. Apparent inhibition constant values (K' from the Hill equation) for ACCase1 were 0.054 microM for quizalofop and 21.8 microM for fluazifop. On the other hand, binding of quizalofop or fluazifop to ACCase2 exhibited positive co-operativity, as shown by the (napp) values of 1.85 and 1.59 for quizalofop and fluazifop respectively. K' values for ACCase2 were 1.7 mM for quizalofop and 140 mM for fluazifop. Kinetic parameters for the co-operative binding of quizalofop to maize ACCase2 were close to those of another multifunctional ACCase of limited sensitivity to graminicide, ACC220 from pea. Inhibition of ACCase1 by quizalofop was mixed-type with respect to acetyl-CoA or ATP, but the concentration of acetyl-CoA had the greater effect on the level of inhibition. Neither ACCase1 nor ACCase2 was appreciably sensitive to CoA esters of palmitic acid (16:0) or oleic acid (18:1). Approximate IC50 values were 10 microM (ACCase2) and 50 microM (ACCase1) for both CoA esters. Citrate concentrations up to 1 mM had no effect on ACCase1 activity. Above this concentration, citrate was inhibitory. ACCase2 activity was slightly stimulated by citrate over a broad concentration range (0.25-10 mM). The significance of possible effects of acyl-CoAs or citrate in vivo is discussed.
Herbert, D; Price, L J; Alban, C; Dehaye, L; Job, D; Cole, D J; Pallett, K E; Harwood, J L
Germline Deletion of Pantothenate Kinases 1 and 2 Reveals the Key Roles for CoA in Postnatal Metabolism
Pantothenate kinase (PanK) phosphorylates pantothenic acid (vitamin B5) and controls the overall rate of coenzyme A (CoA) biosynthesis. Pank1 gene deletion in mice results in a metabolic phenotype where fatty acid oxidation and gluconeogenesis are impaired in the fasted state, leading to mild hypoglycemia. Inactivating mutations in the human PANK2 gene lead to childhood neurodegeneration, but Pank2 gene inactivation in mice does not elicit a phenotype indicative of the neuromuscular symptoms or brain iron accumulation that accompany the human disease. Pank1/Pank2 double knockout (dKO) mice were derived to determine if the mild phenotypes of the single knockout mice are due to the ability of the two isoforms to compensate for each other in CoA biosynthesis. Postnatal development was severely affected in the dKO mice. The dKO pups developed progressively severe hypoglycemia and hyperketonemia by postnatal day 10 leading to death by day 17. Hyperketonemia arose from impaired whole-body ketone utilization illustrating the requirement for CoA in energy generation from ketones. dKO pups had reduced CoA and decreased fatty acid oxidation coupled with triglyceride accumulation in liver. dKO hepatocytes could not maintain the NADH levels compared to wild-type hepatocytes. These results revealed an important link between CoA and NADH levels, which was reflected by deficiencies in hepatic oleate synthesis and gluconeogenesis. The data indicate that PanK1 and PanK2 can compensate for each other to supply tissue CoA, but PanK1 is more important to CoA levels in liver whereas PanK2 contributes more to CoA synthesis in the brain.
Garcia, Matthew; Leonardi, Roberta; Zhang, Yong-Mei; Rehg, Jerold E.; Jackowski, Suzanne
Germline deletion of pantothenate kinases 1 and 2 reveals the key roles for CoA in postnatal metabolism.
Pantothenate kinase (PanK) phosphorylates pantothenic acid (vitamin B(5)) and controls the overall rate of coenzyme A (CoA) biosynthesis. Pank1 gene deletion in mice results in a metabolic phenotype where fatty acid oxidation and gluconeogenesis are impaired in the fasted state, leading to mild hypoglycemia. Inactivating mutations in the human PANK2 gene lead to childhood neurodegeneration, but Pank2 gene inactivation in mice does not elicit a phenotype indicative of the neuromuscular symptoms or brain iron accumulation that accompany the human disease. Pank1/Pank2 double knockout (dKO) mice were derived to determine if the mild phenotypes of the single knockout mice are due to the ability of the two isoforms to compensate for each other in CoA biosynthesis. Postnatal development was severely affected in the dKO mice. The dKO pups developed progressively severe hypoglycemia and hyperketonemia by postnatal day 10 leading to death by day 17. Hyperketonemia arose from impaired whole-body ketone utilization illustrating the requirement for CoA in energy generation from ketones. dKO pups had reduced CoA and decreased fatty acid oxidation coupled with triglyceride accumulation in liver. dKO hepatocytes could not maintain the NADH levels compared to wild-type hepatocytes. These results revealed an important link between CoA and NADH levels, which was reflected by deficiencies in hepatic oleate synthesis and gluconeogenesis. The data indicate that PanK1 and PanK2 can compensate for each other to supply tissue CoA, but PanK1 is more important to CoA levels in liver whereas PanK2 contributes more to CoA synthesis in the brain. PMID:22815849
Garcia, Matthew; Leonardi, Roberta; Zhang, Yong-Mei; Rehg, Jerold E; Jackowski, Suzanne
hCOA3 Stabilizes Cytochrome c Oxidase 1 (COX1) and Promotes Cytochrome c Oxidase Assembly in Human Mitochondria*
Cytochrome c oxidase (COX) or complex IV of the mitochondrial respiratory chain plays a fundamental role in energy production of aerobic cells. In humans, COX deficiency is the most frequent cause of mitochondrial encephalomyopathies. Human COX is composed of 13 subunits of dual genetic origin, whose assembly requires an increasing number of nuclear-encoded accessory proteins known as assembly factors. Here, we have identified and characterized human CCDC56, an 11.7-kDa mitochondrial transmembrane protein, as a new factor essential for COX biogenesis. CCDC56 shares sequence similarity with the yeast COX assembly factor Coa3 and was termed hCOA3. hCOA3-silenced cells display a severe COX functional alteration owing to a decreased stability of newly synthesized COX1 and an impairment in the holoenzyme assembly process. We show that hCOA3 physically interacts with both the mitochondrial translation machinery and COX structural subunits. We conclude that hCOA3 stabilizes COX1 co-translationally and promotes its assembly with COX partner subunits. Finally, our results identify hCOA3 as a new candidate when screening for genes responsible for mitochondrial diseases associated with COX deficiency.
Clemente, Paula; Peralta, Susana; Cruz-Bermudez, Alberto; Echevarria, Lucia; Fontanesi, Flavia; Barrientos, Antoni; Fernandez-Moreno, Miguel A.; Garesse, Rafael
Swelling of acetylated wood in organic solvents
NASA Astrophysics Data System (ADS)
To investigate the affinity of acetylated wood for organic liquids, Yezo spruce wood specimens were acetylated with acetic anhydride, and their swelling in various liquids were compared to those of untreated specimens. The acetylated wood was rapidly and remarkably swollen in aprotic organic liquids such as benzene and toluene in which the untreated wood was swollen only slightly and/or very slowly. On the other hand, the swelling of wood in water, ethylene glycol and alcohols remained unchanged or decreased by the acetylation. Consequently the maximum volume of wood swollen in organic liquids was always larger than that in water. The effect of acetylation on the maximum swollen volume of wood was greater in liquids having smaller solubility parameters. The easier penetration of aprotic organic liquids into the acetylated wood was considered to be due to the scission of hydrogen bonds among the amorphous wood constituents by the substitution of hydroxyl groups with hydrophobic acetyl groups.
Obataya, E.; Shibutani, S.
Hydrolysis of the GlcNAc oxazoline: deamidation and acyl rearrangement.
The specific deamidation of 2-acetamido-1,3,4,6-tetra-O-acetyl-alpha-D-glucopyranose is achieved by p-toluenesulfonic acid-promoted hydrolysis of 2-methyl-(3,4,6-tri-O-acetyl-1,2-dideoxy-alpha-D-glucopyrano)-[2,1 -d]-2- oxazoline 2 to give quantitative formation of the 1,3,4,6-tetra-O-acetyl-2-amino-2-deoxy-alpha-D-glucopyranose p-toluenesulfonate (5d). This two-step procedure provides an amino sugar which may be readily acylated to give novel glycoconjugates. Alternatively, base-catalyzed O-1-->N-2 acyl rearrangement of the amino tosylate 5d gives the 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-D-glucopyranose 4 as a 9:1 mixture of alpha and beta anomers. Thus, hydrolysis of GlcNAc oxazoline 2 gives the amino-ester 5 as the kinetic product and the amido-alcohol 4 as the thermodynamic product. PMID:8548785
Jha, R; Davis, J T
Regulation of synapse composition by protein acetylation: the role of acetylated cortactin.
Protein acetylation affects synaptic plasticity and memory, but its effects on synapse composition have not been addressed. We found that protein acetylation promotes the dendritic clustering of the excitatory postsynaptic scaffold protein PSD95 in hippocampal neurons, without affecting the total levels of this protein. Cortactin, an F-actin-binding protein enriched in dendritic spines, is a substrate for acetylation and has a role in spine morphogenesis. Recent studies showed that cortactin acetylation changes its ability to bind F-actin and regulates cellular motility, but the function of cortactin acetylation in neuronal cells is so far unknown. We tested whether acetylation of cortactin influences its morphogenic function by overexpressing wild-type cortactin, or the mimetic mutants for acetylated or deacetylated cortactin, in hippocampal neurons, and found that cortactin acetylation has an impact on PSD95 clustering, independent from its function as actin dynamics regulator. Moreover, acetylated cortactin can rescue the reduction in PSD95 clustering mediated by knockdown of cortactin. We also found that acetylation of cortactin is correlated with decreased cortactin interaction with p140Cap and Shank1, and with lower cortactin phosphorylation at tyrosine 421. The neurotrophin BDNF promoted the acetylation of cortactin in hippocampal neurons, suggesting that BDNF may regulate excitatory synapses and PSD95 dendritic clustering at least in part by changing the acetylation level of cortactin. Our findings unravel an unsuspected role for cortactin acetylation in the regulation of PSD95 dendritic clustering, which may work in concert with cortactin's role in spine development. PMID:23038781
Catarino, Tatiana; Ribeiro, Lu�s; Santos, Sandra D; Carvalho, Ana Lu�sa
Economics of Enzymatic Hydrolysis Processes.
National Technical Information Service (NTIS)
Enzymatic hydrolysis processes have the ability to produce high yields of sugars for fermentation to fuel ethanol from lignocellulosic biomass. However, these systems have been plagued with yields, product concentrations, and reactions rates far below tho...
J. D. Wright
Mitochondrial Acetylation and Diseases of Aging
In recent years, protein lysine acetylation has emerged as a prominent and conserved regulatory posttranslational modification that is abundant on numerous enzymes involved in the processes of intermediary metabolism. Well-characterized mitochondrial processes of carbon utilization are enriched in acetyl-lysine modifications. Although seminal discoveries have been made in the basic biology of mitochondrial acetylation, an understanding of how acetylation states influence enzyme function and metabolic reprogramming during pathological states remains largely unknown. This paper will examine our current understanding of eukaryotic acetate metabolism and present recent findings in the field of mitochondrial acetylation biology. The implications of mitochondrial acetylation for the aging process will be discussed, as well as its potential implications for the unique and localized metabolic states that occur during the aging-associated conditions of heart failure and cancer growth.
Wagner, Gregory R.; Payne, R. Mark
Occurrence of naturally acetylated lignin units.
This work examines the occurrence of native acetylated lignin in a large set of vascular plants, including both angiosperms and gymnosperms, by a modification of the so-called Derivatization Followed by Reductive Cleavage (DFRC) method. Acetylated lignin units were found in the milled wood lignins of all angiosperms selected for this study, including mono- and eudicotyledons, but were absent in the gymnosperms analyzed. In some plants (e.g., abaca, sisal, kenaf, or hornbeam), lignin acetylation occurred at a very high extent, exceeding 45% of the uncondensed (alkyl-aryl ether linked) syringyl lignin units. Acetylation was observed exclusively at the gamma-carbon of the lignin side chain and predominantly on syringyl units, although a predominance of acetylated guaiacyl over syringyl units was observed in some plants. In all cases, acetylation appears to occur at the monomer stage, and sinapyl and coniferyl acetates seem to behave as real lignin monomers participating in lignification. PMID:17552541
Del R�o, Jos� C; Marques, Gisela; Rencoret, Jorge; Mart�nez, Angel T; Guti�rrez, Ana
Hydrolysis reactor for hydrogen production
In accordance with certain embodiments of the present disclosure, a method for hydrolysis of a chemical hydride is provided. The method includes adding a chemical hydride to a reaction chamber and exposing the chemical hydride in the reaction chamber to a temperature of at least about 100.degree. C. in the presence of water and in the absence of an acid or a heterogeneous catalyst, wherein the chemical hydride undergoes hydrolysis to form hydrogen gas and a byproduct material.
Davis, Thomas A.; Matthews, Michael A.
Enzymatic hydrolysis of bacterial cellulose
Native cellulose from the bacterium Acetobacter xylinum as well as acid-treated bacterial cellulose prepared from partial hydrolysis of the native bacterial cellulose with 2.5 N HCl were subjected to enzymatic hydrolysis by Trichoderma viride cellobiohydrolase I (CBH I) and endoglucanase II (EG II). The activities of the two enzymes were continuously monitored with an oxidation-reduction potential electrode based on the
Masahiro Samejima; Junji Sugiyama; Kiyohiko Igarashi; Karl-Erik L. Eriksson
AcK-knowledge Reversible Acetylation
NSDL National Science Digital Library
In 1966, the histone was identified as the first protein subject to reversible acetylation. The ensuing 30 years of research on histone acetylation has been critical for elucidating how gene transcription and chromatin remodeling are regulated at the molecular level. This central focus on histones, however, has also restricted our understanding of reversible acetylation, and therefore the enzymes that catalyze this reaction, to cellular processes predominantly associated with chromatin. The study of reversible acetylation has become more or less synonymous with histone acetylation. Recent developments—including increased ability to detect acetylated proteins, the characterization of novel acetyltransferases and deacetylases, and the identification of specific inhibitors for these enzymes—have revealed that this histone-central paradigm probably reflects only a fraction of the cellular processes regulated by reversible acetylation. New studies have uncovered unexpected roles for reversible acetylation in many diverse areas, thereby establishing protein acetylation as a highly versatile signaling modification that has functions beyond gene transcription and chromatin remodeling.
Todd Cohen (Duke University;Department of Pharmacology and Cancer Biology REV); Tso-Pang Yao (Duke University;Department of Pharmacology and Cancer Biology REV)
Age-associated mitochondrial oxidative decay: Improvement of carnitine acetyltransferase substrate-binding affinity and activity in brain by feeding old rats acetyl-l- carnitine and/or R-?-lipoic acid
We test whether the dysfunction with age of carnitine acetyltransferase (CAT), a key mitochondrial enzyme for fuel utilization, is due to decreased binding affinity for substrate and whether this substrate, fed to old rats, restores CAT activity. The kinetics of CAT were analyzed by using the brains of young and old rats and of old rats supplemented for 7 weeks with the CAT substrate acetyl-l-carnitine (ALCAR) and/or the mitochondrial antioxidant precursor R-?-lipoic acid (LA). Old rats, compared with young rats, showed a decrease in CAT activity and in CAT-binding affinity for both substrates, ALCAR and CoA. Feeding ALCAR or ALCAR plus LA to old rats significantly restored CAT-binding affinity for ALCAR and CoA, and CAT activity. To explore the underlying mechanism, lipid peroxidation and total iron and copper levels were assayed; all increased in old rats. Feeding old rats LA or LA plus ALCAR inhibited lipid peroxidation but did not decrease iron and copper levels. Ex vivo oxidation of young-rat brain with Fe(II) caused loss of CAT activity and binding affinity. In vitro oxidation of purified CAT with Fe(II) inactivated the enzyme but did not alter binding affinity. However, in vitro treatment of CAT with the lipid peroxidation products malondialdehyde or 4-hydroxy-nonenal caused a decrease in CAT-binding affinity and activity, thus mimicking age-related change. Preincubation of CAT with ALCAR or CoA prevented malondialdehyde-induced dysfunction. Thus, feeding old rats high levels of key mitochondrial metabolites can ameliorate oxidative damage, enzyme activity, substrate-binding affinity, and mitochondrial dysfunction.
Liu, Jiankang; Killilea, David W.; Ames, Bruce N.
Stable thiobarbituric acid chromophore with dimethyl sulphoxide. Application to sialic acid assay in analytical de-O-acetylation.
With dimethyl sulphoxide instead of butanol in the thiobarbituric acid assay for sialic acid, a non-fading chromophore with lambdamax. = 549 nm was produced in a homogeneous solution, allowing dilution of the test mixture in case of high colour yield. This test adapted well to studies on alkaline de-O-acetylation. Bovine and rat submaxillary mucins, and rabbit Tamm-Horsfall urinary sialoproteins contain O-acetyl isomers of neuramine acid that are resistant to the thiobarbituric acid assay. Alkaline de-O-acetylation converted resistant O-acetylneuraminic acid into thiobarbituric acid-reactive sialic acid, and such conversion paralleled de-O-acetylation as measured by the ferric hydroxamate method. The colour increment was similar when the alkaline treatment of bovine submaxillary mucin either preceded or followed the acid hydrolysis. Only alkaline preptreatment was effective with rat submaxillary mucin. By selecting optimal conditions for alkaline de-O-acetylation, O-acetyl isomers can be accurately assessed by the thiobarbituric acid assay.
Skoza, L; Mohos, S
Molecular cloning of acetyl coenzyme A: deacetylcephalosporin C o-acetyltransferase cDNA from Acremonium chrysogenum: sequence and expression of catalytic activity in yeast.
Acetyl CoA: deacetylcephalosporin C o-acetyltransferase(DCPC-ATF) catalyses the final step in the biosynthesis of cephalosporin C, the conversion of deacetylcephalosporin C to cephalosporin C. A cDNA encoding DCPC-ATF has been isolated from a cDNA library of a cephalosporin C producing fungus Acremonium chrysogenum using oligonucleotide probes based on N-terminal amino acid sequences of the enzyme. The cDNA contains a single large open reading frame for a putative precursor consisting of 12 amino acid(AA) leader peptide of unknown function, 274 AA large subunit and 126 AA small subunit at the carboxyl end. The cDNA was expressed in yeast exhibiting a functional DCPC-ATF activity. It was also indicated that the leader peptide was not essential for expression of the enzyme activity. The primary structure of DCPC-ATF shows significant homology with those of acetyl CoA: homoserine o-acetyltransferase in Saccharomyces cerevisiae and Ascobolas immersus. PMID:1540196
Matsuda, A; Sugiura, H; Matsuyama, K; Matsumoto, H; Ichikawa, S; Komatsu, K
Insulin Signaling Regulates Fatty Acid Catabolism at the Level of CoA Activation
The insulin/IGF signaling pathway is a highly conserved regulator of metabolism in flies and mammals, regulating multiple physiological functions including lipid metabolism. Although insulin signaling is known to regulate the activity of a number of enzymes in metabolic pathways, a comprehensive understanding of how the insulin signaling pathway regulates metabolic pathways is still lacking. Accepted knowledge suggests the key regulated step in triglyceride (TAG) catabolism is the release of fatty acids from TAG via the action of lipases. We show here that an additional, important regulated step is the activation of fatty acids for beta-oxidation via Acyl Co-A synthetases (ACS). We identify pudgy as an ACS that is transcriptionally regulated by direct FOXO action in Drosophila. Increasing or reducing pudgy expression in vivo causes a decrease or increase in organismal TAG levels respectively, indicating that pudgy expression levels are important for proper lipid homeostasis. We show that multiple ACSs are also transcriptionally regulated by insulin signaling in mammalian cells. In sum, we identify fatty acid activation onto CoA as an important, regulated step in triglyceride catabolism, and we identify a mechanistic link through which insulin regulates lipid homeostasis.
Xu, Xiaojun; Gopalacharyulu, Peddinti; Seppanen-Laakso, Tuulikki; Ruskeepaa, Anna-Liisa; Aye, Cho Cho; Carson, Brian P.; Mora, Silvia; Oresic, Matej; Teleman, Aurelio A.
Enzymatic Hydrolysis of Cellulosic Biomass
Biological conversion of cellulosic biomass to fuels and chemicals offers the high yields to products vital to economic success and the potential for very low costs. Enzymatic hydrolysis that converts lignocellulosic biomass to fermentable sugars may be the most complex step in this process due to substrate-related and enzyme-related effects and their interactions. Although enzymatic hydrolysis offers the potential for higher yields, higher selectivity, lower energy costs, and milder operating conditions than chemical processes, the mechanism of enzymatic hydrolysis and the relationship between the substrate structure and function of various glycosyl hydrolase components are not well understood. Consequently, limited success has been realized in maximizing sugar yields at very low cost. This review highlights literature on the impact of key substrate and enzyme features that influence performance to better understand fundamental strategies to advance enzymatic hydrolysis of cellulosic biomass for biological conversion to fuels and chemicals. Topics are summarized from a practical point of view including characteristics of cellulose (e.g., crystallinity, degree of polymerization, and accessible surface area) and soluble and insoluble biomass components (e.g., oligomeric xylan, lignin, etc.) released in pretreatment, and their effects on the effectiveness of enzymatic hydrolysis. We further discuss the diversity, stability, and activity of individual enzymes and their synergistic effects in deconstructing complex lignocellulosic biomass. Advanced technologies to discover and characterize novel enzymes and to improve enzyme characteristics by mutagenesis, post-translational modification, and over-expression of selected enzymes and modifications in lignocellulosic biomass are also discussed.
Yang, Bin; Dai, Ziyu; Ding, Shi-You; Wyman, Charles E.
Dispersed fluorescence observations of the CO/A 1Pi to X 1Sigma+/ transitions from photodissociation of CO2
NASA Technical Reports Server (NTRS)
The spectra of vacuum ultraviolet (vuv) fluorescence resulting from the excitation of CO2 by photons from an intense line emission source at 15 wavelengths in the range 449-955 A were obtained. The vibrational population distributions for the v = 0, 1, and 2 levels of the CO(A 1Pi) fragments were obtained at several incident photon wavelengths from 700 to 923 A. At incident photon wavelengths of 901 and 923, the relative intensities of the CO(A to X) bands were determined, permitting examination of the variation of the electronic transition moment with the r centroid.
Phillips, E.; Lee, C. L.; Judge, D. L.
40 CFR 721.10520 - Acetylated fatty acid glycerides (generic).
Code of Federal Regulations, 2013 CFR
...2013-07-01 false Acetylated fatty acid glycerides (generic). 721...Substances § 721.10520 Acetylated fatty acid glycerides (generic). (a...identified generically as acetylated fatty acid glycerides (PMN...
Reverse genetic characterization of two paralogous acetoacetyl CoA thiolase genes in Arabidopsis reveals their importance in plant growth and development
Acetoacetyl CoA thiolase (AACT, EC 18.104.22.168) catalyzes the condensation of two acetyl?CoA molecules to form acetoacetyl?CoA. Two AACT?encoding genes, At5g47720 (AACT1) and At5g48230 (AACT2), were functionally identified in the Arabidopsis genome by direct enzymological assays and functional expression in yeast. Promoter::GUS fusion experiments indicated that AACT1 is primarily expressed in the vascular system and AACT2 is highly expressed in root tips, young leaves, top stems and anthers. Characterization of T?DNA insertion mutant alleles at each AACT locus established that AACT2 function is required for embryogenesis and for normal male gamete transmission. In contrast, plants lacking AACT1 function are completely viable and show no apparent growth phenotypes, indicating that AACT1 is functionally redundant with respect to AACT2 function. RNAi lines that express reduced levels of AACT2 show pleiotropic phenotypes, including reduced apical dominance, elongated life span and flowering duration, sterility, dwarfing, reduced seed yield and shorter root length. Microscopic analysis reveals that the reduced stature is caused by a reduction in cell size and fewer cells, and male sterility is caused by loss of the pollen coat and premature degeneration of the tapetal cells. Biochemical analyses established that the roots of AACT2 RNAi plants show quantitative and qualitative alterations in phytosterol profiles. These phenotypes and biochemical alterations are reversed when AACT2 RNAi plants are grown in the presence of mevalonate, which is consistent with the role of AACT2 in generating the bulk of the acetoacetyl?CoA precursor required for the cytosol?localized, mevalonate?derived isoprenoid biosynthetic pathway.
Jin, Huanan; Song, Zhihong; Nikolau, Basil J.
The cardiac acetyl-lysine proteome.
In the heart, lysine acetylation has been implicated in processes ranging from transcriptional control of pathological remodeling, to cardioprotection arising from caloric restriction. Given the emerging importance of this post-translational modification, we used a proteomic approach to investigate the broader role of lysine acetylation in the heart using a guinea pig model. Briefly, hearts were fractionated into myofilament-, mitochondrial- and cytosol-enriched fractions prior to proteolysis and affinity-enrichment of acetylated peptides. LC-MS/MS analysis identified 1075 acetylated peptides, harboring 994 acetylation sites that map to 240 proteins with a global protein false discovery rate <0.8%. Mitochondrial targets account for 59% of identified proteins and 64% of sites. The majority of the acetyl-proteins are enzymes involved in fatty acid metabolism, oxidative phosphorylation or the TCA cycle. Within the cytosolic fraction, the enzymes of glycolysis, fatty acid synthesis and lipid binding are prominent. Nuclear targets included histones and the transcriptional regulators E1A(p300) and CREB binding protein. Comparison of our dataset with three previous global acetylomic studies uniquely revealed 53 lysine-acetylated proteins. Specifically, newly-identified acetyl-proteins include Ca(2+)-handling proteins, RyR2 and SERCA2, and the myofilament proteins, myosin heavy chain, myosin light chains and subunits of the Troponin complex, among others. These observations were confirmed by anti-acetyl-lysine immunoblotting. In summary, cardiac lysine acetylation may play a role in cardiac substrate selection, bioenergetic performance, and maintenance of redox balance. New sites suggest a host of potential mechanisms by which excitation-contraction coupling may also be modulated. PMID:23844019
Foster, D Brian; Liu, Ting; Rucker, Jasma; O'Meally, Robert N; Devine, Lauren R; Cole, Robert N; O'Rourke, Brian
De-acetyl-cinobufalactam monohydrate.
The title compound, C24H33NO4�H2O, the reaction product of de-acetyl-cinobufagin with ammonium acetate, consists of three cyclo-hexane rings (A, B and C), one five-membered ring (D), one six-membered lactone ring (E) and an epoxide ring (F). The stereochemistry of the ring junctures are A/B cis, B/C trans, C/D cis and D/F cis. Cyclo-hexane rings A, B and C have normal chair conformations. The five-membered ring D adopts an envelope conformation (with the C atom bearing the lactone ring as the flap) and the lactone ring E is planar. In the crystal, hy-droxy and water O-H?O and amine N-H?O hydrogen bonds involving carbonyl, hy-droxy and water O-atom acceptors link the mol-ecules into a three-dimensional network. PMID:24940236
Tang, Hong-Jin; Yuan, Xiao-Feng; Tian, Hai-Yan; Ruan, Li-Jun; Jiang, Ren-Wang
De-acetyl-cinobufalactam monohydrate
The title compound, C24H33NO4�H2O, the reaction product of de�acetyl�cinobufagin with ammonium acetate, consists of three cyclo�hexane rings (A, B and C), one five-membered ring (D), one six-membered lactone ring (E) and an epoxide ring (F). The stereochemistry of the ring junctures are A/B cis, B/C trans, C/D cis and D/F cis. Cyclo�hexane rings A, B and C have normal chair conformations. The five-membered ring D adopts an envelope conformation (with the C atom bearing the lactone ring as the flap) and the lactone ring E is planar. In the crystal, hy�droxy and water O�H?O and amine N�H?O hydrogen bonds involving carbonyl, hy�droxy and water O-atom acceptors link the mol�ecules into a three-dimensional network.
Tang, Hong-Jin; Yuan, Xiao-Feng; Tian, Hai-Yan; Ruan, Li-Jun; Jiang, Ren-Wang
The COA supplies drinking water to a number of municipalities in central Oklahoma. Two major stratigraphic units in the COA, the Garber Sandstone and Wellington Formation, contain naturally occurring arsenic that exceeds government mandated drinking-water standards (EPA, 2001). ...
The hydrolysis kinetics of 4-chlorostilbene oxide (CSO) in buffered distilled water, in natural waters, and in sediment associated water are reported. he disappearance of CSO followed pseudo-first-order kinetics in buffered water over the experimental pH range of 3 to 11. elow pH...
Succinyl CoA: 3-oxoacid CoA transferase (SCOT): human cDNA cloning, human chromosomal mapping to 5p13, and mutation detection in a SCOT-deficient patient.
Succinyl CoA: 3-oxoacid CoA transferase (SCOT; E.C.22.214.171.124) mediates the rate-determining step of ketolysis in extrahepatic tissues, the esterification of acetoacetate to CoA for use in energy production. Hereditary SCOT deficiency in humans causes episodes of severe ketoacidosis. We obtained human-heart SCOT cDNA clones spanning the entire 1,560-nt coding sequence. Sequence alignment of the human SCOT peptides with other known CoA transferases revealed several conserved regions of potential functional importance. A single approximately 3.2-kb SCOT mRNA is present in human tissues (heart > leukocytes >> fibroblasts), but no signal is detectable in the human hepatoma cell line HepG2. We mapped the human SCOT locus (OXCT) to the cytogenetic band 5p13 by in situ hybridization. From fibroblasts of a patient with hereditary SCOT deficiency, we amplified and cloned cDNA fragments containing the entire SCOT coding sequence. We found a homozygous C-to-G transversion at nt 848, which changes the Ser 283 codon to a stop codon. This mutation (S283X) is incompatible with normal enzyme function and represents the first documentation of a pathogenic mutation in SCOT deficiency. Images Figure 2 Figure 6
Kassovska-Bratinova, S.; Fukao, T.; Song, X. Q.; Duncan, A. M.; Chen, H. S.; Robert, M. F.; Perez-Cerda, C.; Ugarte, M.; Chartrand, C.; Vobecky, S.; Kondo, N.; Mitchell, G. A.
Succinyl CoA: 3-oxoacid CoA transferase (SCOT): Human cDNA cloning, human chromosomal mapping to 5p13, and mutation detection in a SCOT-deficient patient
Succinyl CoA: 3-oxoacid CoA transferase (SCOT; E.C.22.214.171.124) mediates the rate-determining step of ketolysis in extrahepatic tissues, the esterification of acetoacetate to CoA for use in energy production. Hereditary SCOT deficiency in humans causes episodes of severe ketoacidosis. We obtained human-heart SCOT cDNA clones spanning the entire 1,560-nt coding sequence. Sequence alignment of the human SCOT peptides with other known CoA transferases revealed several conserved regions of potential functional importance. A single {approximately}3.2-kb SCOT mRNA is present in human tissues (heart > leukocytes {much_gt} fibroblasts), but no signal is detectable in the human hepatoma cell line HepG2. We mapped the human SCOT locus (OXCT) to the cytogenetic band 5p13 by in situ hybridization. From fibroblasts of a patient with hereditary SCOT deficiency, we amplified and cloned cDNA fragments containing the entire SCOT coding sequence. We found a homozygous C-to-G transversion at nt 848, which changes the Ser 283 codon to a stop codon. This mutation (S283X) is incompatible with normal enzyme function and represents the first documentation of a pathogenic mutation in SCOT deficiency. 45 refs., 6 figs.
Kassovska-Bratinova, S.; Robert, M.F.; Mitchell, G.A. [Gifu Univ. School of Medicine (Japan)] [and others
Identification of a Colorectal Tumor-Associated Antigen (COA-1) Recognized by CD4+ T Lymphocytes
Only a limited number of target molecules have been shown to be recognized by colon tumor-reactive T cells, limiting the options for the development of immunotherapies for patients with colon cancer. The current studies were undertaken in an attempt to generate tumor-reactive T cells that could be used to identify and characterize novel colon tumor-associated antigens. Multiple CD4+ T-cell clones isolated either from tumor-infiltrating lymphocytes or peripheral blood mononuclear cells that were sensitized in vitro with autologous tumor cells from a colon cancer patient, 1869, recognized autologous tumor cells in a class II HLA-DR-restricted manner. One of the peripheral blood mononuclear cell clones, clone C111, was used to screen pools of clones that were generated from an autologous colon tumor cell line cDNA library. A cDNA clone that was isolated encoded a protein that was termed colorectal tumor-associated antigen-1 (COA-1). This product was recognized in the context of the two autologous HLA-DR?1 alleles, HLA-DR?1*0402 and DR?1*1301. The nucleotide sequence of the COA-1 transcript was nearly identical to multiple expressed sequence tag sequences that encode variants of Socius, a protein that was found recently to bind to members of the Rnd family of GTPases. The COA-1 gene was expressed at relatively comparable levels in colorectal and melanoma tumor cells, EBV-infected B cells, normal B cells, and cultured fibroblast cell lines. However, the gene that was isolated from normal cell types contained a single nucleotide substitution, resulting in an amino acid change near the COOH terminus of the protein. Although the minimal epitope recognized by CD4+ cells was encoded by sequences that were upstream from this substitution, C111 T cells did not appear to recognize the normal gene product. Therefore, this alteration may either affect the localization or the processing of this gene product, which may at least in part be responsible for the differential recognition of tumor and normal cells.
Maccalli, Cristina; Li, Yong F.; El-Gamil, Mona; Rosenberg, Steven A.; Robbins, Paul F.
Comparative effects of (?)-hydroxycitrate and (+)- allo -hydroxycitrate on acetyl CoA carboxylase and fatty acid and cholesterol synthesis in vivo
(?)-Hydroxycitrate and (+)-allo-hydroxycitrate were investigated for their effects on lipid synthesis in vivo under conditions of either high carbohydrate\\u000a feeding or 24 hr fasting. Changes in rates of lipid synthesis resulting from the oral administration of these compounds were\\u000a monitored with the use of radiolabeled H2O, alanine, and acetate. In the fed rat, (?)-hydroxycitrate significantly reduced the incorporation of H2O
Joseph Triscari; Ann C. Sullivan
Levoglucosan, cellobiose and their acetates as model compounds for the thermally assisted hydrolysis and methylation of cellulose and cellulose acetate
Levoglucosan (1,6-anhydro-?-d-glucopyranose), cellobiose (?-d-glucopyranosyl-[1?4]-d-glucopyranose), tri-O-acetyl-levoglucosan (1,6-anhydro-?-d-glucopyranose-2,3,4-triacetate) and cellobiose octaacetate have been studied with regard to their suitability as model compounds for thermally assisted hydrolysis and methylation with tetramethylammonium hydroxide of cellulose and cellulose acetate. In addition, the results of analytical pyrolysis of methyl cellulose were compared with those of thermally assisted hydrolysis and methylation, to distinguish between products arising from
C Schwarzinger; I Tanczos; H Schmidt
Nonhistone protein acetylation as cancer therapy targets
Acetylation and deacetylation are counteracting, post-translational modifications that affect a large number of histone and nonhistone proteins. The significance of histone acetylation in the modification of chromatin structure and dynamics, and thereby gene transcription regulation, has been well recognized. A steadily growing number of nonhistone proteins have been identified as acetylation targets and reversible lysine acetylation in these proteins plays an important role(s) in the regulation of mRNA stability, protein localization and degradation, and protein�protein and protein�DNA interactions. The recruitment of histone acetyltransferases (HATs) and histone deacetylases (HDACs) to the transcriptional machinery is a key element in the dynamic regulation of genes controlling cellular proliferation, differentiation and apoptosis. Many nonhistone proteins targeted by acetylation are the products of oncogenes or tumor-suppressor genes and are directly involved in tumorigenesis, tumor progression and metastasis. Aberrant activity of HDACs has been documented in several types of cancers and HDAC inhibitors (HDACi) have been employed for therapeutic purposes. Here we review the published literature in this field and provide updated information on the regulation and function of nonhistone protein acetylation. While concentrating on the molecular mechanism and pathways involved in the addition and removal of the acetyl moiety, therapeutic modalities of HDACi are also discussed.
Singh, Brahma N; Zhang, Guanghua; Hwa, Yi L; Li, Jinping; Dowdy, Sean C; Jiang, Shi-Wen
Acetylation modulates the STAT signaling code.
A fascinating question of modern biology is how a limited number of signaling pathways generate biological diversity and crosstalk phenomena in vivo. Well-defined posttranslational modification patterns dictate the functions and interactions of proteins. The signal transducers and activators of transcription (STATs) are physiologically important cytokine-induced transcription factors. They are targeted by a multitude of posttranslational modifications that control and modulate signaling responses and gene expression. Beyond phosphorylation of serine and tyrosine residues, lysine acetylation has recently emerged as a critical modification regulating STAT functions. Interestingly, acetylation can determine STAT signaling codes by various molecular mechanisms, including the modulation of other posttranslational modifications. Here, we provide an overview on the acetylation of STATs and how this protein modification shapes cellular cytokine responses. We summarize recent advances in understanding the impact of STAT acetylation on cell growth, apoptosis, innate immunity, inflammation, and tumorigenesis. Furthermore, we discuss how STAT acetylation can be targeted by small molecules and we consider the possibility that additional molecules controlling STAT signaling are regulated by acetylation. Our review also summarizes evolutionary aspects and we show similarities between the acetylation-dependent control of STATs and other important molecules. We propose the concept that, similar to the 'histone code', distinct posttranslational modifications and their crosstalk orchestrate the functions and interactions of STAT proteins. PMID:22795479
Wieczorek, Martin; Ginter, Torsten; Brand, Peter; Heinzel, Thorsten; Kr�mer, Oliver H
Acetylation of rice straw for thermoplastic applications.
An inexpensive and biodegradable thermoplastic was developed through acetylation of rice straw (RS) with acetic anhydride. Acetylation conditions were optimized. The structure and properties of acetylated RS were characterized by fourier transform infrared (FTIR), solid-state (13)C NMR spectroscopy, X-ray diffractometer (XRD), scanning electron microscope (SEM), differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). The results showed that acetylation of RS has successfully taken place, and comparing with raw RS, the degree of crystallinity decreased and the decomposition rate was slow. The acetylated RS has got thermoplasticity when weight ratio of RS and acetic anhydride was 1:3, using sulphuric acid (9% to RS) as catalyst in glacial acetic acid 35�C for 12h, and the dosage of solvent was 9 times RS, in which weight percent gain (WPG) of the modified RS powder was 35.5% and its percent acetyl content was 36.1%. The acetylated RS could be formed into transparent thin films with different amount of plasticizer diethyl phthalate (DEP) using tape casting technology. PMID:23688473
Zhang, Guangzhi; Huang, Kai; Jiang, Xue; Huang, Dan; Yang, Yiqi
Mitochondrial protein acetylation is driven by acetyl-CoA from fatty acid oxidation.
Mitochondria integrate metabolic networks for maintaining bioenergetic requirements. Deregulation of mitochondrial metabolic networks can lead to mitochondrial dysfunction, which is a common hallmark of many diseases. Reversible post-translational protein acetylation modifications are emerging as critical regulators of mitochondrial function and form a direct link between metabolism and protein function, via the metabolic intermediate acetyl-CoA. Sirtuins catalyze protein deacetylation, but how mitochondrial acetylation is determined is unclear. We report here a mechanism that explains mitochondrial protein acetylation dynamics in vivo. Food withdrawal in mice induces a rapid increase in hepatic protein acetylation. Furthermore, using a novel LC-MS/MS method, we were able to quantify protein acetylation in human fibroblasts. We demonstrate that inducing fatty acid oxidation in fibroblasts increases protein acetylation. Furthermore, we show by using radioactively labeled palmitate that fatty acids are a direct source for mitochondrial protein acetylation. Intriguingly, in a mouse model that resembles human very-long chain acyl-CoA dehydrogenase (VLCAD) deficiency, we demonstrate that upon food-withdrawal, hepatic protein hyperacetylation is absent. This indicates that functional fatty acid oxidation is necessary for protein acetylation to occur in the liver upon food withdrawal. Furthermore, we now demonstrate that protein acetylation is abundant in human liver peroxisomes, an organelle where acetyl-CoA is solely generated by fatty acid oxidation. Our findings provide a mechanism for metabolic control of protein acetylation, which provides insight into the pathophysiogical role of protein acetylation dynamics in fatty acid oxidation disorders and other metabolic diseases associated with mitochondrial dysfunction. PMID:24516071
Pougovkina, Olga; Te Brinke, Heleen; Ofman, Rob; van Cruchten, Arno G; Kulik, Wim; Wanders, Ronald J A; Houten, Sander M; de Boer, Vincent C J
Semicontinuous enzymatic hydrolysis of lignocelluloses.
Lignocelluloses (steamed hardwood and hardwood kraft pulp) were semicontinuously hydrolyzed on a large scale [2-2. 5 kg of substrate vs. 20, 000 IU filter paperase (FPase)] using a 10-L hydrolysis reactor with an ultrafiltration unit for the recovery and reuse of cellulases. The substrate was added to the reactor at appropriate intervals to keep a concentration of approximately 5% (w/v). All of the enzyme was added at the beginning and no further addition was done. The ultrafiltration unit was operated intermittently rather than continuously due to its enough capacity (dilution rate of 2.5 h(-1)) and making the enzyme durable. The enzyme required to produce one gram of reducing sugar in this reactor was 27.3 FPase IU/g RS for steamed hardwood and 7.4 FPase IU/g RS for hardwood kraft pulp. The sugar composition of hydrolyzate was unaltered virtually from beginning to end of the hydrolysis in spite of the progressive loss of enzyme activities. The analysis of the enzyme composition in the hydrolyzate during hydrolysis revealed that an exo-beta-D-glucanase component was adsorbed selectively at the stages of advanced hydrolysis extent. PMID:18597319
Ishihara, M; Uemura, S; Hayashi, N; Shimizu, K
National Technical Information Service (NTIS)
The Energetics and Warheads Division of the U.S. Army Armament Research, Development and Engineering Center has been involved in the development of CL-20. An alkaline hydrolysis study was performed to better understand the fate and transport of CL-2O thro...
P. Karakaya M. Sidhourn C. Christodoulatos W. Balas S. Nicolich
Improved enzymatic hydrolysis of hair.
An enzymatic hair extraction method is proposed for drug analysis. Pronase digestion of various aliquots of hair from a cocaine abuser was preceded by a 2-h incubation with a dithiothreitol solution. The extraction solution was tested to identify possible interferences in the radioimmunoassay and was compared with other hydrolysis methods to assess the results of extraction. PMID:8138218
Offidani, C; Strano Rossi, S; Chiarotti, M
Stearoyl CoA desaturase (SCD) gene polymorphisms in Italian cattle breeds.
Stearoyl CoA desaturase (SCD) is the key enzyme involved in the endogenous synthesis of conjugated linoleic acid (CLA) in ruminants. Changes in the enzymatic activity as a result of SCD gene polymorphism and regulation have been hypothesized to cause diet-independent variations of CLA content in milk. Evidences for the direct influence of SCD polymorphism on fatty acid composition of milk and beef have also been reported. To evaluate genetic differences because of breed and/or selection goal, we investigated the polymorphism of three previously reported single nucleotide polymorphisms located in exon 5 of the SCD gene in 11 cattle breeds raised in Italy and selected for different production goals. Results obtained: (i) evidenced a high variability in the allele frequencies across breeds; (ii) detected three novel haplotypes, one of which is private to indigenous beef breeds, and (iii) showed a significant association between haplotypes and selective goal. PMID:18254828
Milanesi, E; Nicoloso, L; Crepaldi, P
Determination of 3-hydroxy-3-methylglutaryl CoA reductase activity in plants.
The enzyme 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) reductase catalyzes the NADPH-mediated reductive deacylation of HMG-CoA to mevalonic acid, which is the first committed step of the mevalonate pathway for isoprenoid biosynthesis. In agreement with its key regulatory role in the pathway, plant HMG-CoA reductase is modulated by many diverse external stimuli and endogenous factors and can be detected to variable levels in every plant tissue. A fine determination of HMG-CoA reductase activity levels is required to understand its contribution to plant development and adaptation to changing environmental conditions. Here, we report a procedure to reliably determine HMG-CoA reductase activity in plants. The method includes the sample collection and homogenization strategies as well as the specific activity determination based on a classical radiochemical assay. PMID:24777788
Campos, Narciso; Arr�, Montserrat; Ferrer, Albert; Boronat, Albert
Phthalate Ester Hydrolysis: Linear Free Energy Relationships.
National Technical Information Service (NTIS)
Alkaline hydrolysis rate constants were measured for dimethyl, diethyl, di-n-butyl, di-iso-butyl, and di-(2-ethylhexyl) phthalate esters in water. A linear free energy relationship (LFER) was established for estimating alkaline hydrolysis rate constants f...
N. L. Wolfe W. C. Steen L. A. Burns
Stability of acetyl-1-carnitine in 5% dextrose using a high-performance liquid chromatography-mass spectrometry times 2 method.
A stability-indicating high-performance liquid chromatography-mass spectrometry times 2 method was developed to establish the stability of acetyl-l-carnitine dissolved in 5% dextrose in water; quantitation of acetyl-l-carnitine and its hydrolysis product I-carnitine was performed using this method. Acetyl-l-carnitine dissolved in water was stress-degraded at a pH range of 3 to 12, and conversion to l-carnitine was quantified over 18 hours. The method was further validated by stressing the acetyl-l-carnitine solution at 68 degrees C, 82 degrees C, and 90 degrees C for up to 10 days, yielding a temperature-dependent hydrolysis rate constant. Acetyl-l-carnitine solutions were stored at 25 degrees C and 4 degrees C to 8 degrees C for 33 days to validate the kinetics prediction. The liquid chromatography-mass spectrometry times 2 method was sensitive and specific, allowing rapid separation and simultaneous quantitation of acetyl-l-carnitine and l-carnitine. Acetyl-l-carnitine dissolved in aqueous solutions is stable at neutral to acidic pH, but unstable at pH > 9. After 1 hour storage at room temperature, only 72.6% of acetyl-l-carnitine was left at pH 11 and 4.2% left at pH 12. The kinetics relationship between temperature and rate constant was In(k) = -8650.1 /T + 20.344 (r2 = 0.9851) at pH 5.2. The time required to degrade 15% of acetyl-I-carnitine was estimated to be 38 days at 25 degrees C or 234 days at 8 degrees C, and was confirmed with actual storage stability testing. Acetyl-l-carnitine dissolved in water (pH 5.2) at concentrations of 1 and 10 mg/mL was found stable at room temperature or refrigerated for at least 33 days using the established stability-indicating method. Acetyl-l-carnitine solutions are not stable at basic pH. When reconstituted in water, acetyl-l-carnitine is stable for over 30 days at room temperature or under refrigeration. PMID:23050330
Zhang, Yang; Jiang, Hongliang; Hutson, Paul
Dilute-Acid Hydrolysis of Lignocellulosic Biomass
In recent years, treatment of lignocellulosic biomass with dilute sulfuric acid has been primarily used as a means of hemicellulose\\u000a hydrolysis and pretreatment for enzymatic hydrolysis of cellulose. A significant advancement has also been made in the area\\u000a of dilute acid hydrolysis of cellulose. An overview of reactor theory as it applies to the dilute acid hydrolysis and recent\\u000a developments
Y. Y. Lee; Prashant Iyer; R. W. Torget
In the title salt, C6H13N2O+�C6H2N3O7 ? (systematic name: 4-acetyl�piperazin-1-ium 2,4,6-tri�nitro�phenolate), the piperazin-1-ium ring has a slightly distorted chair conformation. In the picrate anion, the mean planes of the two o-NO2 and p-NO2 groups are twisted with respect to the benzene ring by 15.0?(2), 68.9?(4) and 4.4?(3)�, respectively. In the crystal, N�H?O hydrogen bonds are observed, linking the ions into an infinite chain along [010]. In addition, weak cation�anion C�H?O inter�molecular inter�actions and a weak ?�? stacking inter�action between the benzene rings of the anions, with an inter-centroid distance of 3.771?(8)?�, help to stabilize the crystal packing, giving an overall sheet structure lying parallel to (100). Disorder was modelled for one of the O atoms in one of the o-NO2 groups over two sites with an occupancy ratio of 0.57?(6):0.43?(6).
Kavitha, Channappa N.; Kaur, Manpreet; Jasinski, Jerry P.; Yathirajan, Hemmige S.
4-Acetyl-piperazinium picrate.
In the title salt, C6H13N2O(+)�C6H2N3O7 (-) (systematic name: 4-acetyl-piperazin-1-ium 2,4,6-tri-nitro-phenolate), the piperazin-1-ium ring has a slightly distorted chair conformation. In the picrate anion, the mean planes of the two o-NO2 and p-NO2 groups are twisted with respect to the benzene ring by 15.0?(2), 68.9?(4) and 4.4?(3)�, respectively. In the crystal, N-H?O hydrogen bonds are observed, linking the ions into an infinite chain along [010]. In addition, weak cation-anion C-H?O inter-molecular inter-actions and a weak ?-? stacking inter-action between the benzene rings of the anions, with an inter-centroid distance of 3.771?(8)?�, help to stabilize the crystal packing, giving an overall sheet structure lying parallel to (100). Disorder was modelled for one of the O atoms in one of the o-NO2 groups over two sites with an occupancy ratio of 0.57?(6):0.43?(6). PMID:24940287
Kavitha, Channappa N; Kaur, Manpreet; Jasinski, Jerry P; Yathirajan, Hemmige S
Regulation of RAS oncogenicity by acetylation
Members of the RAS small GTPase family regulate cellular responses to extracellular stimuli by mediating the flux through downstream signal transduction cascades. RAS activity is strongly dependent on its subcellular localization and its nucleotide-binding status, both of which are modulated by posttranslational modification. We have determined that RAS is posttranslationally acetylated on lysine 104. Molecular dynamics simulations suggested that this modification affects the conformational stability of the Switch II domain, which is critical for the ability of RAS to interact with guanine nucleotide exchange factors. Consistent with this model, an acetylation-mimetic mutation in K-RAS4B suppressed guanine nucleotide exchange factor-induced nucleotide exchange and inhibited in vitro transforming activity. These data suggest that lysine acetylation is a negative regulatory modification on RAS. Because mutations in RAS family members are extremely common in cancer, modulation of RAS acetylation may constitute a therapeutic approach.
Yang, Moon Hee; Nickerson, Seth; Kim, Eric T.; Liot, Caroline; Laurent, Gaelle; Spang, Robert; Philips, Mark R.; Shan, Yibing; Shaw, David E.; Bar-Sagi, Dafna; Haigis, Marcia C.; Haigis, Kevin M.
Targeting bromodomains: epigenetic readers of lysine acetylation.
Lysine acetylation is a key mechanism that regulates chromatin structure; aberrant acetylation levels have been linked to the development of several diseases. Acetyl-lysine modifications create docking sites for bromodomains, which are small interaction modules found on diverse proteins, some of which have a key role in the acetylation-dependent assembly of transcriptional regulator complexes. These complexes can then initiate transcriptional programmes that result in phenotypic changes. The recent discovery of potent and highly specific inhibitors for the BET (bromodomain and extra-terminal) family of bromodomains has stimulated intensive research activity in diverse therapeutic areas, particularly in oncology, where BET proteins regulate the expression of key oncogenes and anti-apoptotic proteins. In addition, targeting BET bromodomains could hold potential for the treatment of inflammation and viral infection. Here, we highlight recent progress in the development of bromodomain inhibitors, and their potential applications in drug discovery. PMID:24751816
Filippakopoulos, Panagis; Knapp, Stefan
Bromodomain: an acetyl-lysine binding domain
Bromodomains, an extensive family of evolutionarily conserved protein modules originally found in proteins associated with chromatin and in nearly all nuclear histone acetyltransferases, have been recently discovered to function as acetyl-lysine binding domains. More recent structural studies of bromodomain\\/peptide ligand complexes have enriched our understanding of differences in ligand selectivity of bromodomains. These new findings demonstrate that bromodomain\\/acetyl-lysine recognition can
Lei Zeng; Ming-Ming Zhou
Conformational analysis of 1-acetyl-2-methylhydrazine
The conformational analysis of 1-acetyl-2-methylhydrazine was performed using DNMR spectroscopy and quantum chemical calculations (B3LYP\\/6-31+G*, AM1 and PM3). Activation barrier for interconversion of Z and E conformers was measured to be 16.1kcal\\/mol by DNMR. DFT calculations indicate that there are four minima on the 1-acetyl-2-methylhydrazine potential energy surface and the most stabile is Z conformer with the hydrogen bond between
O T�ubrik; P Burk; T Pehk; U M�eorg
Protein Acetylation in Procaryotes Increases Stress Resistance
Acetylation of lysine residues is conserved in all three kingdoms; however, its role in prokaryotes is unknown. Here we demonstrate that acetylation enables the reference bacterium Escherichia coli to withstand environmental stress. Specifically, the bacterium reaches higher cell densities and becomes more resistant to heat and oxidative stress when its proteins are acetylated as shown by deletion of the gene encoding acetyltransferase YfiQ and the gene encoding deacetylase CobB as well as by overproducing YfiQ and CobB. Furthermore, we show that the increase in oxidative stress resistance with acetylation is due to the induction of catalase activity through enhanced katG expression. We also found that two-component system proteins CpxA, PhoP, UvrY, and BasR are associated with cell catalase activity and may be responsible as the connection between bacterial acetylation and the stress response. This is the first demonstration of a specific environmental role of acetylation in prokaryotes.
Ma, Qun; Wood, Thomas K.
Protein acetylation in prokaryotes increases stress resistance.
Acetylation of lysine residues is conserved in all three kingdoms; however, its role in prokaryotes is unknown. Here we demonstrate that acetylation enables the reference bacterium Escherichia coli to withstand environmental stress. Specifically, the bacterium reaches higher cell densities and becomes more resistant to heat and oxidative stress when its proteins are acetylated as shown by deletion of the gene encoding acetyltransferase YfiQ and the gene encoding deacetylase CobB as well as by overproducing YfiQ and CobB. Furthermore, we show that the increase in oxidative stress resistance with acetylation is due to the induction of catalase activity through enhanced katG expression. We also found that two-component system proteins CpxA, PhoP, UvrY, and BasR are associated with cell catalase activity and may be responsible as the connection between bacterial acetylation and the stress response. This is the first demonstration of a specific environmental role of acetylation in prokaryotes. PMID:21703240
Ma, Qun; Wood, Thomas K
Studies on the Mechanism of Ring Hydrolysis in Phenylacetate Degradation
The widespread, long sought-after bacterial aerobic phenylalanine/phenylacetate catabolic pathway has recently been elucidated. It proceeds via coenzyme A (CoA) thioesters and involves the epoxidation of the aromatic ring of phenylacetyl-CoA, subsequent isomerization to an uncommon seven-membered C-O-heterocycle (oxepin-CoA), and non-oxygenolytic ring cleavage. Here we characterize the hydrolytic oxepin-CoA ring cleavage catalyzed by the bifunctional fusion protein PaaZ. The enzyme consists of a C-terminal (R)-specific enoyl-CoA hydratase domain (formerly MaoC) that cleaves the ring and produces a highly reactive aldehyde and an N-terminal NADP+-dependent aldehyde dehydrogenase domain that oxidizes the aldehyde to 3-oxo-5,6-dehydrosuberyl-CoA. In many phenylacetate-utilizing bacteria, the genes for the pathway exist in a cluster that contains an NAD+-dependent aldehyde dehydrogenase in place of PaaZ, whereas the aldehyde-producing hydratase is encoded outside of the cluster. If not oxidized immediately, the reactive aldehyde condenses intramolecularly to a stable cyclic derivative that is largely prevented by PaaZ fusion in vivo. Interestingly, the derivative likely serves as the starting material for the synthesis of antibiotics (e.g. tropodithietic acid) and other tropone/tropolone related compounds as well as for ?-cycloheptyl fatty acids. Apparently, bacteria made a virtue out of the necessity of disposing the dead-end product with ring hydrolysis as a metabolic branching point.
Teufel, Robin; Gantert, Carla; Voss, Michaela; Eisenreich, Wolfgang; Haehnel, Wolfgang; Fuchs, Georg
Purification and Characterization of Acetyl-Coenzyme A Carboxylase from Diclofop-Resistant and -Susceptible Lolium multiflorum.
Acetyl-coenzyme A carboxylase (ACCase) was purified >100-fold (specific activity 3.5 units mg-1) from leaf tissue of diclofopresistant and -susceptible biotypes of Lolium multiflorum. As determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the purified fractions from both biotypes contained a single 206-kD biotinylated polypeptide. The molecular mass of the native enzyme from both biotypes was approximately 520 kD. In some cases the native dimer from both biotypes dissociated during gel filtration to form a subunit of approximately 224 kD. The inclusion of 5% (w/v) polyethylene glycol 3350 (PEG) in the elution buffer prevented this dissociation. Steady-state substrate kinetics were analyzed in both the presence and absence of 5% PEG. For ACCase from both biotypes, addition of PEG increased the velocity 22% and decreased the apparent Km values for acetyl-coenzyme A (acetyl-CoA), but increased the Km values for bicarbonate and ATP. In the presence of PEG, the Km values for bicarbonate and ATP were approximately 35% higher for the enzyme from the susceptible biotype compared with the resistant enzyme. In the absence of PEG, no differences in apparent Km values were observed for the enzymes from the two biotypes. Inhibition constants (Ki app) were determined for CoA, malonyl-CoA, and diclofop. CoA was an S-hyperbolic (slope replots)-I-hyperbolic (intercept replots) noncompetitive inhibitor with respect to acetyl-CoA, with Ki app values of 711 and 795 [mu]M for enzymes from the resistant and susceptible biotypes, respectively. Malonyl-CoA competitively inhibited both enzymes (versus acetyl-CoA) with Ki app values of 140 and 104 [mu]M for ACCase from resistant and susceptible biotypes, respectively. Diclofop was a linear noncompetitive inhibitor of ACCase from the susceptible biotype and a nonlinear, or S-hyperbolic-I-hyperbolic, noncompetitive inhibitor of ACCase from the resistant biotype. For ACCase from the susceptible biotype the slope (Kis) and intercept (Kii) inhibition constants for diclofop versus acetyl-CoA were 0.08 and 0.44 [mu]M, respectively. ACCase from the resistant biotype had a Ki app value of 6.5 [mu]M. At a subsaturating acetyl-CoA concentration of 50 [mu]M, the Hill coefficients for diclofop binding were 0.61 and 1.2 for ACCase from the resistant and susceptible biotypes, respectively. The Hill coefficients for diclofop binding and the inhibitor replots suggest that the resistant form of ACCase exhibits negative cooperativity in binding diclofop. However, the possibility that the nonlinear inhibition of ACCase activity by diclofop in the enzyme fraction isolated from the resistant biotype is due to the presence of both resistant and susceptible forms of ACCase cannot be excluded.
Evenson, K. J.; Gronwald, J. W.; Wyse, D. L.
The chlorination of hydrolysis lignin
Summary 1.Technical hydrolysis lignin is easily chlorinated at 20� by solutions of chlorine in carbon tetrachloride or by chlorine water. When 25�27% of chlorine is introduced, the latter is quantitatively removed from the solution by the lignin.2.The chlorine content of products chlorinated in carbon tetrachloide is very close to that required by calculation for the substitution reaction.3.When lignin is chlorinated
N. N. Shorygina; L. I. Kolotova
Dilute acid hydrolysis of softwoods
Whole tree chips obtained from softwood forest thinnings were converted to ethanol via a two-stage dilute acid hydrolysis\\u000a followed by yeast fermentation. The chips were first impregnated with dilute sulfuric acid, then pretreated in a steam explosion\\u000a reactor to hydrolyze, more than 90% of the hemicellulose and approx 10% of the cellulose. The hydrolysate was filtered and\\u000a washed with water
Quang A. Nguyen; Melvin P. Tucker; Fred A. Keller; Delicia A. Beaty; Kevin M. Connors; Fannie P. Eddy
Crystal Structure of the N-Acetyltransferase Domain of Human N-Acetyl-L-Glutamate Synthase in Complex with N-Acetyl-L-Glutamate Provides Insights into Its Catalytic and Regulatory Mechanisms
N-acetylglutamate synthase (NAGS) catalyzes the conversion of AcCoA and L-glutamate to CoA and N-acetyl-L-glutamate (NAG), an obligate cofactor for carbamyl phosphate synthetase I (CPSI) in the urea cycle. NAGS deficiency results in elevated levels of plasma ammonia which is neurotoxic. We report herein the first crystal structure of human NAGS, that of the catalytic N-acetyltransferase (hNAT) domain with N-acetyl-L-glutamate bound at 2.1 � resolution. Functional studies indicate that the hNAT domain retains catalytic activity in the absence of the amino acid kinase (AAK) domain. Instead, the major functions of the AAK domain appear to be providing a binding site for the allosteric activator, L-arginine, and an N-terminal proline-rich motif that is likely to function in signal transduction to CPS1. Crystalline hNAT forms a dimer similar to the NAT-NAT dimers that form in crystals of bifunctional N-acetylglutamate synthase/kinase (NAGS/K) from Maricaulis maris and also exists as a dimer in solution. The structure of the NAG binding site, in combination with mutagenesis studies, provide insights into the catalytic mechanism. We also show that native NAGS from human and mouse exists in tetrameric form, similar to those of bifunctional NAGS/K.
Zhao, Gengxiang; Jin, Zhongmin; Allewell, Norma M.; Tuchman, Mendel; Shi, Dashuang
Heterologous expression and biochemical characterization of acetyl xylan esterase from Coprinopsis cinerea.
Acetyl xylan esterase (AXE) from basidiomycete Coprinopsis cinerea Okayama 7 (#130) was functionally expressed in Pichia pastoris with a C-terminal tag under the alcohol oxidase 1 (AOX1) promoter and secreted into the medium at 1.5 mg l(-1). Its molecular mass was estimated to be 65.5 kDa based on the SDS-PAGE analysis, which is higher than the calculated molecular mass of 40 kDa based on amino acid composition. In-silico analysis of the amino acid sequence predicted two potential N-glycosylation sites. Results from PNGase F deglycosylation and mass spectrum confirmed the presence of N-glycosylation on the recombinant AXE with predominant N-glycans HexNAc2Hex9-16. The recombinant AXE showed best activity at 40 �C and pH 8. It showed not only acetyl esterase activity with a Km of 4.3 mM and a Vmax of 2.15 U mg(-1) for hydrolysis of 4-nitrophenyl acetate but also a butyl esterase activity for hydrolysis of 4-nitrophenyl butyrate with a Km of 0.11 mM and Vmax of 0.78 U mg(-1). The presence of two additional amino acid residues at its native N-terminus was found to help stabilize the enzyme against the protease cleavages without affecting its activity. PMID:23180549
Juturu, Veeresh; Aust, Christina; Wu, Jin Chuan
Photosensitized [2 + 2] cycloaddition of N-acetylated cytosine affords stereoselective formation of cyclobutane pyrimidine dimer
Photocycloaddition between two adjacent bases in DNA produces a cyclobutane pyrimidine dimer (CPD), which is one of the major UV-induced DNA lesions, with either the cis-syn or trans-syn structure. In this study, we investigated the photosensitized intramolecular cycloaddition of partially-protected thymidylyl-(3??5?)-N4-acetyl-2?-deoxy-5-methylcytidine, to clarify the effect of the base modification on the cycloaddition reaction. The reaction resulted in the stereoselective formation of the trans-syn CPD, followed by hydrolysis of the acetylamino group. The same result was obtained for the photocycloaddition of thymidylyl-(3??5?)-N4-acetyl-2?-deoxycytidine, whereas both the cis-syn and trans-syn CPDs were formed from thymidylyl-(3??5?)-thymidine. Kinetic analyses revealed that the activation energy of the acid-catalyzed hydrolysis is comparable to that reported for the thymine-cytosine CPD. These findings provided a new strategy for the synthesis of oligonucleotides containing the trans-syn CPD. Using the synthesized oligonucleotide, translesion synthesis by human DNA polymerase ? was analyzed.
Yamamoto, Junpei; Nishiguchi, Kosuke; Manabe, Koichiro; Masutani, Chikahide; Hanaoka, Fumio; Iwai, Shigenori
Photosensitized [2 + 2] cycloaddition of N-acetylated cytosine affords stereoselective formation of cyclobutane pyrimidine dimer.
Photocycloaddition between two adjacent bases in DNA produces a cyclobutane pyrimidine dimer (CPD), which is one of the major UV-induced DNA lesions, with either the cis-syn or trans-syn structure. In this study, we investigated the photosensitized intramolecular cycloaddition of partially-protected thymidylyl-(3'?5')-N(4)-acetyl-2'-deoxy-5-methylcytidine, to clarify the effect of the base modification on the cycloaddition reaction. The reaction resulted in the stereoselective formation of the trans-syn CPD, followed by hydrolysis of the acetylamino group. The same result was obtained for the photocycloaddition of thymidylyl-(3'?5')-N(4)-acetyl-2'-deoxycytidine, whereas both the cis-syn and trans-syn CPDs were formed from thymidylyl-(3'?5')-thymidine. Kinetic analyses revealed that the activation energy of the acid-catalyzed hydrolysis is comparable to that reported for the thymine-cytosine CPD. These findings provided a new strategy for the synthesis of oligonucleotides containing the trans-syn CPD. Using the synthesized oligonucleotide, translesion synthesis by human DNA polymerase ? was analyzed. PMID:20880992
Yamamoto, Junpei; Nishiguchi, Kosuke; Manabe, Koichiro; Masutani, Chikahide; Hanaoka, Fumio; Iwai, Shigenori
Novel domain arrangement in the crystal structure of a truncated acetyl-CoA synthase fromMoorella thermoacetica��
Ni-dependent Acetyl-CoA synthase (ACS) and CO dehydrogenase (CODH) constitute the central enzyme complex of the Wood-Ljungdahl pathway of acetyl-CoA formation. The crystal structure of a recombinant bacterial ACS lacking the N-terminal domain that interacts with CODH shows a large reorganization of the remaining two globular domains, producing a narrow cleft of suitable size, shape and nature to bind CoA. Sequence comparisons with homologous archaeal enzymes that naturally lack the N-terminal domain show that many amino acids lining this cleft are conserved. Besides the typical [4Fe-4S] center, the A-cluster contains only one proximal metal ion that, according to anomalous scattering data, is most likely Cu or Zn. Incorporation of a functional Ni2Fe4S4 A-cluster would require only minor structural rearrangements. Using available structures, a plausible model of the interaction between CODH and the smaller ACS in archaeal multi-enzyme complexes is presented, along with a discussion of evolutionary relationships of the archaeal and bacterial enzymes.
Volbeda, Anne; Darnault, Claudine; Tan, Xiangshi; Lindahl, Paul A.; Fontecilla-Camps, Juan C.
Yeast phospholipase C is required for normal acetyl-CoA homeostasis and global histone acetylation.
Phospholipase C (Plc1p) is required for the initial step of inositol polyphosphate (InsP) synthesis, and yeast cells with deletion of the PLC1 gene are completely devoid of any InsPs and display aberrations in transcriptional regulation. Here we show that Plc1p is required for a normal level of histone acetylation; plc1? cells that do not synthesize any InsPs display decreased acetylation of bulk histones and global hypoacetylation of chromatin histones. In accordance with the role of Plc1p in supporting histone acetylation, plc1? mutation is synthetically lethal with mutations in several subunits of SAGA and NuA4 histone acetyltransferase (HAT) complexes. Conversely, the growth rate, sensitivity to multiple stresses, and the transcriptional defects of plc1? cells are partially suppressed by deletion of histone deacetylase HDA1. The histone hypoacetylation in plc1? cells is due to the defect in degradation of repressor Mth1p, and consequently lower expression of HXT genes and reduced conversion of glucose to acetyl-CoA, a substrate for HATs. The histone acetylation and transcriptional defects can be partially suppressed and the overall fitness improved in plc1? cells by increasing the cellular concentration of acetyl-CoA. Together, our data indicate that Plc1p and InsPs are required for normal acetyl-CoA homeostasis, which, in turn, regulates global histone acetylation. PMID:23913687
Galdieri, Luciano; Chang, Jennifer; Mehrotra, Swati; Vancura, Ales
The fluvial Garber Sandstone and the underlying Wellington Formation are important sources of drinking water in central Oklahoma. These formations, which make up much of the COA, consist of amalgamated sandstones with some interbedded mudstones, siltstones, and local mudstone- a...
Structural, Kinetic and Proteomic Characterization of Acetyl Phosphate-Dependent Bacterial Protein Acetylation
The emerging view of N?-lysine acetylation in eukaryotes is of a relatively abundant post-translational modification (PTM) that has a major impact on the function, structure, stability and/or location of thousands of proteins involved in diverse cellular processes. This PTM is typically considered to arise by the donation of the acetyl group from acetyl-coenzyme A (acCoA) to the ?-amino group of a lysine residue that is reversibly catalyzed by lysine acetyltransferases and deacetylases. Here, we provide genetic, mass spectrometric, biochemical and structural evidence that N?-lysine acetylation is an equally abundant and important PTM in bacteria. Applying a recently developed, label-free and global mass spectrometric approach to an isogenic set of mutants, we detected acetylation of thousands of lysine residues on hundreds of Escherichia coli proteins that participate in diverse and often essential cellular processes, including translation, transcription and central metabolism. Many of these acetylations were regulated in an acetyl phosphate (acP)-dependent manner, providing compelling evidence for a recently reported mechanism of bacterial N?-lysine acetylation. These mass spectrometric data, coupled with observations made by crystallography, biochemistry, and additional mass spectrometry showed that this acP-dependent acetylation is both non-enzymatic and specific, with specificity determined by the accessibility, reactivity and three-dimensional microenvironment of the target lysine. Crystallographic evidence shows acP can bind to proteins in active sites and cofactor binding sites, but also potentially anywhere molecules with a phosphate moiety could bind. Finally, we provide evidence that acP-dependent acetylation can impact the function of critical enzymes, including glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase, and RNA polymerase.
Sahu, Alexandria; Sorensen, Dylan; Minasov, George; Lima, Bruno P.; Scholle, Michael; Mrksich, Milan; Anderson, Wayne F.; Gibson, Bradford W.; Schilling, Birgit; Wolfe, Alan J.
Beating the acetyl coenzyme A-pathway to the origin of life
Attempts to draft plausible scenarios for the origin of life have in the past mainly built upon palaeogeochemical boundary conditions while, as detailed in a companion article in this issue, frequently neglecting to comply with fundamental thermodynamic laws. Even if demands from both palaeogeochemistry and thermodynamics are respected, then a plethora of strongly differing models are still conceivable. Although we have no guarantee that life at its origin necessarily resembled biology in extant organisms, we consider that the only empirical way to deduce how life may have emerged is by taking the stance of assuming continuity of biology from its inception to the present day. Building upon this conviction, we have assessed extant types of energy and carbon metabolism for their appropriateness to conditions probably pertaining in those settings of the Hadean planet that fulfil the thermodynamic requirements for life to come into being. Wood�Ljungdahl (WL) pathways leading to acetyl CoA formation are excellent candidates for such primordial metabolism. Based on a review of our present understanding of the biochemistry and biophysics of acetogenic, methanogenic and methanotrophic pathways and on a phylogenetic analysis of involved enzymes, we propose that a variant of modern methanotrophy is more likely than traditional WL systems to date back to the origin of life. The proposed model furthermore better fits basic thermodynamic demands and palaeogeochemical conditions suggested by recent results from extant alkaline hydrothermal seeps.
Nitschke, Wolfgang; Russell, Michael J.
Beating the acetyl coenzyme A-pathway to the origin of life.
Attempts to draft plausible scenarios for the origin of life have in the past mainly built upon palaeogeochemical boundary conditions while, as detailed in a companion article in this issue, frequently neglecting to comply with fundamental thermodynamic laws. Even if demands from both palaeogeochemistry and thermodynamics are respected, then a plethora of strongly differing models are still conceivable. Although we have no guarantee that life at its origin necessarily resembled biology in extant organisms, we consider that the only empirical way to deduce how life may have emerged is by taking the stance of assuming continuity of biology from its inception to the present day. Building upon this conviction, we have assessed extant types of energy and carbon metabolism for their appropriateness to conditions probably pertaining in those settings of the Hadean planet that fulfil the thermodynamic requirements for life to come into being. Wood-Ljungdahl (WL) pathways leading to acetyl CoA formation are excellent candidates for such primordial metabolism. Based on a review of our present understanding of the biochemistry and biophysics of acetogenic, methanogenic and methanotrophic pathways and on a phylogenetic analysis of involved enzymes, we propose that a variant of modern methanotrophy is more likely than traditional WL systems to date back to the origin of life. The proposed model furthermore better fits basic thermodynamic demands and palaeogeochemical conditions suggested by recent results from extant alkaline hydrothermal seeps. PMID:23754811
Nitschke, Wolfgang; Russell, Michael J
Differential Properties of 4-Coumarate: CoA Ligase Related to Growth Suppression by Chalcone in Maize and Rice
Lignin and related metabolites have diverse and important functions for plant growth and development. 4-Coumarate: CoA ligase (4CL, EC 184.108.40.206) is one of the key enzymes in phenylpropanoid metabolism and lignin biosynthesis. In a previous study, maize (Zea maize L. cv. Yellowcorn) growth was suppressed to a greater extent by root-applied chalcone than rice (Oryza sativa L. cv. Nipponbare). The
Min-Soo Yun; Weijun Chen; Fan Deng; Yasuhiro Yogo
Pyruvate Dehydrogenase Activity and Malonyl CoA Levels in Normal and Ischemic Swine Myocardium: Effects of Dichloroacetate
The pruposes of this study were to: (1) assess myocardial pyruvate dehydrogenase (PDH) activity and substrate exchange under well-perfused and eschemic conditions; (2) determine the metabolic effects of an intra-coronary infusion of the PDH activator, dichloroacetate (DCA); and (3) measure the effects of ischemia and DCA on malonyl CoA levels. Experiments were performed in anesthetised open-chest swine under non-ischemic conditions,
William C. Stanley; Lisa A. Hernandez; Doreen Spires; John Bringas; Sonja Wallace; James G. McCormack
Identification and cloning of two forms of liver peroxisomal fatty Acyl CoA Oxidase from the koala ( Phascolarctos cinereus)
In the present study, the cloning, expression and characterization of the rate-limiting enzyme of the peroxisomal ?-oxidation spiral, acyl CoA oxidase (AOX), from koala (Phascolarctos cinereus) liver is described. It has been previously reported that peroxisomal cyanide-insensitive palmitoyl-CoA oxidation activity was absent in koala liver [Comp. Biochem. Physiol. (C) 127 (2000) 327]. This activity is a measure of the overall
Suong Ngoc Thi Ngo; Ross Allan McKinnon; Ieva Stupans
Acetyl-L-carnitine in hepatic encephalopathy.
Hepatic encephalopathy is a common complication of hepatic cirrhosis. The clinical diagnosis is based on two concurrent types of symptoms: impaired mental status and impaired neuromotor function. Impaired mental status is characterized by deterioration in mental status with psychomotor dysfunction, impaired memory, and increased reaction time, sensory abnormalities, poor concentration, disorientation and coma. Impaired neuromotor function include hyperreflexia, rigidity, myoclonus and asterixis. The pathogenesis of hepatic encephalopathy has not been clearly defined. The general consensus is that elevated levels of ammonia and an inflammatory response work in synergy to cause astrocyte to swell and fluid to accumulate in the brain which is thought to explain the symptoms of hepatic encephalopathy. Acetyl-L-carnitine, the short-chain ester of carnitine is endogenously produced within mitochondria and peroxisomes and is involved in the transport of acetyl-moieties across the membranes of these organelles. Acetyl-L-carnitine administration has shown the recovery of neuropsychological activities related to attention/concentration, visual scanning and tracking, psychomotor speed and mental flexibility, language short-term memory, attention, and computing ability. In fact, Acetyl-L-carnitine induces ureagenesis leading to decreased blood and brain ammonia levels. Acetyl-L-carnitine treatment decreases the severity of mental and physical fatigue, depression cognitive impairment and improves health-related quality of life. The aim of this review was to provide an explanation on the possible toxic effects of ammonia in HE and evaluate the potential clinical benefits of ALC. PMID:23389620
Malaguarnera, Michele
Contribution of CoA ligases to benzenoid biosynthesis in petunia flowers.
Biosynthesis of benzoic acid from Phe requires shortening of the side chain by two carbons, which can occur via the ?-oxidative or nonoxidative pathways. The first step in the ?-oxidative pathway is cinnamoyl-CoA formation, likely catalyzed by a member of the 4-coumarate:CoA ligase (4CL) family that converts a range of trans-cinnamic acid derivatives into the corresponding CoA thioesters. Using a functional genomics approach, we identified two potential CoA-ligases from petunia (Petunia hybrida) petal-specific cDNA libraries. The cognate proteins share only 25% amino acid identity and are highly expressed in petunia corollas. Biochemical characterization of the recombinant proteins revealed that one of these proteins (Ph-4CL1) has broad substrate specificity and represents a bona fide 4CL, whereas the other is a cinnamate:CoA ligase (Ph-CNL). RNA interference suppression of Ph-4CL1 did not affect the petunia benzenoid scent profile, whereas downregulation of Ph-CNL resulted in a decrease in emission of benzylbenzoate, phenylethylbenzoate, and methylbenzoate. Green fluorescent protein localization studies revealed that the Ph-4CL1 protein is localized in the cytosol, whereas Ph-CNL is in peroxisomes. Our results indicate that subcellular compartmentalization of enzymes affects their involvement in the benzenoid network and provide evidence that cinnamoyl-CoA formation by Ph-CNL in the peroxisomes is the committed step in the ?-oxidative pathway. PMID:22649270
Klempien, Antje; Kaminaga, Yasuhisa; Qualley, Anthony; Nagegowda, Dinesh A; Widhalm, Joshua R; Orlova, Irina; Shasany, Ajit Kumar; Taguchi, Goro; Kish, Christine M; Cooper, Bruce R; D'Auria, John C; Rhodes, David; Pichersky, Eran; Dudareva, Natalia
Acyl CoA profiles of transgenic plants that accumulate medium-chain fatty acids indicate inefficient storage lipid synthesis in developing oilseeds.
Several Brassica napus lines transformed with genes responsible for the synthesis of medium- or long-chain fatty acids were examined to determine limiting factor(s) for the subsequent accumulation of these fatty acids in seed lipids. Examination of a decanoic acid (10:0) accumulating line revealed a disproportionately high concentration of 10:0 CoA during seed development compared to long-chain acyl CoAs isolated from the same tissues, suggesting that poor incorporation of 10:0 CoA into seed lipids limits 10:0 fatty acid accumulation. This relationship was also seen for dodecanoyl (12:0) CoA and fatty acid in a high 12:0 line, but not for octadecanoic (18:0) CoA and fatty acid in a high 18:0 line. Comparison of 10:0 CoA and fatty acid proportions from seeds at different developmental stages for transgenic B. napus and Cuphea hookeriana, the source plant for the medium-chain thioesterase and 3-ketoacyl-ACP synthase transgenes, revealed that C. hookeriana incorporates 10:0 CoA into seed lipids more efficiently than transgenic B. napus. Furthermore, beta-oxidation and glyoxylate cycle activities were not increased above wild type levels during seed development in the 8:0/10:0 line, suggesting that lipid catabolism was not being induced in response to the elevated 10:0 CoA concentrations. Taken together, these data suggest that transgenic plants that are engineered to synthesize medium-chain fatty acids may lack the necessary mechanisms, such as specific acyltransferases, to incorporate these fatty acids efficiently into seed lipids. PMID:12445123
Larson, Tony R; Edgell, Teresa; Byrne, James; Dehesh, Katayoon; Graham, Ian A
Identification and characterisation of the alpha and beta subunits of succinyl CoA ligase of tomato.
Despite the central importance of the TCA cycle in plant metabolism not all of the genes encoding its constituent enzymes have been functionally identified. In yeast, the heterodimeric protein succinyl CoA ligase is encoded for by two single-copy genes. Here we report the isolation of two tomato cDNAs coding for alpha- and one coding for the beta-subunit of succinyl CoA ligase. These three cDNAs were used to complement the respective Saccharomyces cerevisiae mutants deficient in the alpha- and beta-subunit, demonstrating that they encode functionally active polypeptides. The genes encoding for the subunits were expressed in all tissues, but most strongly in floral and leaf tissues, with equivalent expression of the two alpha-subunit genes being expressed to equivalent levels in all tissues. In all instances GFP fusion expression studies confirmed an expected mitochondrial location of the proteins encoded. Following the development of a novel assay to measure succinyl CoA ligase activity, in the direction of succinate formation, the evaluation of the maximal catalytic activities of the enzyme in a range of tissues revealed that these paralleled those of mRNA levels. We also utilized this assay to perform a preliminary characterisation of the regulatory properties of the enzyme suggesting allosteric control of this enzyme which may regulate flux through the TCA cycle in a manner consistent with its position therein. PMID:16270230
Studart-Guimar�es, Claudia; Gibon, Yves; Frankel, Nicol�s; Wood, Craig C; Zanor, Mar�a In�s; Fernie, Alisdair R; Carrari, Fernando
High consistency enzymatic hydrolysis of hardwood substrates.
The feasibility of using a laboratory peg mixer to carry out high consistency enzymatic hydrolysis of lignocellulosic substrates was investigated. Two hardwood substrates, unbleached hardwood pulp (UBHW) and organosolv pretreated poplar (OPP), were used in this study. Hydrolysis of UBHW and OPP at 20% substrate consistency led to a high glucose concentration in the final hydrolysate. For example, a 48 h enzymatic hydrolysis of OPP resulted in a hydrolysate with 158 g/L of glucose. This is the highest glucose concentration ever obtained from enzymatic hydrolysis of lignocellulosic substrates. Fermentation of UBHW and OPP hydrolysates with high glucose content led to high ethanol concentrations, 50.4 and 63.1 g/L, respectively after fermentation. Our results demonstrate that using common pulping equipment to carry out high consistency hydrolysis can overcome the rheological problems and greatly increase the sugar and ethanol concentrations after the hydrolysis and fermentation. PMID:19643602
Zhang, Xiao; Qin, Wenjuan; Paice, Michael G; Saddler, John N
??Acetyl?N?heterocycles in the Maillard reaction
??Acetyl?N?heterocycles are a special class of Maillard reaction products with typical roasty flavor characteristic and generally low odor thresholds. A total of 12 ??acetyl?N?heterocycles have so far been reported to occur in heated foodstuffs or process flavorings. The partially unsaturated ??acetyl?N?heterocycles 2?acetyltetrahydropyri?dine, 2?acetyl?1?pyrroline, 2?acetyl?2?thiazoline, and 5?acetyl?2,3?di?hydro?1,4?thiazine are reported to have very low odor thresholds (in water) between 0.1 and 1.6
Josef Kerler; Jos G. M. van der Ven; Hugo Weenen
Enhancement of lysine acetylation accelerates wound repair.
In physiopathological conditions, such as diabetes, wound healing is significantly compromised and chronic complications, including ulcers, may occur. In a mouse model of skin repair, we recently reported that wound treatment with Sirtuin activators and class I HDAC inhibitors induced keratinocyte proliferation and enhanced healing via a nitric oxide (NO) dependent mechanism. We observed an increase in total protein acetylation in the wound area, as determined by acetylation of ?-tubulin and histone H3 Lysine 9. We reasoned that this process activated cell function as well as regulated gene expression to foster tissue repair. We report here that the direct activation of P300/CBP-associated factor (PCAF) by the histone acetylase activator pentadecylidenemalonate 1b (SPV-106) induced Lysine acetylation in the wound area. This intervention was sufficient to enhance repair process by a NO-independent mechanism. Hence, an impairment of PCAF and/or other GCN5 family acetylases may delay skin repair in physiopathological conditions. PMID:24265859
Spallotta, Francesco; Cencioni, Chiara; Straino, Stefania; Sbardella, Gianluca; Castellano, Sabrina; Capogrossi, Maurizio C; Martelli, Fabio; Gaetano, Carlo
Enhancement of lysine acetylation accelerates wound repair
In physiopathological conditions, such as diabetes, wound healing is significantly compromised and chronic complications, including ulcers, may occur. In a mouse model of skin repair, we recently reported that wound treatment with Sirtuin activators and class I HDAC inhibitors induced keratinocyte proliferation and enhanced healing via a nitric oxide (NO) dependent mechanism. We observed an increase in total protein acetylation in the wound area, as determined by acetylation of ?-tubulin and histone H3 Lysine 9. We reasoned that this process activated cell function as well as regulated gene expression to foster tissue repair. We report here that the direct activation of P300/CBP-associated factor (PCAF) by the histone acetylase activator pentadecylidenemalonate 1b (SPV-106) induced Lysine acetylation in the wound area. This intervention was sufficient to enhance repair process by a NO-independent mechanism. Hence, an impairment of PCAF and/or other GCN5 family acetylases may delay skin repair in physiopathological conditions.
Spallotta, Francesco; Cencioni, Chiara; Straino, Stefania; Sbardella, Gianluca; Castellano, Sabrina; Capogrossi, Maurizio C; Martelli, Fabio; Gaetano, Carlo
Validation of Protein Acetylation by Mass Spectrometry
Summary Due to the key role of post-translational modifications (PTMs) such as lysine acetylation in numerous signaling and regulatory pathways, the ability to identify novel PTMs and to quantify their abundances provides invaluable information for understanding these signaling networks. Currently, mass spectrometry (MS) arguably serves as the most high-throughput and unbiased platform for studying post-translational modifications. Here we detail experimental and analytical procedures for the characterization of lysine acetylation on proteins in general and on histones in particular, which are among the most highly modified proteins in eukaryotic cells.
Zee, Barry M.; Garcia, Benjamin A.
The hydrolysis of bile acid conjugates.
Studies were made of a) the relationship of bile acid structure and analytical recoveries (measured by 3-hydroxysteroid oxidoreductase) following vigorous alkaline hydrolysis of bile acid conjugates and b) the relationship of structure and hydrolysis time of taurine- and glycine bile acid conjugates in a reaction catalyzed by glycocholic acid hydrolase. Alkaline hydrolysis resulted in good recoveries of hydroxy and 7 and 12- oxo-bile acids but poor recoveries of 3-oxo-bile acids. Borohydride reduction of the 3-oxo-acids prevented these losses. Complete enzymatic hydrolysis of glycine conjugated bile acids was about five times more rapid than that of taurine conjugates. Hydrolysis of conjugates containing oxo groups was slow. Borohydride reduction of oxo-acids corrected this and did not inhibit enzymatic hydrolysis. It was concluded that both vigorous alkaline and enzymatic hydrolysis are satisfactory in bile acid assays if borohydride reduction is instituted before the hydrolytic step. However, due to the presence of possible enzyme inhibitors and solubility difficulties, strong alkaline hydrolysis is preferable to enzymatic hydrolysis in fecal bile acid determinations at this time. PMID:715825
Beher, W T; Stradnieks, S; Beher, G R; Lin, G J
Acid Hydrolysis of Trioxalatocobaltate (III) Ion
ERIC Educational Resources Information Center
Describes an investigation involving acid hydrolysis and using both volumetric and kinetic techniques. Presents examples of the determination of the rate constant and its variation with temperature. (GS)
Wiggans, P. W.
New improved synthesis of 2-deoxy-2-(/sup 18/F)fluoro-d-glucose from /sup 18/F-labeled acetyl hypofluorite
A new improved synthesis of 2-deoxy-2-(/sup 18/F)fluoro-D-glucose (2-/sup 18/FDG, 4) from /sup 18/F-labeled acetyl hypofluorite (1) is described. Reaction of 3,4,6-tri-O-acetyl-D-glucal (2) with 1, generated in situ from (/sup 18/F)F/sub 2/ and ammonium acetate in acetic acid solution at room temperature, gave 2-deoxy-2-(/sup 18/F)fluoro-1,3,4,6-tetra-O-acetyl-..cap alpha..-D-glucopyranose (3). Hydrolysis of compound 3 with 2N HCl gave 2-deoxy-2-(/sup 18/F)fluoro-D-glucose (4) in radiochemical yields of approx.20% in a synthesis time of 70 min from EOB.
Shiue, C.Y. (Brookhaven National Lab., Upton, NY); Salvadori, P.A.; Wolf, A.P.; Fowler, J.S.; MacGregor, R.R.
4-coumarate: CoA ligase partitions metabolites for eugenol biosynthesis.
Biosynthesis of eugenol shares its initial steps with that of lignin, involving conversion of hydroxycinnamic acids to their corresponding coenzyme A (CoA) esters by 4-coumarate:CoA ligases (4CLs). In this investigation, a 4CL (OS4CL) was identified from glandular trichome-rich tissue of Ocimum sanctum with high sequence similarity to an isoform (OB4CL_ctg4) from Ocimum basilicum. The levels of OS4CL and OB4CL_ctg4-like transcripts were highest in O. sanctum trichome, followed by leaf, stem and root. The eugenol content in leaf essential oil was positively correlated with the expression of OS4CL in the leaf at different developmental stages. Recombinant OS4CL showed the highest activity with p-coumaric acid, followed by ferulic, caffeic and trans-cinnamic acids. Transient RNA interference (RNAi) suppression of OS4CL in O. sanctum leaves caused a reduction in leaf eugenol content and trichome transcript level, with a considerable increase in endogenous p-coumaric, ferulic, trans-cinnamic and caffeic acids. A significant reduction in the expression levels was observed for OB4CL_ctg4-related transcripts in suppressed trichome compared with transcripts similar to the other four isoforms (OB4CL_ctg1, 2, 3 and 5). Sinapic acid and lignin content were also unaffected in RNAi suppressed leaf samples. Transient expression of OS4CL-green fluorescent protein fusion protein in Arabidopsis protoplasts was associated with the cytosol. These results indicate metabolite channeling of intermediates towards eugenol by a specific 4CL and is the first report demonstrating the involvement of 4CL in creation of virtual compartments through substrate utilization and committing metabolites for eugenol biosynthesis at an early stage of the pathway. PMID:23677922
Rastogi, Shubhra; Kumar, Ritesh; Chanotiya, Chandan S; Shanker, Karuna; Gupta, Madan M; Nagegowda, Dinesh A; Shasany, Ajit K
Intracellular Long-Chain Acyl CoAs Activate TRPV1 Channels.
TRPV1 channels are an important class of membrane proteins that play an integral role in the regulation of intracellular cations such as calcium in many different tissue types. The anionic phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) is a known positive modulator of TRPV1 channels and the negatively charged phosphate groups interact with several basic amino acid residues in the proximal C-terminal TRP domain of the TRPV1 channel. We and other groups have shown that physiological sub-micromolar levels of long-chain acyl CoAs (LC-CoAs), another ubiquitous anionic lipid, can also act as positive modulators of ion channels and exchangers. Therefore, we investigated whether TRPV1 channel activity is similarly regulated by LC-CoAs. Our results show that LC-CoAs are potent activators of the TRPV1 channel and interact with the same PIP2-binding residues in TRPV1. In contrast to PIP2, LC-CoA modulation of TRPV1 is independent of Ca2+i, acting in an acyl side-chain saturation and chain-length dependent manner. Elevation of LC-CoAs in intact Jurkat T-cells leads to significant increases in agonist-induced Ca2+i levels. Our novel findings indicate that LC-CoAs represent a new fundamental mechanism for regulation of TRPV1 channel activity that may play a role in diverse cell types under physiological and pathophysiological conditions that alter fatty acid transport and metabolism such as obesity and diabetes. PMID:24798548
Yu, Yi; Carter, Chris R J; Youssef, Nermeen; Dyck, Jason R B; Light, Peter E
FGFR3 stimulates stearoyl CoA desaturase 1 activity to promote bladder tumor growth.
Fibroblast growth factor receptor 3 (FGFR3) belongs to a family of receptor tyrosine kinases that control cell proliferation, differentiation, and survival. Aberrant activation of FGFR3 via overexpression or mutation is a frequent feature of bladder cancer; however, its molecular and cellular consequences and functional relevance to carcinogenesis are not well understood. Through transcriptional profiling of bladder carcinoma cells subjected to short hairpin RNA knockdown of FGFR3, we identified a gene-signature linking FGFR3 signaling with de novo sterol and lipid biosynthesis and metabolism. We found that FGFR3 signaling promotes the cleavage and activation of the master transcriptional regulator of lipogenesis, sterol regulatory element-binding protein 1(SREBP1/SREBF1), in a PI3K-mTORC1-dependent fashion. In turn, SREBP1 regulates the expression of key lipogenic enzymes, including stearoyl CoA desaturase 1 (SCD1/SCD). SCD1 is the rate-limiting enzyme in the biosynthesis of monounsaturated fatty acids and is crucial for lipid homeostasis. In human bladder cancer cell lines expressing constitutively active FGFR3, knockdown of SCD1 by siRNA markedly attenuated cell-cycle progression, reduced proliferation, and induced apoptosis. Furthermore, inducible knockdown of SCD1 in a bladder cancer xenograft model substantially inhibited tumor progression. Pharmacologic inhibition of SCD1 blocked fatty acid desaturation and also exerted antitumor activity in vitro and in vivo. Together, these findings reveal a previously unrecognized role of FGFR3 in regulating lipid metabolism to maintain tumor growth and survival, and also identify SCD1 as a potential therapeutic target for FGFR3-driven bladder cancer. PMID:23019225
Du, Xiangnan; Wang, Qian-Rena; Chan, Emily; Merchant, Mark; Liu, Jinfeng; French, Dorothy; Ashkenazi, Avi; Qing, Jing
Intracellular Long-Chain Acyl CoAs Activate TRPV1 Channels
TRPV1 channels are an important class of membrane proteins that play an integral role in the regulation of intracellular cations such as calcium in many different tissue types. The anionic phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) is a known positive modulator of TRPV1 channels and the negatively charged phosphate groups interact with several basic amino acid residues in the proximal C-terminal TRP domain of the TRPV1 channel. We and other groups have shown that physiological sub-micromolar levels of long-chain acyl CoAs (LC-CoAs), another ubiquitous anionic lipid, can also act as positive modulators of ion channels and exchangers. Therefore, we investigated whether TRPV1 channel activity is similarly regulated by LC-CoAs. Our results show that LC-CoAs are potent activators of the TRPV1 channel and interact with the same PIP2-binding residues in TRPV1. In contrast to PIP2, LC-CoA modulation of TRPV1 is independent of Ca2+i, acting in an acyl side-chain saturation and chain-length dependent manner. Elevation of LC-CoAs in intact Jurkat T-cells leads to significant increases in agonist-induced Ca2+i levels. Our novel findings indicate that LC-CoAs represent a new fundamental mechanism for regulation of TRPV1 channel activity that may play a role in diverse cell types under physiological and pathophysiological conditions that alter fatty acid transport and metabolism such as obesity and diabetes.
Youssef, Nermeen; Dyck, Jason R. B.; Light, Peter E.
Gene encoding acetyl-coenzyme A carboxylase
A DNA encoding an acetyl-coenzyme A carboxylase (ACCase) from a photosynthetic organism and functional derivatives are disclosed which are resistant to inhibition from certain herbicides. This gene can be placed in organisms to increase their fatty acid content or to render them resistant to certain herbicides. 5 figs.
Roessler, P.G.; Ohlrogge, J.B.
Gene encoding acetyl-coenzyme A carboxylase
A DNA encoding an acetyl-coenzyme A carboxylase (ACCase) from a photosynthetic organism and functional derivatives thereof which are resistant to inhibition from certain herbicides. This gene can be placed in organisms to increase their fatty acid content or to render them resistant to certain herbicides.
Roessler, Paul G. (Golden, CO); Ohlrogge, John B. (Okemos, MI)
A slow dance for microtubule acetylation.
Microtubules contribute to diverse cellular processes through balancing dynamic, short-lived and stable, long-lived populations. One way in which long-lived microtubules are marked is by posttranslational acetylation of ?-tubulin by tubulin acetyltransferase (TAT). Szyk et�al. now provide insight into TAT's mechanism of action and its unique time-stamping ability. PMID:24906144
Kull, F Jon; Sloboda, Roger D
Evidence for a blockwise distribution of acetyl groups onto homogalacturonans from a commercial sugar beet (Beta vulgaris) pectin.
Commercial acid-extracted sugar beet pectin was extensively hydrolysed using an endo-polygalacturonase (AnPGI from Aspergillus niger or AnPGII from A. niger or FmPG from Fusarium moniliforme) in combination with Aspergillus aculeatus pectin methyl-esterase (AaPME). The homogalacturonan-derived oligogalacturonates released were quantified by high-performance anion-exchange chromatography and their structure determined by mass spectrometry. The different endo-polygalacturonases exhibited variable tolerance towards acetyl groups. AnPGI was the most active and FmPG the less. A hypothetical homogalacturonan was constructed using the AnPGI-recovered oligogalacturonates as building blocks and the validity of the model was checked taking into account FmPG observed requirements and hydrolysis products. A blockwise repartition of the acetyl groups onto sugar beet pectin homogalacturonan is proposed. PMID:18448141
Ralet, Marie-Christine; Cr�peau, Marie-Jeanne; Bonnin, Estelle
Fast and single solid phase fluorescence spectroscopic batch procedure for (acetyl) salicylic acid determination in drug formulations.
A solid phase fluorescence spectroscopic batch procedure for (acetyl) salicylic acid in drug formulations have been developed. The procedure is based on the sorption of salicylic acid (SA) on Sephadex DEAE A-25 anion exchanger gel (100 mg) by equilibration from an aqueous solution (10 or 25 ml) for 5 min; the equilibrated gel is transferred into an 1 mm quartz cell and the native fluorescence of SA sorbed on it is directly measured (lambda(ex)=297 nm; lambda(em)=405 nm). Good linearity was found in the 10-200 and 5-100 microg l(-1) ranges (for 10 and 25 ml sample volume, respectively) with R.S.D. (%) of 2.8 and 1.1. The procedure was successfully applied to the determination of acetyl salicylic acid (ASA) in drug formulations after alkaline hydrolysis to yield SA. PMID:12615230
Ortega Algar, S; Ramos Martos, N; Molina D�az, A
Property enhancement of optically transparent bionanofiber composites by acetylation
NASA Astrophysics Data System (ADS)
The authors studied acetylation of bacterial cellulose (BC) nanofibers to widen the applications of BC nanocomposites in optoelectronic devices. The slight acetylation of BC nanofibers significantly reduces the hygroscopicity of BC nanocomposites, while maintaining their high optical transparency and thermal stability. Furthermore, the degradation in optical transparency at elevated temperature (200 �C) was significantly reduced by acetylation treatment. Therefore, the acetylation of bionanofibers has an extraordinary potential as treatment for property enhancement of bionanofiber composites.
Nogi, Masaya; Abe, Kentaro; Handa, Keishin; Nakatsubo, Fumiaki; Ifuku, Shinsuke; Yano, Hiroyuki
The Crystal Structure of N-Acetyl-L-glutamate Synthase from Neisseria gonorrhoeae Provides Insights into Mechanisms of Catalysis and Regulation
The crystal structures of N-acetylglutamate synthase (NAGS) in the arginine biosynthetic pathway of Neisseria gonorrhoeae complexed with acetyl-CoA and with CoA plus N-acetylglutamate have been determined at 2.5- and 2.6-A resolution, respectively. The monomer consists of two separately folded domains, an amino acid kinase (AAK) domain and an N-acetyltransferase (NAT) domain connected through a 10-A linker. The monomers assemble into a hexameric ring that consists of a trimer of dimers with 32-point symmetry, inner and outer ring diameters of 20 and 100A, respectively, and a height of 110A(.) Each AAK domain interacts with the cognate domains of two adjacent monomers across two 2-fold symmetry axes and with the NAT domain from a second monomer of the adjacent dimer in the ring. The catalytic sites are located within the NAT domains. Three active site residues, Arg316, Arg425, and Ser427, anchor N-acetylglutamate in a position at the active site to form hydrogen bond interactions to the main chain nitrogen atoms of Cys356 and Leu314, and hydrophobic interactions to the side chains of Leu313 and Leu314. The mode of binding of acetyl-CoA and CoA is similar to other NAT family proteins. The AAK domain, although catalytically inactive, appears to bind arginine. This is the first reported crystal structure of any NAGS, and it provides insights into the catalytic function and arginine regulation of NAGS enzymes.
Shi, Dashuang; Sagar, Vatsala; Jin, Zhongmin; Yu, Xiaolin; Caldovic, Ljubica; Morizono, Hiroki; Allewell, Norma M.; Tuchman, Mendel (Maryland); (GWU); (Georgia)
The Crystal Structure of N-Acetyl-l-glutamate Synthase from Neisseria gonorrhoeae Provides Insights into Mechanisms of Catalysis and Regulation*�
The crystal structures of N-acetylglutamate synthase (NAGS) in the arginine biosynthetic pathway of Neisseria gonorrhoeae complexed with acetyl-CoA and with CoA plus N-acetylgluta-mate have been determined at 2.5- and 2.6-� resolution, respectively. The monomer consists of two separately folded domains, an amino acid kinase (AAK) domain and an N-acetyltransferase (NAT) domain connected through a 10-� linker. The monomers assemble into a hexameric ring that consists of a trimer of dimers with 32-point symmetry, inner and outer ring diameters of 20 and 100�, respectively, and a height of 110�. Each AAK domain interacts with the cognate domains of two adjacent monomers across two 2-fold symmetry axes and with the NAT domain from a second monomer of the adjacent dimer in the ring. The catalytic sites are located within the NAT domains. Three active site residues, Arg316, Arg425, and Ser427, anchor N-acetylglutamate in a position at the active site to form hydrogen bond interactions to the main chain nitrogen atoms of Cys356 and Leu314, and hydrophobic interactions to the side chains of Leu313 and Leu314. The mode of binding of acetyl-CoA and CoA is similar to other NAT family proteins. The AAK domain, although catalytically inactive, appears to bind argi-nine. This is the first reported crystal structure of any NAGS, and it provides insights into the catalytic function and arginine regulation of NAGS enzymes.
Shi, Dashuang; Sagar, Vatsala; Jin, Zhongmin; Yu, Xiaolin; Caldovic, Ljubica; Morizono, Hiroki; Allewell, Norma M.; Tuchman, and Mendel
A Secondary Isotope Effect Study of Equine Serum Butyrylcholinesterase-Catalyzed Hydrolysis of Acetylthiocholine
?-Secondary deuterium isotope effects have been measured for equine serum butyrylcholinesterase-catalyzed hydrolysis of acetyl-L3-thiocholine (L = H or 2H). The dependencies of initial rates on isotopic substrate concentrations show close adherence to Michaelis-Menten kinetics, and yield the following isotope effects: D3kcat/Km = 0.98 � 0.02 and D3kcat = 1.10 � 0.02. The modestly inverse isotope effect on kcat/Km is consistent with partial rate limitation by a step that converts the sp2-hybridized ester carbonyl of the E + A reactant state into a quasi-tetrahedral transition state in the acylation stage of catalysis. On the other hand, the markedly normal isotope effect on kcat indicates that the Michaelis complex that accumulates at substrate saturation of the active site during catalytic turnover is a tetrahedral intermediate, whose decomposition is the rate-limiting step. These results compliment a previous report [J. R. Tormos et al., J. Am. Chem. Soc. 127 (2005) 14538�14539] that showed that substrate-activated hydrolysis of acetylthiocholine, catalyzed by recombinant human butyrylcholinesterase, is also rate limited by decomposition of an accumulating tetrahedral intermediate.
Wiley, Kenneth L.; Tormos, Jose R.; Quinn, Daniel M.
Microwave Pretreatment For Hydrolysis Of Cellulose
NASA Technical Reports Server (NTRS)
Microwave pretreatment enhances enzymatic hydrolysis of cellulosic wastes into soluble saccharides used as feedstocks for foods, fuels, and other products. Low consumption of energy, high yield, and low risk of proposed hydrolysis process incorporating microwave pretreatment makes process viable alternative to composting.
Cullingford, Hatice S.; George, Clifford E.; Lightsey, George R.
Cotton cellulose: enzyme adsorption and enzymic hydrolysis
The adsorption of a crude cellulase complex from Trichoderma viride on variously pretreated cotton cellulose samples was studied in the framework of the Langmuir approach at 2-8 degrees. The saturation amount of adsorbed enzyme was related to the susceptibility of the substrates to hydrolysis. In every case the adsorption process was faster by 2-3 orders of magnitude than the hydrolysis
P. L. Beltrame; P. Carniti; B. Focher; A. Marzetti; M. Cattaneo
Progress towards synthetic enzymes for phosphoester hydrolysis
Synthesis of artificial enzymes for catalyzing phosphoester hydrolysis has been attracting interest for a long time. The remarkable discovery that lanthanide ions catalyze the hydrolysis of DNA and RNA spurred the trend. Currently, progress is being made, mainly in the preparation of homogeneous catalysts, the promotion of catalytic activity by using acid\\/base cooperation within catalysts, the detailed understanding of the
Makoto Komiyama; Jun Sumaoka
Formation of the N(2)-acetyl-2,6-diaminopurine oligonucleotide impurity caused by acetyl capping.
The acetyl 'capping' reaction routinely employed during phosphorothioate oligonucleotide synthesis has been implicated in the formation of an impurity species with a mass 41amu greater than the expected oligonucleotide molecule. The impurity has been found to arise by conversion of a protected guanine nucleobase to N(2)-acetyl-2,6-diaminopurine. A two-part mechanism is proposed consisting of transamidation of the protecting group on guanine and substitution of guanine's O(6) atom. PMID:24980055
Rodriguez, Andrew A; Cedillo, Isaiah; Mowery, Brendan P; Gaus, Hans J; Krishnamoorthy, Seetha S; McPherson, Andrew K
Acetylation and characterization of spruce ( Picea abies) galactoglucomannans
Acetylated galactoglucomannans (GGMs) are the main hemicellulose type in most softwood species and can be utilized as, for example, bioactive polymers, hydrocolloids, papermaking chemicals, or coating polymers. Acetylation of spruce GGM using acetic anhydride with pyridine as catalyst under different conditions was conducted to obtain different degrees of acetylation on a laboratory scale, whereas, as a classic method, it can
Chunlin Xu; Ann-Sofie Lepp�nen; Patrik Eklund; Peter Holmlund; Rainer Sj�holm; Kenneth Sundberg; Stefan Willf�r
The Development of a Decision Analytic Model of Changes in Mean Deviation in People with Glaucoma: The COA Model
Purpose To create and validate a statistical model predicting progression of primary open angle glaucoma (POAG) assessed by loss of visual field as measured in mean deviation (MD) using three landmark studies of glaucoma progression and treatment. Design A Markov decision analytic model using patient level data described longitudinal MD changes over seven years. Participants Patient level data from the Collaborative Initial Glaucoma Treatment Study (CIGTS, n=607), the Ocular Hypertension Treatment Study (OHTS, n=148, only those who developed POAG in the first five years of OHTS) and Advanced Glaucoma Intervention Study (AGIS, n=591), the COA model. Methods We developed a Markov model with transition matrices stratified by current MD, age, race and intraocular pressure categories and used a microsimulation approach to estimate change in MD over seven years. Internal validation compared model prediction for seven years to actual MD for COA participants. External validation used a cohort of glaucoma patients drawn from university clinical practices. Main Outcome Measures Change in visual field as measured in MD in decibels (dB). Results Regressing the actual MD against the predicted produced an R2 of 0.68 for the right eye and 0.63 for the left. The model predicted ending MD for right eyes of 65% of participants and for 63% of left eyes within 3 dB of actual results at seven years. In external validation the model had an R2 of 0.79 in the right eye and 0.77 in the left at five years. Conclusion The COA model is a validated tool for clinicians, patients and health policy makers seeking to understand longitudinal changes in mean deviation in people with glaucoma..
Kymes, Steven M.; Lambert, Dennis L.; Lee, Paul P.; Musch, David C.; Siegfried, Carla J.; Kotak, Sameer V.; Stwalley, Dustin L.; Fain, Joel; Johnson, Chris; Gordon, Mae O.
Insights into the Regulatory Characteristics of the Mycobacterial Dephosphocoenzyme A Kinase: Implications for the Universal CoA Biosynthesis Pathway
Being vastly different from the human counterpart, we suggest that the last enzyme of the Mycobacterium tuberculosis Coenzyme A biosynthetic pathway, dephosphocoenzyme A kinase (CoaE) could be a good anti-tubercular target. Here we describe detailed investigations into the regulatory features of the enzyme, affected via two mechanisms. Enzymatic activity is regulated by CTP which strongly binds the enzyme at a site overlapping that of the leading substrate, dephosphocoenzyme A (DCoA), thereby obscuring the binding site and limiting catalysis. The organism has evolved a second layer of regulation by employing a dynamic equilibrium between the trimeric and monomeric forms of CoaE as a means of regulating the effective concentration of active enzyme. We show that the monomer is the active form of the enzyme and the interplay between the regulator, CTP and the substrate, DCoA, affects enzymatic activity. Detailed kinetic data have been corroborated by size exclusion chromatography, dynamic light scattering, glutaraldehyde crosslinking, limited proteolysis and fluorescence investigations on the enzyme all of which corroborate the effects of the ligands on the enzyme oligomeric status and activity. Cysteine mutagenesis and the effects of reducing agents on mycobacterial CoaE oligomerization further validate that the latter is not cysteine-mediated or reduction-sensitive. These studies thus shed light on the novel regulatory features employed to regulate metabolite flow through the last step of a critical biosynthetic pathway by keeping the latter catalytically dormant till the need arises, the transition to the active form affected by a delicate crosstalk between an essential cellular metabolite (CTP) and the precursor to the pathway end-product (DCoA).
Walia, Guneet; Surolia, Avadhesha
Human Histone Chaperone Nucleophosmin Enhances Acetylation-Dependent Chromatin Transcription�
Histone chaperones are a group of proteins that aid in the dynamic chromatin organization during different cellular processes. Here, we report that the human histone chaperone nucleophosmin interacts with the core histones H3, H2B, and H4 but that this histone interaction is not sufficient to confer the chaperone activity. Significantly, nucleophosmin enhances the acetylation-dependent chromatin transcription and it becomes acetylated both in vitro and in vivo. Acetylation of nucleophosmin and the core histones was found to be essential for the enhancement of chromatin transcription. The acetylated NPM1 not only shows an increased affinity toward acetylated histones but also shows enhanced histone transfer ability. Presumably, nucleophosmin disrupts the nucleosomal structure in an acetylation-dependent manner, resulting in the transcriptional activation. These results establish nucleophosmin (NPM1) as a human histone chaperone that becomes acetylated, resulting in the enhancement of chromatin transcription.
Swaminathan, V.; Kishore, A. Hari; Febitha, K. K.; Kundu, Tapas K.
Acetylator phenotype in Iraqi patients with systemic lupus erythematosus.
The study was designed to determine the acetylator status in patients with systemic lupus erythematosus (SLE) and compare it to a matched group of healthy volunteers. Disease severity was determined using the revised American College of Rheumatology criteria for classification and the SLE disease activity index. After an overnight fast, each participant received a single oral dose of 100 mg dapsone. After 3 hours, plasma dapsone/monoacetyldapsone ratio was determined. In the control group, frequency of slow acetylators was 73.3%; frequency of rapid acetylators was 26.7%. In SLE patients, frequency of slow acetylators was 78.0%; frequency of rapid acetylators was 12.0%. However, 8.0% were non-acetylators (monoacetyldapsone not detected in plasma). There was no association between acetylator status and severity of SLE. PMID:16761671
Najim, R A; Farid, Y Y Z; Samad, T Abed; Shihab, S A R
Enzymatic sequencing of partially acetylated chitosan oligomers.
Chitosan oligosaccharides have diverse biological activities with potentially valuable applications, for example, in the fields of medicine and agriculture. These functionalities are thought to depend on their degree of polymerization and acetylation, and possibly on specific patterns of acetylation. Chitosan oligomers with fully defined architecture are difficult to produce, and their complete analysis is demanding. Analysis is typically done using MS or NMR, requiring access to expensive infrastructure, and yielding unequivocal results only in the case of rather small oligomers. We here describe a simple and cost-efficient method for the sequencing of ?g amounts of chitosan oligosaccharides which is based on the sequential action of two recombinant glycosidases, namely an exo-?-N-acetylhexosaminidase (GlcNAcase) from Bacillus subtilis 168 and an exo-?-d-glucosaminidase (GlcNase) from Thermococcus kodakarensis KOD1. Starting from the non-reducing end, GlcNAcase and GlcNase specifically remove N-acetyl glucosamine (A) and glucosamine (D) units, respectively. By the sequential addition and removal of these enzymes in an alternating way followed by analysis of the products using high-performance thin-layer chromatography, the sequence of chitosan oligosaccharides can be revealed. Importantly, both enzymes work under identical conditions so that no buffer exchange is required between steps, and the enzyme can be removed conveniently using simple ultra-filtration devices. As proof-of-principle, the method was used to sequence the product of enzymatic deacetylation of chitin pentamer using a recombinant chitin deacetylase from Vibrio cholerae which specifically removes the acetyl group from the second unit next to the non-reducing end of the substrate, yielding mono-deacetylated pentamer with the sequence ADAAA. PMID:24824785
Hamer, Stefanie Nicole; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan
Fragrance material review on acetyl cedrene.
A toxicologic and dermatologic review of acetyl cedrene when used as a fragrance ingredient is presented. Acetyl cedrene is a member of the fragrance structural group Alkyl Cyclic Ketones. The generic formula for this group can be represented as (R1)(R2)CO. These fragrances can be described as being composed of an alkyl, R1, and various substituted and bicyclic saturated or unsaturated cyclic hydrocarbons, R2, in which one of the rings may include up to 12 carbons. Alternatively, R2 may be a carbon bridge of C2-C4 carbon chain length between the ketone and cyclic hydrocarbon. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for acetyl cedrene were evaluated then summarized and includes physical properties, acute toxicity, skin irritation, mucous membrane (eye) irritation, skin sensitization, elicitation, phototoxicity, photoallergy, toxicokinetics, repeated dose, reproductive toxicity, and genotoxicity data. A safety assessment of the entire Alkyl Cyclic Ketones will be published simultaneously with this document; please refer to Belsito et al. (2013) (Belsito, D., Bickers, D., Bruze, M., Calow, P., Dagli, M., Fryer, A.D., Greim, H., Miyachi, Y., Saurat, J.H., Sipes, I.G., 2013. A Toxicologic and Dermatologic Assessment of Alkyl Cyclic Ketones When Used as Fragrance Ingredients. Submitted with this manuscript.) for an overall assessment of the safe use of this material and all Alkyl Cyclic Ketones in fragrances. PMID:23907023
Scognamiglio, J; Letizia, C S; Politano, V T; Api, A M
Fragrance material review on acetyl carene.
A toxicologic and dermatologic review of acetyl carene when used as a fragrance ingredient is presented. Acetyl carene is a member of the fragrance structural group Alkyl Cyclic Ketones. These fragrances can be described as being composed of an alkyl, R1, and various substituted and bicyclic saturated or unsaturated cyclic hydrocarbons, R2, in which one of the rings may include up to 12 carbons. Alternatively, R2 may be a carbon bridge of C2-C4 carbon chain length between the ketone and cyclic hydrocarbon. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for acetyl carene were evaluated then summarized and includes physical properties, acute toxicity, skin irritation, and skin sensitization data. A safety assessment of the entire Alkyl Cyclic Ketones will be published simultaneously with this document; please refer to Belsito et al. (Belsito, D., Bickers, D., Bruze, M., Calow, P., Dagli, M., Fryer, A.D., Greim, H., Miyachi, Y., Saurat, J.H., Sipes, I.G., 2013A Toxicologic and dermatologic assessment of alkyl cyclic ketones when used as fragrance ingredients. (submitted for publication).) for an overall assessment of the safe use of this material and all Alkyl Cyclic Ketones in fragrances. PMID:23911801
Scognamiglio, J; Letizia, C S; Api, A M
Poly-acetylated chromatin signatures are preferred epitopes for site-specific histone H4 acetyl antibodies
Antibodies specific for histone post-translational modifications (PTMs) have been central to our understanding of chromatin biology. Here, we describe an unexpected and novel property of histone H4 site-specific acetyl antibodies in that they prefer poly-acetylated histone substrates. By all current criteria, these antibodies have passed specificity standards. However, we find these site-specific histone antibodies preferentially recognize chromatin signatures containing two or more adjacent acetylated lysines. Significantly, we find that the poly-acetylated epitopes these antibodies prefer are evolutionarily conserved and are present at levels that compete for these antibodies over the intended individual acetylation sites. This alarming property of acetyl-specific antibodies has far-reaching implications for data interpretation and may present a challenge for the future study of acetylated histone and non-histone proteins.
Rothbart, Scott B.; Lin, Shu; Britton, Laura-Mae; Krajewski, Krzysztof; Keogh, Michael-C; Garcia, Benjamin A.; Strahl, Brian D.
Formyl-CoA transferase encloses the CoA binding site at the interface of an interlocked dimer
Formyl-CoA transferase catalyses transfer of CoA from formate to oxalate in the first step of oxalate degradation by Oxalobacter formigenes, a bacterium present in the intestinal flora which is implicated in oxalate catabolism in mammals. Formyl-CoA transferase is a member of a family of CoA-transferases for which no structural information is available. We now report the three-dimensional structure of O.formigenes formyl-CoA transferase, which reveals a novel fold and a very striking assembly of the homodimer. The subunit is composed of a large and a small domain where residues from both the N- and C-termini of the subunit are part of the large domain. The linkers between the domains give the subunit a circular shape with a hole in the middle. The enzyme monomers are tightly interacting and are interlocked. This fold requires drastic rearrangement of ?75 residues at the C-terminus for formation of the dimer. The structure of a complex of formyl-CoA transferase with CoA is also reported and sets the scene for a mechanistic understanding of enzymes of this family of CoA-transferases.
Ricagno, Stefano; Jonsson, Stefan; Richards, Nigel; Lindqvist, Ylva
Screening, identification, and characterization of mechanistically diverse inhibitors of the Mycobacterium tuberculosis enzyme, pantothenate kinase (CoaA).
The authors describe the discovery of anti-mycobacterial compounds through identifying mechanistically diverse inhibitors of the essential Mycobacterium tuberculosis (Mtb) enzyme, pantothenate kinase (CoaA). Target-driven drug discovery technologies often work with purified enzymes, and inhibitors thus discovered may not optimally inhibit the form of the target enzyme predominant in the bacterial cell or may not be available at the desired concentration. Therefore, in addition to addressing entry or efflux issues, inhibitors with diverse mechanisms of inhibition (MoI) could be prioritized before hit-to-lead optimization. The authors describe a high-throughput assay based on protein thermal melting to screen large numbers of compounds for hits with diverse MoI. Following high-throughput screening for Mtb CoaA enzyme inhibitors, a concentration-dependent increase in protein thermal stability was used to identify true binders, and the degree of enhancement or reduction in thermal stability in the presence of substrate was used to classify inhibitors as competitive or non/uncompetitive. The thermal shift-based MoI assay could be adapted to screen hundreds of compounds in a single experiment as compared to traditional biochemical approaches for MoI determination. This MoI was confirmed through mechanistic studies that estimated K(ie) and K(ies) for representative compounds and through nuclear magnetic resonance-based ligand displacement assays. PMID:22086722
Venkatraman, Janani; Bhat, Jyothi; Solapure, Suresh M; Sandesh, Jatheendranath; Sarkar, Debasmita; Aishwarya, Sundaram; Mukherjee, Kakoli; Datta, Santanu; Malolanarasimhan, Krishnan; Bandodkar, Balachandra; Das, Kaveri S
Synthesis and biological activity of O-N-acetyl-beta-muramoyl-L-alanyl-D-isoglutamine)-(1 leads to 6)-2-acylamino-2-deoxy-D-glucoses.
Benzoylation of benzyl 2-acetamido-2-deoxy-4,6-O-isopropylidene-alpha-D-glucopyranoside, benzyl 2-deoxy-2-(DL-3-hydroxytetradecanoylamino)-4,6-O-isopropylidene-alpha-D-glucopyranoside, and benzyl 2-deoxy-4,6-O-isopropylidene-2-octadecanoylamino-beta-D-glucopyranoside, with subsequent hydrolysis of the 4,6-O-isopropylidene group, gave the corresponding 3-O-benzoyl derivatives (4, 5, and 7). Hydrogenation of benzyl 2-acetamido-4,6-di-O-acetyl-2-deoxy-3-O-[D-1-(methoxycarbonyl)ethyl]-alpha-D-glucopyranoside, followed by chlorination, gave a product that was treated with mercuric actate to yield 2-acetamido-1,4,6-tri-O-acetyl-2-deoxy-3-O-[D-1-(methoxy-carbonyl)ethyl]-beta-D -glucopyranose (11). Treatment of 11 with ferric chloride afforded the oxazoline derivative, which was condensed with 4, 5, and 7 to give the (1 goes to 6)-beta-linked disaccharide derivative 13, 15, and 17. Hydrolysis of the methyl ester group in the compounds derived from 13, 15, and 17 by 4-O-acetylation gave the corresponding free acids, which were coupled with L-alanyl-D-isoglutamine benzyl ester, to yield the dipeptide derivatives 19-21 in excellent yields. Hydrolysis of 19-21, followed by hydrogenation, gave the respective O-(N-acetyl-beta-muramoyl-L-alanyl-D-isoglutamine)-(1 goes to 6)-2-acylamino-2-deoxy-D-glucoses in good yields. The immuno-adjuvant activity of these compounds was examined in guinea-pigs. PMID:7083252
Hasegawa, A; Ozaki, M; Goh, Y; Kiso, M; Azuma, I
Sucrose Hydrolysis at Limited Water Concentration.
National Technical Information Service (NTIS)
To enable development of a model describing reaction kinetics in dehydrated foods, sucrose hydrolysis was studied at limited water concentration. Saturated sucrose solutions containing various acids and inert solid materials gave identical rate constants ...
T. Schoebel S. R. Tannenbaum T. P. Labuza
Effect of surfactants on cellulose hydrolysis
The effect of surfactants on the heterogeneous enzymatic hydrolysis of Sigmacell 100 cellulose and of steam-exploded wood was studied. Certain biosurfactants (sophorolipid, rhamnolipid, bacitracin) and Tween 80 increased the rate of hydrolysis of Sigmacell 100, as measured by the amount of reducing sugar produced, by as much as seven times. The hydrolysis of steam-exploded wood was increased by 67% in the presence of sophorolipid. At the same time, sophorolipid was found to decrease the amount of enzyme absorbed onto the cellulose at equilibrium. Sophorolipid had the greatest effect on cellulose hydrolysis when it was present from the beginning of the experiment and when the enzyme/cellulose ratio was low.
Helle, S.S.; Duff, S.J.B. (Univ. of British Columbia, Vancouver, British Columbia (Canada). Pulp and Paper Centre and Dept. of Chemical Engineering); Cooper, D.G. (McGill Univ., Montreal, Quebec (Canada). Chemical Engineering Dept.)
The biology of lysine acetylation integrates transcriptional programming and metabolism
The biochemical landscape of lysine acetylation has expanded from a small number of proteins in the nucleus to a multitude of proteins in the cytoplasm. Since the first report confirming acetylation of the tumor suppressor protein p53 by a lysine acetyltransferase (KAT), there has been a surge in the identification of new, non-histone targets of KATs. Added to the known substrates of KATs are metabolic enzymes, cytoskeletal proteins, molecular chaperones, ribosomal proteins and nuclear import factors. Emerging studies demonstrate that no fewer than 2000 proteins in any particular cell type may undergo lysine acetylation. As described in this review, our analyses of cellular acetylated proteins using DAVID 6.7 bioinformatics resources have facilitated organization of acetylated proteins into functional clusters integral to cell signaling, the stress response, proteolysis, apoptosis, metabolism, and neuronal development. In addition, these clusters also depict association of acetylated proteins with human diseases. These findings not only support lysine acetylation as a widespread cellular phenomenon, but also impel questions to clarify the underlying molecular and cellular mechanisms governing target selectivity by KATs. Present challenges are to understand the molecular basis for the overlapping roles of KAT-containing co-activators, to differentiate between global versus dynamic acetylation marks, and to elucidate the physiological roles of acetylated proteins in biochemical pathways. In addition to discussing the cellular 'acetylome', a focus of this work is to present the widespread and dynamic nature of lysine acetylation and highlight the nexus that exists between epigenetic-directed transcriptional regulation and metabolism.
O-Acetylation of Plant Cell Wall Polysaccharides
Plant cell walls are composed of structurally diverse polymers, many of which are O-acetylated. How plants O-acetylate wall polymers and what its function is remained elusive until recently, when two protein families were identified in the model plant Arabidopsis that are involved in the O-acetylation of wall polysaccharides � the reduced wall acetylation (RWA) and the trichome birefringence-like (TBL) proteins. This review discusses the role of these two protein families in polysaccharide O-acetylation and outlines the differences and similarities of polymer acetylation mechanisms in plants, fungi, bacteria, and mammals. Members of the TBL protein family had been shown to impact pathogen resistance, freezing tolerance, and cellulose biosynthesis. The connection of TBLs to polysaccharide O-acetylation thus gives crucial leads into the biological function of wall polymer O-acetylation. From a biotechnological point understanding the O-acetylation mechanism is important as acetyl-substituents inhibit the enzymatic degradation of wall polymers and released acetate can be a potent inhibitor in microbial fermentations, thus impacting the economic viability of, e.g., lignocellulosic based biofuel production.
Gille, Sascha; Pauly, Markus
Hydrolysis of steam-pretreated lignocellulose
The mechanism of hydrolysis of cellulose is important for improving the enzymatic conversion in bioprocesses based on lignocellulose.\\u000a Adsorption and hydrolysis experiments were performed with cellobiohydrolase I (CBH I) and endoglucanase II (EG II) from Trichoderma reesei on a realistic lignocellulose substrates: steam-pretreated willow. The enzymes were studied both alone and in equimolar mixtures.\\u000a Adsorption isotherms were determined at 4
Johan Karlsson; J�zsef Medve; Folke Tjerneld
The complete enzymic hydrolysis of crosslinked proteins.
Procedures used for the complete enzymic hydrolysis of proteins are reviewed. The successful application of complete enzymic hydrolysis in the detection of naturally occurring isopeptide crosslinks and various other types of chemically introduced crosslinks is described. The method may fail if the level of crosslinking is too high, or if crosslinking is accompanied by racemization. Although it is usual to cleave disulphide crosslinks prior to enzymic digestion, the necessity for this in all cases is questioned. PMID:906920
Milligan, B; Holt, L A
Insertional Inactivation of Methylmalonyl Coenzyme A (CoA) Mutase and Isobutyryl-CoA Mutase Genes in Streptomyces cinnamonensis: Influence on Polyketide Antibiotic Biosynthesis
The coenzyme B12-dependent isobutyryl coenzyme A (CoA) mutase (ICM) and methylmalonyl-CoA mutase (MCM) catalyze the isomerization of n-butyryl-CoA to isobutyryl-CoA and of methylmalonyl-CoA to succinyl- CoA, respectively. The influence that both mutases have on the conversion of n- and isobutyryl-CoA to methylmalonyl-CoA and the use of the latter in polyketide biosynthesis have been investigated with the polyether antibiotic (monensin) producer
Acid mediated hydrolysis of blueberry anthocyanins.
Acid mediated hydrolysis of anthocyanins was studied using capillary zone electrophoresis (CZE). A commercially available wild blueberry (Bilberry) extract was dissolved in different concentrations of TFA (0.1, 1, 3, 9%), then was subjected to thermodecomposition reaction at 95 degrees C. After the reaction, the samples were analyzed by CZE. The hydrolysis rate of each anthocyanin and the formation of the aglycon were determined by the change in the peak pattern of the anthocyanins in the electropherogram. Each anthocyanin peak decreased time dependently in a first order kinetic fashion. It was revealed that the hydrolysis rate of each anthocyanin was determined primarily by the type of conjugated sugar and not by the aglycon structure. The rate constant of anthocyanin hydrolysis was in the following order, arabinoside>galactoside>glucoside without regard to the aglycon structure. The kinetic behavior of this anthocyanin hydrolysis together with the CZE mobility allowed us to identify an unknown CZE peak as delphinidin 3-O-beta-arabinoside. At low TFA concentration, significant decomposition of the anthocyanidin nucleus occurred, but the glycoside hydrolysis predominated at high TFA concentration. It was further revealed that the aglycon released reacted successively to form polymeric products at higher TFA conditions. PMID:11201215
Ichiyanagi, T; Oikawa, K; Tateyama, C; Konishi, T
Molecular Bases for Sensitivity to Acetyl-Coenzyme A Carboxylase Inhibitors in Black-Grass1
In grasses, residues homologous to residues Ile-1,781 and Ile-2,041 in the carboxyl-transferase (CT) domain of the chloroplastic acetyl-coenzyme A (CoA) carboxylase (ACCase) from the grass weed black-grass (Alopecurus myosuroides [Huds.]) are critical determinants for sensitivity to two classes of ACCase inhibitors, aryloxyphenoxypropionates (APPs) and cyclohexanediones. Using natural mutants of black-grass, we demonstrated through a molecular, biological, and biochemical approach that residues Trp-2,027, Asp-2,078, and Gly-2,096 are also involved in sensitivity to ACCase inhibitors. In addition, residues Trp-2,027 and Asp-2,078 are very likely involved in CT activity. Using three-dimensional modeling, we found that the side chains of the five residues are adjacent, located at the surface of the inside of the cavity of the CT active site, in the vicinity of the binding site for APPs. Residues 1,781 and 2,078 are involved in sensitivity to both APPs and cyclohexanediones, whereas residues 2,027, 2,041, and 2,096 are involved in sensitivity to APPs only. This suggests that the binding sites for these two classes of compounds are overlapping, although distinct. Comparison of three-dimensional models for black-grass wild-type and mutant CTs and for CTs from organisms with contrasted sensitivity to ACCase inhibitors suggested that inhibitors fitting into the cavity of the CT active site of the chloroplastic ACCase from grasses to reach their active sites may be tight. The three-dimensional shape of this cavity is thus likely of high importance for the efficacy of ACCase inhibitors.
Delye, Christophe; Zhang, Xiao-Qi; Michel, Severine; Matejicek, Annick; Powles, Stephen B.
Biochemical and Crystallographic Analysis of Substrate Binding and Conformational Changes in Acetyl-CoA Synthetase
The adenylate-forming enzymes, including acyl-CoA synthetases, the adenylation domains of non-ribosomal peptide synthetases (NRPS), and firefly luciferase, perform two half-reactions in a ping-pong mechanism. We have proposed a domain alternation mechanism for these enzymes whereby, upon completion of the initial adenylation reaction, the C-terminal domain of these enzymes undergoes a 140{sup o} rotation to perform the second thioester-forming half-reaction. Structural and kinetic data of mutant enzymes support this hypothesis. We present here mutations to Salmonella enterica acetyl-CoA synthetase (Acs) and test the ability of the enzymes to catalyze the complete reaction and the adenylation half-reaction. Substitution of Lys609 with alanine results in an enzyme that is unable to catalyze the adenylate reaction, while the Gly524 to leucine substitution is unable to catalyze the complete reaction yet catalyzes the adenylation half-reaction with activity comparable to the wild-type enzyme. The positions of these two residues, which are located on the mobile C-terminal domain, strongly support the domain alternation hypothesis. We also present steady-state kinetic data of putative substrate-binding residues and demonstrate that no single residue plays a dominant role in dictating CoA binding. We have also created two mutations in the active site to alter the acyl substrate specificity. Finally, the crystallographic structures of wild-type Acs and mutants R194A, R584A, R584E, K609A, and V386A are presented to support the biochemical analysis.
Reger,A.; Carney, J.; Gulick, A.
A yeast acetyl coenzyme A carboxylase mutant links very-long-chain fatty acid synthesis to the structure and function of the nuclear membrane-pore complex.
The conditional mRNA transport mutant of Saccharomyces cerevisiae, acc1-7-1 (mtr7-1), displays a unique alteration of the nuclear envelope. Unlike nucleoporin mutants and other RNA transport mutants, the intermembrane space expands, protuberances extend from the inner membrane into the intermembrane space, and vesicles accumulate in the intermembrane space. MTR7 is the same gene as ACC1, encoding acetyl coenzyme A (CoA) carboxylase (Acc1p), the rate-limiting enzyme of de novo fatty acid synthesis. Genetic and biochemical analyses of fatty acid synthesis mutants and acc1-7-1 indicate that the continued synthesis of malonyl-CoA, the enzymatic product of acetyl-CoA carboxylase, is required for an essential pathway which is independent from de novo synthesis of fatty acids. We provide evidence that synthesis of very-long-chain fatty acids (C26 atoms) is inhibited in acc1-7-1, suggesting that very-long-chain fatty acid synthesis is required to maintain a functional nuclear envelope.
Schneiter, R; Hitomi, M; Ivessa, A S; Fasch, E V; Kohlwein, S D; Tartakoff, A M
Acetylation and characterization of spruce (Picea abies) galactoglucomannans.
Acetylated galactoglucomannans (GGMs) are the main hemicellulose type in most softwood species and can be utilized as, for example, bioactive polymers, hydrocolloids, papermaking chemicals, or coating polymers. Acetylation of spruce GGM using acetic anhydride with pyridine as catalyst under different conditions was conducted to obtain different degrees of acetylation on a laboratory scale, whereas, as a classic method, it can be potentially transferred to the industrial scale. The effects of the amount of catalyst and acetic anhydride, reaction time, temperature and pretreatment by acetic acid were investigated. A fully acetylated product was obtained by refluxing GGM for two hours. The structures of the acetylated GGMs were determined by SEC-MALLS/RI, (1)H and (13)C NMR and FTIR spectroscopy. NMR studies also indicated migration of acetyl groups from O-2 or O-3 to O-6 after a heating treatment in a water bath. The thermal stability of the products was investigated by DSC-TGA. PMID:20144827
Xu, Chunlin; Lepp�nen, Ann-Sofie; Eklund, Patrik; Holmlund, Peter; Sj�holm, Rainer; Sundberg, Kenneth; Willf�r, Stefan
Function and regulation of isoforms of carbon monoxide dehydrogenase/acetyl coenzyme A synthase in Methanosarcina acetivorans.
Conversion of acetate to methane (aceticlastic methanogenesis) is an ecologically important process carried out exclusively by methanogenic archaea. An important enzyme for this process as well as for methanogenic growth on carbon monoxide is the five-subunit archaeal CO dehydrogenase/acetyl coenzyme A (CoA) synthase multienzyme complex (CODH/ACS) catalyzing both CO oxidation/CO(2) reduction and cleavage/synthesis of acetyl-CoA. Methanosarcina acetivorans C2A contains two very similar copies of a six-gene operon (cdh genes) encoding two isoforms of CODH/ACS (Cdh1 and Cdh2) and a single CdhA subunit, CdhA3. To address the role of the CODH/ACS system in M. acetivorans, mutational as well as promoter/reporter gene fusion analyses were conducted. Phenotypic characterization of cdh disruption mutants (three single and double mutants, as well as the triple mutant) revealed a strict requirement of either Cdh1 or Cdh2 for acetotrophic or carboxidotrophic growth, as well as for autotrophy, which demonstrated that both isoforms are bona fide CODH/ACS. While expression of the Cdh2-encoding genes was generally higher than that of genes encoding Cdh1, both appeared to be regulated differentially in response to growth phase and to changing substrate conditions. While dispensable for growth, CdhA3 clearly affected expression of cdh1, suggesting that it functions in signal perception and transduction rather than in catabolism. The data obtained argue for a functional hierarchy and regulatory cross talk of the CODH/ACS isoforms. PMID:22865842
Matschiavelli, Nicole; Oelgeschl�ger, Ellen; Cocchiararo, Berardino; Finke, Johannes; Rother, Michael
Acetylation pharmacogenetics and renal function in diabetes mellitus patients.
Activities of human hepatic drug metabolizing enzymes N-acetyl transferase (NATS) had earlier been recognized as a cause of inter-individual variation in the metabolism of drugs. Therefore acetylation of many drugs in human exhibit genetic polymorphism. The aim of the study was to investigate if acetylator status predispose diabetic mellitus patients more to the complications of renal disease, One hundred and twenty (120) diabetics consisting of (50) Type 1 (T(1)) and 70 Type 2 (T(2)) diabetes mellitus patients and 100 healthy individuals as controls were classified as slow or rapid acetylator using sulphamethazine (SMZ) as an in vivo probe. The percentage acetylation, recovery of SMZ, creatinine clearance and presence of urinary albumin were determined. A significant difference (P�<�0.05) was observed in the percentage of SMZ acetylated between slow and rapid acetylators in control, T(1) and T(2) subjects. The ratios of slow to rapid acetylators for T(1), T(2) and control subjects were 1:4, 3:2 and 2:3 respectively. No significant differences were observed in the percentage of SMZ recovered in the urine of slow and rapid acetylators that are diabetics. The difference in creatinine clearance of slow and rapid acetylators in T(1) and T(2) were significant (P�<�0.05). 29% out of 120 (24.2%) diabetics (T(1) and T(2)) exhibited albuminuria out of which 25 (86.2%) had slow acetylator status. These findings suggest that slow acetylator status in diabetes mellitus could be a predisposing factor in the development of renal complications. This underscores the need for a rapid pharmacogenetic testing and therapeutic drug monitoring in such patients. However this inference could be further validated with a larger sample size. PMID:21731200
Banjoko, S O; Akinlade, K S
Acetylation Regulates Transcription Factor Activity at Multiple Levels
CREB-binding protein (CBP) possesses an intrinsic acetyltransferase activity capable of acetylating nucleosomal histones as well as several nonhistone proteins. Here, it is shown that CBP can acetylate hepatocyte nuclear factor-4 (HNF-4), a member of the nuclear hormone receptor family, at lysine residues within the nuclear localization sequence. CBP-mediated acetylation is crucial for the proper nuclear retention of HNF-4, which is
Evi Soutoglou; Nitsa Katrakili; Iannis Talianidis
Dynamic Histone Acetylation of Late Embryonic Genes during Seed Germination
Histone acetylation is involved in the regulation of gene expression in plants and eukaryotes. Histone deacetylases (HDACs)\\u000a are enzymes that catalyze the removal of acetyl groups from histones, which is associated with the repression of gene expression.\\u000a To study the role of histone acetylation in the regulation of gene expression during seed germination, trichostatin A (TSA),\\u000a a specific inhibitor of
Helen H. Tai; George C. C. Tai; Tannis Beardmore
[The acetylator phenotype in patients with urinary bladder cancer].
In 53 patients suffering from bladder cancer the acelylization phenotype was examined. The frequency of slow acetylizers (70%) was significantly higher than in the control group. The more frequently occurrence of slow acetylizers in the group with professional pollutant exposition is an important reference to a higher bladder cancer morbidity in the case of carcinogen contact. A screening of the acetylization phenotype in professions with carcinogen contact may be important in the prophylaxis of bladder cancer. PMID:2618184
Wich, H; Franke, G; Grimm, U; Siegmund, W
Acetylation Pharmacogenetics and Renal Function in Diabetes Mellitus Patients
Activities of human hepatic drug metabolizing enzymes N-acetyl transferase (NATS) had earlier been recognized as a cause of inter-individual variation in the metabolism of drugs.\\u000a Therefore acetylation of many drugs in human exhibit genetic polymorphism. The aim of the study was to investigate if acetylator\\u000a status predispose diabetic mellitus patients more to the complications of renal disease, One hundred and
S. O. Banjoko; K. S. Akinlade
3-Acetyl-4-hydroxy-phenyl acrylate
In the title compound, C12H12O4, the hydr�oxy O and the C and O atoms of the acetyl group are almost coplanar [maximum deviation = 0.0356?(1)?�] with the benzene ring. The dihedral angle between the benzene ring and the plane through the non-H atoms of the methacrylo�yloxy group is 86.1?(1)�. In the crystal structure, mol�ecules are linked by two C�H?O hydrogen bonds, forming dimers with graph-set descriptor R 2 2(16). A strong intra�molecular O�H?O hydrogen bond is also observed.
Azeezaa, V.; Usha, G.; Bhaskaran, Sundari; Anthonysamy, A.; Balasubramanian, S.
New Targets for Acetylation in Autophagy
NSDL National Science Digital Library
Macroautophagy is an evolutionarily conserved homeostatic process that mediates the degradation of long-lived cytoplasmic components in eukaryotes, which allows cells to survive stresses such as inflammation, hypoxia, and deprivation of nutrients or growth factors. At least 30 members of the Atg (autophagy-related) protein family orchestrate this degradative process. Additional complexity resides in the signaling networks controlling the autophagic process, which include various posttranslational modifications of key components. Evidence is accumulating that protein acetylation represents an evolutionarily conserved mechanism tightly regulating macroautophagy.
Ahmed Hamai (Paris;INSERM REV); Patrice Codogno (Paris;INSERM REV)
Role of transcription factor acetylation in diabetic kidney disease.
Nuclear factor (NF)-?B and signal transducer and activator of transcription 3 (STAT3) play a critical role in diabetic nephropathy (DN). Sirtuin-1 (SIRT1) regulates transcriptional activation of target genes through protein deacetylation. Here, we determined the roles of Sirt1 and the effect of NF-?B (p65) and STAT3 acetylation in DN. We found that acetylation of p65 and STAT3 was increased in both mouse and human diabetic kidneys. In human podocytes, advanced glycation end products (AGEs) induced p65 and STAT3 acetylation and overexpression of acetylation-incompetent mutants of p65 and STAT3 abrogated AGE-induced expression of NF-?B and STAT3 target genes. Inhibition of AGE formation in db/db mice by pyridoxamine treatment attenuated proteinuria and podocyte injury, restored SIRT1 expression, and reduced p65 and STAT3 acetylation. Diabetic db/db mice with conditional deletion of SIRT1 in podocytes developed more proteinuria, kidney injury, and acetylation of p65 and STAT3 compared with db/db mice without SIRT1 deletion. Treatment of db/db mice with a bromodomain and extraterminal (BET)-specific bromodomain inhibitor (MS417) which blocks acetylation-mediated association of p65 and STAT3 with BET proteins, attenuated proteinuria, and kidney injury. Our findings strongly support a critical role for p65 and STAT3 acetylation in DN. Targeting protein acetylation could be a potential new therapy for DN. PMID:24608443
Liu, Ruijie; Zhong, Yifei; Li, Xuezhu; Chen, Haibing; Jim, Belinda; Zhou, Ming-Ming; Chuang, Peter Y; He, John Cijiang
Measurement of acetylation turnover at distinct lysines in human histones identifies long-lived acetylation sites.
Histone acetylation has long been determined as a highly dynamic modification associated with open chromatin and transcriptional activation. Here we develop a metabolic labelling scheme using stable isotopes to study the kinetics of acetylation turnover at 19 distinct lysines on histones H3, H4 and H2A. Using human HeLa S3 cells, the analysis reveals 12 sites of histone acetylation with fast turnover and 7 sites stable over a 30 h experiment. The sites showing fast turnover (anticipated from classical radioactive measurements of whole histones) have half-lives between ~1-2 h. To support this finding, we use a broad-spectrum deacetylase inhibitor to verify that only fast turnover sites display 2-10-fold increases in acetylation whereas long-lived sites clearly do not. Most of these stable sites lack extensive functional studies or localization within global chromatin, and their role in non-genetic mechanisms of inheritance is as yet unknown. PMID:23892279
Zheng, Yupeng; Thomas, Paul M; Kelleher, Neil L
Enzymatic saccharification of pretreated wheat straw: comparison of solids-recycling, sequential hydrolysis and batch hydrolysis.
In the enzymatic hydrolysis of lignocellulose materials, the recycling of the solid residue has previously been considered within the context of enzyme recycling. In this study, a steady state investigation of a solids-recycling process was made with pretreated wheat straw and compared to sequential and batch hydrolysis at constant reaction times, substrate feed and liquid and enzyme consumption. Compared to batch hydrolysis, the recycling and sequential processes showed roughly equal hydrolysis yields, while the volumetric productivity was significantly increased. In the 72h process the improvement was 90% due to an increased reaction consistency, while the solids feed was 16% of the total process constituents. The improvement resulted primarily from product removal, which was equally efficient in solids-recycling and sequential hydrolysis processes. No evidence of accumulation of enzymes beyond the accumulation of the substrate was found in recycling. A mathematical model of solids-recycling was constructed, based on a geometrical series. PMID:24333697
Pihlajaniemi, Ville; Sipponen, Satu; Sipponen, Mika H; Pastinen, Ossi; Laakso, Simo
Rapid increase in hepatic HMG CoA reductase activity and in vivo cholesterol synthesis after Triton WR 1339 injection1
Triton WR 1339, injected intravenously into rats, caused a 12% decrease in hepatic cholesterol within 30 minutes and a 34% decrease after 2 hours. An early and progressive increase in plasma cholesterol and tri- glycerides was also confirmed. Although hepatic HMG CoA reductase activity was unchanged after 30 minutes, it had increased seven-fold after 105 minutes. In vivo cho- lesterol
Stanley Goldfarb
HMG CoA Reductase Inhibitor-Induced Myotoxicity: Pravastatin and Lovastatin Inhibit the Geranylgeranylation of Low-Molecular-Weight Proteins in Neonatal Rat Muscle Cell Culture
In previous studies, inhibition of cholesterol synthesis by HMG CoA reductase inhibitors (HMGRI) was associated with myotoxicity in cultures of neonatal rat skeletal myotubes, and rhabdomyolysis in rats, rabbits, and humansin vivo. In vitromyotoxicity was directly related to HMGRI-induced depletion of mevalonate, farnesol, and geranylgeraniol, since supplementation with these intermediate metabolites abrogated the toxicity. Both farnesol and geranylgeraniol are required
Oliver P. Flint; Barbara A. Masters; Richard E. Gregg; Stephen K. Durham
Review: Enzymatic Hydrolysis of Cellulosic Biomass
Biological conversion of cellulosic biomass to fuels and chemicals offers the high yields to products vital to economic success and the potential for very low costs. Enzymatic hydrolysis that converts lignocellulosic biomass to fermentable sugars may be the most complex step in this process due to substrate-related and enzyme-related effects and their interactions. Although enzymatic hydrolysis offers the potential for higher yields, higher selectivity, lower energy costs, and milder operating conditions than chemical processes, the mechanism of enzymatic hydrolysis and the relationship between the substrate structure and function of various glycosyl hydrolase components are not well understood. Consequently, limited success has been realized in maximizing sugar yields at very low cost. This review highlights literature on the impact of key substrate and enzyme features that influence performance to better understand fundamental strategies to advance enzymatic hydrolysis of cellulosic biomass for biological conversion to fuels and chemicals. Topics are summarized from a practical point of view including characteristics of cellulose (e.g., crystallinity, degree of polymerization, and accessible surface area) and soluble and insoluble biomass components (e.g., oligomeric xylan, lignin, etc.) released in pretreatment, and their effects on the effectiveness of enzymatic hydrolysis. We further discuss the diversity, stability, and activity of individual enzymes and their synergistic effects in deconstructing complex lignocellulosic biomass. Advanced technologies to discover and characterize novel enzymes and to improve enzyme characteristics by mutagenesis, post-translational modification, and over-expression of selected enzymes and modifications in lignocellulosic biomass are also discussed.
Yang, Bin; Dai, Ziyu; Ding, Shi-You; Wyman, Charles E.
QSAR and Molecular Docking Studies of Oxadiazole-Ligated Pyrrole Derivatives as Enoyl-ACP (CoA) Reductase Inhibitors
A quantitative structure-activity relationship model was developed on a series of compounds containing oxadiazole-ligated pyrrole pharmacophore to identify key structural fragments required for anti-tubercular activity. Two-dimensional (2D) and three-dimensional (3D) QSAR studies were performed using multiple linear regression (MLR) analysis and k-nearest neighbour molecular field analysis (kNN-MFA), respectively. The developed QSAR models were found to be statistically significant with respect to training, cross-validation, and external validation. New chemical entities (NCEs) were designed based on the results of the 2D- and 3D-QSAR. NCEs were subjected to Lipinski�s screen to ensure the drug-like pharmacokinetic profile of the designed compounds in order to improve their bioavailability. Also, the binding ability of the NCEs with enoyl-ACP (CoA) reductase was assessed by docking.
Asgaonkar, Kalyani D.; Mote, Ganesh D.; Chitre, Trupti S.
Non-histone lysine acetylated proteins in heart failure.
Both histone-acetylations and histone deacetylases have been shown to play a key role in cardiac remodeling. Recently, it has become abundantly clear that many non-histone proteins are modified by post-translational lysine acetylations and that these acetylations regulate protein activity, conformation, and binding. In the present study, non-histone acetylated proteins associated with heart failure were identified. Global screening for lysine acetylated proteins was performed using 2-dimensional gel electrophoresis coupled with immunoblotting with a primary monoclonal anti-acetyl-lysine antibody. Lysine acetylated proteins were compared in two rodent models of hypertensive heart failure, the Dahl salt-sensitive (SS) and spontaneously hypertensive heart failure prone (SHHF) rats with those in corresponding controls, i.e., the Dahl salt-resistant (SR) and W (W) rat strains, respectively. Forty-one and 66 acetylated proteins were detected in SS and SHHF failing hearts, respectively, but either not detected or detected with less abundance in corresponding control hearts. Twelve of these acetylated proteins were common to both models of heart failure. These were identified using matrix-assisted laser desorption/ionization time of flight (MALDI-TOF/TOF) mass spectrometry followed by Mascot Analysis and included mitochondrial enzymes: ATP synthase, long-chain acyl-CoA dehydrogenase, creatine kinase, malate dehydrogenase, and pyruvate dehydrogenase. The abundance of NAD-dependent deacetylase sirtuin-3 (Sirt3), a mitochondrial deacetylase was reduced in SS and SHHF failing hearts. This is the first description of non-histone protein acetylations associated with heart failure and raises the prospect that acetylations of mitochondrial proteins linked to reduced Sirt3 mediate, in part, metabolic changes in heart failure. PMID:22155497
Grillon, Jean Michel; Johnson, Keven R; Kotlo, Kumar; Danziger, Robert S
Hydrolysis and heat treatment of aluminum dust.
Aluminum dust is a toxic and hazardous byproduct of Al remelting. The present research was performed to characterize and evaluate its behavior in water. The materials obtained by hydrolysis were also characterized, and the gases generated during the process were qualitatively analyzed. The effects of hydrolysis reaction time and temperature on the dust were also explored. The hydrolysis of Al dust is an exothermic reaction that gave rise to a solid composed of aluminum oxide, silicon oxide, and spinel (MgAl2O4). Most of the CH4, NH3, and SH2 gases generated were emitted immediately upon the start of the reaction, though their production continued for a long time. This slow reaction, which was moderately accelerated by temperature, led to the formation of a material less reactive than the untreated dust. On the other hand, heat treatment of the dust gave rise to an inert material composed of spinel, alumina, and magnesium and aluminum silicates. PMID:11417682
L�pez, F A; Pe�a, M C; L�pez-Delgado, A
Structure of a ternary Naa50p (NAT5/SAN) N-terminal acetyltransferase complex reveals the molecular basis for substrate-specific acetylation.
The co-translational modification of N-terminal acetylation is ubiquitous among eukaryotes and has been reported to have a wide range of biological effects. The human N-terminal acetyltransferase (NAT) Naa50p (NAT5/SAN) acetylates the ?-amino group of proteins containing an N-terminal methionine residue and is essential for proper sister chromatid cohesion and chromosome condensation. The elevated activity of NATs has also been correlated with cancer, making these enzymes attractive therapeutic targets. We report the x-ray crystal structure of Naa50p bound to a native substrate peptide fragment and CoA. We found that the peptide backbone of the substrate is anchored to the protein through a series of backbone hydrogen bonds with the first methionine residue specified through multiple van der Waals contacts, together creating an ?-amino methionine-specific pocket. We also employed structure-based mutagenesis; the results support the importance of the ?-amino methionine-specific pocket of Naa50p and are consistent with the proposal that conserved histidine and tyrosine residues play important catalytic roles. Superposition of the ternary Naa50p complex with the peptide-bound Gcn5 histone acetyltransferase revealed that the two enzymes share a Gcn5-related N-acetyltransferase fold but differ in their respective substrate-binding grooves such that Naa50p can accommodate only an ?-amino substrate and not a side chain lysine substrate that is acetylated by lysine acetyltransferase enzymes such as Gcn5. The structure of the ternary Naa50p complex also provides the first molecular scaffold for the design of NAT-specific small molecule inhibitors with possible therapeutic applications. PMID:21900231
Liszczak, Glen; Arnesen, Thomas; Marmorstein, Ronen
Hydrolysis of iodine: equilibria at high temperatures
The hydrolysis (or disproportionation) of molecular iodine to form iodate and iodide ions has been studied by emf measurements over the temperature range, 3.8/sup 0/ to 209.0/sup 0/C. The interpretation of these results required a knowledge of the formation constant for triiodide ion and the acid dissociation constant of iodic acid, both of which were measured as a function of temperature. The resulting thermodynamic data have been incorporated into a general computer model describing the hydrolysis equilibria of iodine as a function of initial concentration, pH and temperature.
Palmer, D.A.; Ramette, R.W.; Mesmer, R.E.
Sorption of spruce O -acetylated galactoglucomannans onto different pulp fibres
Sorption of spruce acetylated galactoglucomannans (GGM) onto different pulps, among which unbleached and peroxide-bleached mechanical pulps, and unbleached and bleached kraft (BK) pulps, was studied as a means of understanding the retention of acetylated GGMs in mechanical pulping and papermaking. The fibre surface coverage of lignin and carbohydrates was estimated by X-ray photoelectron spectroscopy (XPS) or electron spectroscopy for chemical
Tea Hannuksela; Pedro Fardim; Bjarne Holmbom
The biology of lysine acetylation integrates transcriptional programming and metabolism
The biochemical landscape of lysine acetylation has expanded from a small number of proteins in the nucleus to a multitude of proteins in the cytoplasm. Since the first report confirming acetylation of the tumor suppressor protein p53 by a lysine acetyltransferase (KAT), there has been a surge in the identification of new, non-histone targets of KATs. Added to the known
Jigneshkumar Patel; Ravi R Pathak; Shiraz Mujtaba
Globin Intergenic Transcription and Histone Acetylation Dependent on an Enhancer
Histone acetyltransferases are associated with the elongating RNA polymerase II (Pol II) complex, support- ing the idea that histone acetylation and transcription are intertwined mechanistically in gene coding se- quences. Here, we studied the establishment and function of histone acetylation and transcription in noncoding sequences by using a model locus linking the -globin HS2 enhancer and the embryonic -globin gene
A. Kim; H. Zhao; I. Ifrim; A. Dean
Protein lysine acetylation guards metabolic homeostasis to fight against cancer.
Properly coordinated metabolism and maintained metabolite homeostasis are important because altered metabolite homeostasis has a causal role in many human diseases, including cancer. Metabolite homeostasis is maintained by fine-tuned coordination of metabolite generation and utilization. Metabolite deregulation has recently been shown to alter the signaling pathways and reprogram epigenetic factors associated with tumorigenesis. Protein lysine acetylation is emerging as a metabolism-coordinating mechanism. Mechanistic studies have shown that acetylation may have roles in nutrient adaptation and in maintaining metabolite homeostasis by exerting regulatory effects on metabolic enzymes, metabolic pathways and metabolic networks. Here we review recent progress in the determination of the role of acetylation regulation in metabolism coordination. In particular, we review links between deregulated acetylation in metabolic enzymes and tumorigenesis. We further hypothesize on applications of the mediation of acetylation to restore deregulated metabolism coordination and thus develop novel means of cancer treatment. PMID:23665675
Xu, W; Li, Y; Liu, C; Zhao, S
Self-scrolling ability of differentially acetylated chitosan film.
Chitosan film cast on a glass slide was exposed to acetic anhydride vapor, resulting in an acetylation gradient in the film, with preferential acetylation of the exposed surface. The difference in degree of acetylation between the two surfaces of the peeled film was confirmed by attenuated total reflection infrared spectroscopy. Upon immersion of the film in water, differential swelling occurred because the more highly acetylated surface absorbed less water, and the resulting bending moment caused self-scrolling with the more highly acetylated surface inside. Simultaneous peeling and scrolling of films with a thickness of micrometer order, using dilute aqueous hydrofluoric acid, afforded tightly rolled chitosan microtubes. This simple self-scrolling mechanism is potentially applicable for micro-scale design with various naturally occurring polymers. PMID:24815399
Saito, Yukie; Luchnikov, Valeriy; Inaba, Ayano; Tamura, Katsuhito
Cell biology (Communication arising): Tubulin acetylation and cell motility
NASA Astrophysics Data System (ADS)
Although the protein tubulin is known to undergo several post-translational modifications that accumulate in stable but not dynamic microtubules inside cells, the function of these modifications is unknown. Hubbert et al. have shown that the enzyme HDAC6 (for histone deacetylase 6) reverses the post-translational acetylation of tubulin, and provide evidence that reducing tubulin acetylation enhances cell motility. They also suggest that decreasing tubulin acetylation reduces microtubule stability. However, we find that microtubule stabilization is not promoted by tubulin acetylation. We conclude that the alteration in cell motility observed by Hubbert et al. in cells overexpressing HDAC6 results not from changes in the formation of stable microtubules, but from alterations in the degree of tubulin acetylation.
Palazzo, Alexander; Ackerman, Brian; Gundersen, Gregg G.
Formation of the thioester, N-acetyl, S-lactoylcysteine, by reaction of N-acetylcysteine with pyruvaldehyde in aqueous solution. [in prebiotic evolution
NASA Technical Reports Server (NTRS)
N-acetylcysteine reacts efficiently with pyruvaldehyde (methylglyoxal) in aqueous solution (pH 7.0) in the presence of a weak base, like imidazole or phosphate, to give the thioester, N-acetyl, S-lactoylcysteine. Reactions of 100 mM N-acetylcysteine with 14 mM, 24 mM and 41 mM pyruvaldehyde yield, respectively, 86%, 76% and 59% N-acetyl, S-lactoylcysteine based on pyruvaldehyde. The decrease in the percent yield at higher pyruvaldehyde concentrations suggests that during its formation the thioester is not only consumed by hydrolysis, but also by reaction with some substance in the pyruvaldehyde preparation. Indeed, purified N-acetyl, S-lactoylcysteine disappears much more rapidly in the presence of pyruvaldehyde than in its absence. Presumably, N-acetyl, S-lactoylcysteine synthesis occurs by rearrangement of the hemithioacetal of N-acetylcysteine and pyruvaldehyde. The significance of this pathway of thioester formation to molecular evolution is discussed.
Weber, A. L.
Semi-synthetic preparation of 1-O-(1'-/sup 14/C)hexadecyl-2-acetyl-sn-glycero-3-phosphocholine (platelet activating factor) using plant cell cultures
Incubation of photomixotrophic cell suspension cultures of rape (Brassica napus) and heterotrophic cell suspension cultures of soya (Glycine max) with 1-O-(1'-/sup 14/C)hexadecyl-sn-glycerol or rac-1-O-(1'-/sup 14/C)hexadecylglycerol leads in high yield (up to 78%) to labeled 1-O-hexadecyl-2-acyl-sn-glycero-3-phosphocholines. Alkaline hydrolysis of the choline glycerophospholipids yields pure 1-O-(1'-/sup 14/C)hexadecyl-sn-glycero-3-phosphocholine. 1-O-(1'-14C)Hexadecyl-2-acetyl-sn-glycero-3-phosphocholine (platelet activating factor) is obtained by acetylating the lyso compound. The semi-synthetic preparation described leads to labeled platelet activating factor in an overall yield of 50-60% without loss of specific activity.
Weber, N.; Mangold, H.K.
Cellulose hydrolysis in subcritical and supercritical water
In this paper we propose a new method to hydrolyze cellulose rapidly in supercritical water (SCW) to recover glucose, fructose and oligomers (cellobiose, cellotriose, cellotetraose, etc.). Cellulose decomposition experiments were conducted with a flow type reactor in the range of temperature from 290 to 400�C at 25MPa. A high pressure slurry feeder was developed to feed the cellulose�water slurries. Hydrolysis
Mitsuru Sasaki; Bernard Kabyemela; Roberto Malaluan; Satoshi Hirose; Naoko Takeda; Tadafumi Adschiri; Kunio Arai
Thioglycoside hydrolysis catalyzed by {beta}-glucosidase
Sweet almond {beta}-glucosidase (EC 220.127.116.11) has been shown to have significant thioglycohydrolase activity. While the K{sub m} values for the S- and O-glycosides are similar, the k{sub cat} values are about 1000-times lower for the S-glycosides. Remarkably, the pH-profile for k{sub cat}/K{sub m} for hydrolysis of p-nitrophenyl thioglucoside (pNPSG) shows the identical dependence on a deprotonated carboxylate (pK{sub a} 4.5) and a protonated group (pK{sub a} 6.7) as does the pH-profile for hydrolysis of the corresponding O-glycoside. Not surprisingly, in spite of the requirement for the presence of this protonated group in catalytically active {beta}-glucosidase, thioglucoside hydrolysis does not involve general acid catalysis. There is no solvent kinetic isotope effect on the enzyme-catalyzed hydrolysis of pNPSG.
Shen Hong [Department of Chemistry, Tulane University, New Orleans, LA 70118 (United States); Byers, Larry D. [Department of Chemistry, Tulane University, New Orleans, LA 70118 (United States)], E-mail: firstname.lastname@example.com
HD Hydrolysis/Biodegradation Toxicology and Kinetics.
National Technical Information Service (NTIS)
One technology developed for potential use in the disposal of RD (2,2'-dichlorodiethyl sulfide) from the Aberdeen Proving Ground unitary chemical stockpile is hydrolysis followed by biodegradation. Stockpile RD (89.2% pure) was hydrolyzed in 90 deg C wate...
S. P. Harvey L. L. Szafraniec W. T. Beaudry M. V. Haley T. E. Rosso
Hippurate Hydrolysis in Klebsiella-Cloaca Classification
SUMMARY: 169 strains of Klebsiella pneumoniae and 68 strains of Cloaca cloacae were used in an examination of Hajna & Damon's hippurate test and various modi- fications of it. The addition of a pH indicator (phenol red) to the medium enabled hydrolysis to be detected by a change of colour. Clear-cut distinction between K. pneumoniae and C. cloacae was not
The program RATE was developed to aid in the extrapolation and interpretation of hydrolysis rate data to a format that is useful for environmental risk assessment. ydrolysis data typically are reported in the literature as pseudo-first-order rate constants at the temperature and ...
Microstructure of coke briquettes from hydrolysis lignin
The microstructure of pyrolyzed briquettes prepared from hydrolyzed lignins at different final heating temperatures has been investigated. The pore structure of the briquettes of hydrolysis lignins before the thermal treatment is dependent to a large extent upon the granulometric composition of the raw material. The initial structure of the lignins also played an important part on the effect of the
B. N. Zhitov; Yu. G. Korolev; G. N. Makarov; A. M. Myasoedov; I. A. Shimanskaya; V. P. Okladnikov
Optimization of dilute acid hydrolysis of Enteromorpha
NASA Astrophysics Data System (ADS)
Acid hydrolysis is a simple and direct way to hydrolyze polysaccharides in biomass into fermentable sugars. To produce fermentable sugars effectively and economically for fuel ethanol, we have investigated the hydrolysis of Enteromorpha using acids that are typically used to hydrolyze biomass: H2SO4, HCl, H3PO4 and C4H4O4 (maleic acid). 5%(w/w) Enteromorpha biomass was treated for different times (30, 60, and 90 min) and with different acid concentrations (0.6, 1.0, 1.4, 1.8, and 2.2%, w/w) at 121�C. H2SO4 was the most effective acid in this experiment. We then analyzed the hydrolysis process in H2SO4 in detail using high performance liquid chromatography. At a sulfuric acid concentration of 1.8% and treatment time of 60 min, the yield of ethanol fermentable sugars (glucose and xylose) was high, (230.5 mg/g dry biomass, comprising 175.2 mg/g glucose and 55.3 mg/g xylose), with 48.6% of total reducing sugars being ethanol fermentable. Therefore, Enteromorpha could be a good candidate for production of fuel ethanol. In future work, the effects of temperature and biomass concentration on hydrolysis, and also the fermentation of the hydrolysates to ethanol fuel should be focused on.
Feng, Dawei; Liu, Haiyan; Li, Fuchao; Jiang, Peng; Qin, Song
Mechanisms of lactone hydrolysis in acidic conditions.
The acid-catalyzed hydrolysis of linear esters and lactones was studied using a hybrid supermolecule-polarizable continuum model (PCM) approach including up to six water molecules. The compounds studied included two linear esters, four ?-lactones, two ?-lactones, and one ?-lactone: ethyl acetate, methyl formate, ?-propiolactone, ?-butyrolactone, ?-isovalerolactone, diketene (4-methyleneoxetan-2-one), ?-butyrolactone, 2(5H)-furanone, and ?-valerolactone. The theoretical results are in good quantitative agreement with the experimental measurements reported in the literature and also in excellent qualitative agreement with long-held views regarding the nature of the hydrolysis mechanisms at molecular level. The present results help to understand the balance between the unimolecular (A(AC)1) and bimolecular (A(AC)2) reaction pathways. In contrast to the experimental setting, where one of the two branches is often occluded by the requirement of rather extreme experimental conditions, we have been able to estimate both contributions for all the compounds studied and found that a transition from A(AC)2 to A(AC)1 hydrolysis takes place as acidity increases. A parallel work addresses the neutral and base-catalyzed hydrolysis of lactones. PMID:23731203
G�mez-Bombarelli, Rafael; Calle, Emilio; Casado, Julio
Monitoring enzymatic ATP hydrolysis by EPR spectroscopy.
An adenosine triphosphate (ATP) analogue modified with two nitroxide radicals is developed and employed to study its enzymatic hydrolysis by electron paramagnetic resonance spectroscopy. For this application, we demonstrate that EPR holds the potential to complement fluorogenic substrate analogues in monitoring enzymatic activity. PMID:24872080
Hacker, Stephan M; Hintze, Christian; Marx, Andreas; Drescher, Malte
Hydrolysis of ionic cellulose to glucose.
Hydrolysis of ionic cellulose (IC), 1,3-dimethylimidazolium cellulose phosphite, which could be synthesized from cellulose and dimethylimidazolium methylphosphite ([Dmim][(OCH3)(H)PO2]) ionic liquid, was conducted for the synthesis of glucose. The reaction without catalysts at 150�C for 12h produced glucose with 14.6% yield. To increase the hydrolysis yield, various acid catalysts were used, in which the sulfonated active carbon (AC-SO3H) performed the best catalytic activity in the IC hydrolysis. In the presence of AC-SO3H, the yields of glucose reached 42.4% and 53.9% at the reaction condition of 150�C for 12h and 180�C for 1.5h, respectively; however the yield decreased with longer reaction time due to the degradation of glucose. Consecutive catalyst reuse experiments on the IC hydrolysis demonstrated the catalytic activity of AC-SO3H persisted at least through four successive uses. PMID:25011079
Vo, Huyen Thanh; Widyaya, Vania Tanda; Jae, Jungho; Kim, Hoon Sik; Lee, Hyunjoo
Hydrolysis of glycocholic acid by fungi
A range of fungi have been investigated for their ability to hydrolyse glycocholic acid to yield cholic acid. Using thin-layer chromatography the majority of fungi tested have been shown to possess this ability. In the case of Cochliobolus intermedius IMI 52980 the product of hydrolysis has been isolated and characterised as cholic acid. The same organism has been demonstrated to
I. So Maddox; R. Chong
Coupled Ferredoxin and Crotonyl Coenzyme A (CoA) Reduction with NADH Catalyzed by the Butyryl-CoA Dehydrogenase\\/Etf Complex from Clostridium kluyveri
Cell extracts of butyrate-forming clostridia have been shown to catalyze acetyl-coenzyme A (acetyl-CoA)- and ferredoxin-dependent formation of H2 from NADH. It has been proposed that these bacteria contain an NADH:ferredoxin oxidoreductase which is allosterically regulated by acetyl-CoA. We report here that ferre- doxin reduction with NADH in cell extracts from Clostridium kluyveri is catalyzed by the butyryl-CoA dehy- drogenase\\/Etf complex
Fuli Li; Julia Hinderberger; Henning Seedorf; Jin Zhang; Wolfgang Buckel; Rudolf K. Thauer
Kinetic Study of Hydrolysis of Polyester Elastomer in Magnetic Tape.
National Technical Information Service (NTIS)
A useful method for kinetic study of the hydrolysis of polyester elastomer is established which uses the number-average molecular weight. The reasonableness of this method is confirmed and the effect of magnetic particles on hydrolysis is considered.
K. Yamamoto H. Watanabe
Rate constants for hydrolysis in water of ten classes of organic compounds are examined with the objective of establishing new, or expanding existing, property reactivity correlations. These relationships then can be used to predict the environmental hydrolysis of chemicals that ...
A Bacterial Indole3-acetyl-L-aspartic Acid Hydrolase Inhibits Mung Bean ( Vigna radiata L.) Seed Germination Through Arginine-rich Intracellular Delivery
Indole-3-acetyl-L-aspartic acid (IAA-Asp) is a natural product in many plant species and plays many important roles in auxin\\u000a metabolism and plant physiology. IAA-Asp hydrolysis activity is, therefore, believed to affect plant physiology through changes\\u000a in IAA metabolism in plants. We applied a newly discovered technique, arginine-rich intracellular delivery (AID), to deliver\\u000a a bacterial IAA-Asp hydrolase into cells of mung bean
Kevin Liu; Han-Jung Lee; Sio San Leong; Chen-Lun Liu; Jyh-Ching Chou
Removing Hemicellulose from Pulps by Specific Enzymic Hydrolysis
The hemicellulose content (solubility in 18% NaOH) of a delignified mechanical aspen pulp was lowered from 23.4% to 18.2% by one-hour hydrolysis with xylanase isolated from the fungus Schizophpllum commune by fractional precipitation. After 24 h hydrolysis, the hemicellulose content was reduced further to 12.9%. The predominant hydrolysis products, xylose and xylobiose, confirmed the specificity of hydrolysis. A crude mixture
M. G. Paice; L. Jurasek
Acetylation in Nuclear Receptor Signaling and the Role of Sirtuins
It has been known since the early 1970s that nuclear receptor complexes bind DNA in association with coregulatory proteins. Characterization of these nuclear receptor coregulators has revealed diverse enzymatic activities that temporally and spatially coordinate nuclear receptor activity within the context of local chromatin in response to diverse hormone signals. Chromatin-modifying proteins, which dictate the higher-order chromatin structure in which DNA is packaged, in turn orchestrate orderly recruitment of nuclear receptor complexes. Modifications of histones include acetylation, methylation, phosphorylation, ubiquitylation, sumoylation, ADP ribosylation, deimination, and proline isomerization. At this time, we understand how a subset of these modifications regulates nuclear receptor signaling. However, the effects, particularly of acetylation and demethylation, are profound. The finding that nuclear receptors are directly acetylated and that acetylation in turn directly regulates contact-independent growth has broad therapeutic implications. Studies over the past 7 yr have led to the understanding that nuclear receptor acetylation is a conserved function, regulating diverse nuclear receptor activity. Furthermore, we now know that acetylation of multiple and distinct substrates within nuclear receptor signaling pathways, form an acetylation signaling network from the cell surface to the nucleus. The finding that nicotinamide adenine dinucleotide (NAD)-dependent histone deacetylases, the sirtuins, are capable of deacetylating nuclear receptors provides a new level of complexity in the control of nuclear receptor activity in which local intracellular concentrations of NAD may regulate nuclear receptor physiology.
Wang, Chenguang; Powell, Michael J.; Popov, Vladimir M.; Pestell, Richard G.
Regulators of Cellular Levels of Histone Acetylation in Saccharomyces cerevisiae
Histone acetylation levels are regulated through the opposing activities of histone acetyltransferases (HATs) and deacetylases (HDACs). While much is known about gene-specific control of histone acetylation, little is understood about how total or cellular levels of histone acetylation are regulated. To identify regulators of cellular levels of histone acetylation, we developed an immunofluorescence-based approach to screen the single-gene deletion library of Saccharomyces cerevisiae for strains with significant reductions in cellular histone acetylation levels. Of the 4848 mutants screened, we identified 63 strains with considerable cellular hypoacetylation of N-terminal lysines in histones H3 and H4. The cellular hypoacetylation was validated for subsets of the identified strains through secondary screens including mass spectrometric analysis of individual lysines and chromatin immunoprecipitation of specific genomic loci. Among the identified mutants were several members of the Ccr4-Not complex, V-type ATPases, and vacuolar protein-sorting complexes as well as genes with unknown functions. We show that Gcn5, a major HAT in yeast, has diminished histone acetyltransferase activity in particular mutants, providing a plausible explanation for reduction of cellular acetylation levels in vivo. Our findings have revealed unexpected and novel links between histone acetylation, Gcn5 HAT activity, and diverse processes such as transcription, cellular ion homeostasis, and protein transport.
Peng, Weimin; Togawa, Cynthia; Zhang, Kangling; Kurdistani, Siavash K.
Hydrolysis of xylans by enzyme systems from solid cultures of Trichoderma harzianum strains.
Xylanase activity was isolated from crude extracts of Trichoderma harzianum strains C and 4 grown at 28 degree C in a solid medium containing wheat bran as the carbon source. Enzyme activity was demonstrable in the permeate after ultrafiltration of the crude extracts using an Amicon system. The hydrolysis patterns of different xylans and paper pulps by xylanase activity ranged from xylose, xylobiose and xylotriose to higher xylooligosaccharides. A purified ss-xylosidase from the Trichoderma harzianum strain released xylose, xylobiose and xylotriose from seaweed, deacetylated, oat spelt and birchwood xylans. The purified enzyme was not active against acetylated xylan and catalyzed the hydrolysis of xylooligosaccharides, including xylotriose, xylotetraose and xylopentaose. However, the enzyme was not able to degrade xylohexaose. Xylanase pretreatment was effective for hardwood kraft pulp bleaching. Hardwood kraft pulp bleached in the XEOP sequence had its kappa number reduced from 13.2 to 8.9 and a viscosity of 20. 45 cp. The efficiency of delignification was 33%. PMID:10454755
Silveira, F Q; Ximenes, F A; Cacais, A O; Milagres, A M; Medeiros, C L; Puls, J; Filho, E X
Enzymatic hydrolysis of rawhide using papain and neutrase
Rawhide split was hydrolysed separately by two proteolytic enzymes, papain and neutrase. The effects of enzymatic conditions of the hydrolysis reaction were investigated. During the first 10min of the enzymatic hydrolysis, the yield of the hydrolysed protein increased sharply, then it slowly increased or became essentially constant due to the limited availability of the substrate. The optimum hydrolysis conditions of
Siriporn Damrongsakkul; Kongpob Ratanathammapan; Kittinan Komolpis; Wiwut Tanthapanichakoon
Technical bases for precipitate hydrolysis process operating parameters
This report provides the experimental data and rationale in support of the operating parameters for precipitate hydrolysis specified in WSRC-RP-92737. The report is divided into two sections, the first dealing with lab-scale precipitate hydrolysis experimentation while the second part addresses large-scale runs conducted to demonstrate the revised operating parameters in the Precipitate Hydrolysis Experimental Facility (PHEF).
Bannochie, C.J.
The first thermophilic alpha-oxoamine synthase family enzyme that has activities of 2-amino-3-ketobutyrate CoA ligase and 7-keto-8-aminopelargonic acid synthase: cloning and overexpression of the gene from an extreme thermophile, Thermus thermophilus, and characterization of its gene product.
The first thermophilic alpha-oxoamine synthase family enzyme was identified. The gene (ORF TTHA1582), which is annotated to code putative alpha-oxoamine synthase family enzymes, 7-keto-8-aminopelargonic acid (KAPA) synthase (BioF, 8-amino-7-oxononanoate synthase, EC 18.104.22.168) and 2-amino-3-ketobutyrate CoA ligase (KBL, EC 220.127.116.11), in a genomic database, was cloned from an extreme thermophile, Thermus thermophilus, and overexpressed in Escherichia coli. The recombinant TTHA1582 protein was purified and characterized. It exhibited activity of BioF, which catalyzes the condensation of pimeloyl-CoA and L-alanine to produce a biotin intermediate KAPA, CoASH, and CO(2) with pyridoxal 5'-phosphate as a cofactor. The protein is a dimer with a subunit of 43 kDa that shows an amino acid sequence identity of 35% with E. coli BioF. The optimum temperature and pH were about 70 degrees C and about 6.0. The enzyme showed high thermostability at temperatures of up to 70 degrees C for 1 h, and a half-life of 1 h at 80 degrees C. Thus the TTHA1582 protein was found to have the highest optimum temperature and thermostablility of the alpha-oxoamine synthase family enzymes so far reported. Substrate specificity experiments revealed that it was also able to catalyze the KBL reaction, which used acetyl-CoA and glycine as substrates, and that enzyme activity was seen with the following combinations of substrates: acetyl-CoA and glycine, L-alanine, or L-serine; pimeloyl-CoA and L-alanine, glycine, or L-serine; palmitoyl-CoA and L-alanine. This suggests that the recombinant TTHA1582 protein has broad substrate specificity, unlike the reported mesophilic enzymes of the alpha-oxoamine synthase family. PMID:18071260
Kubota, Takaaki; Shimono, Jyunpei; Kanameda, Chie; Izumi, Yoshikazu
Protein N-terminal acetylation: NAT 2007-2008 Symposia.
Protein N-terminal acetylation is a very common modification, but has during the past decades received relatively little attention. In order to put this neglected field back on the scientific map, we have in May 2007 and September 2008 arranged two international NAT symposia in Bergen, Norway. This supplement contains selected proceedings from these symposia reflecting the current status of the field, including an overview of protein N-terminal acetylation in yeast and humans, a novel nomenclature system for the N-terminal acetyltransferases (NATs) and methods for studying protein N-terminal acetylation in vitro and in vivo. PMID:19660094
Arnesen, Thomas
Protein N-terminal acetylation: NAT 2007-2008 Symposia
Protein N-terminal acetylation is a very common modification, but has during the past decades received relatively little attention. In order to put this neglected field back on the scientific map, we have in May 2007 and September 2008 arranged two international NAT symposia in Bergen, Norway. This supplement contains selected proceedings from these symposia reflecting the current status of the field, including an overview of protein N-terminal acetylation in yeast and humans, a novel nomenclature system for the N-terminal acetyltransferases (NATs) and methods for studying protein N-terminal acetylation in vitro and in vivo.
Arnesen, Thomas
Obesity, cancer, and acetyl-CoA metabolism
As rates of obesity soar in the Unites States and around the world, cancer attributed to obesity has emerged as major threat to public health. The link between obesity and cancer can be attributed in part to the state of chronic inflammation that develops in obesity. Acetyl-CoA production and protein acetylation patterns are highly sensitive to metabolic state and are significantly altered in obesity. In this article, we explore the potential role of nutrient-sensitive lysine acetylation in regulating inflammatory processes in obesity-linked cancer.
Lee, Joyce V.; Shah, Supriya A.; Wellen, Kathryn E.
Studies of Human 2,4-Dienoyl CoA Reductase Shed New Light on Peroxisomal ?-Oxidation of Unsaturated Fatty Acids
Peroxisomes play an essential role in maintaining fatty acid homeostasis. Although mitochondria are also known to participate in the catabolism of fatty acids via ?-oxidation, differences exist between the peroxisomal and mitochondrial ?-oxidation. Only peroxisomes, but not mitochondrion, can shorten very long chain fatty acids. Here, we describe the crystal structure of a ternary complex of peroxisomal 2,4-dienoyl CoA reductases (pDCR) with hexadienoyl CoA and NADP, as a prototype for comparison with the mitochondrial 2,4-dienoyl CoA reductase (mDCR) to shed light on the differences between the enzymes from the two organelles at the molecular level. Unexpectedly, the structure of pDCR refined to 1.84 � resolution reveals the absence of the tyrosine-serine pair seen in the active site of mDCR, which together with a lysine and an asparagine have been deemed a hallmark of the SDR family of enzymes. Instead, aspartate hydrogen-bonded to the C? hydroxyl via a water molecule seems to perturb the water molecule for protonation of the substrate. Our studies provide the first structural evidence for participation of water in the DCR-catalyzed reactions. Biochemical studies and structural analysis suggest that pDCRs can catalyze the shortening of six-carbon-long substrates in vitro. However, the Km values of pDCR for short chain acyl CoAs are at least 6-fold higher than those for substrates with 10 or more aliphatic carbons. Unlike mDCR, hinge movements permit pDCR to process very long chain polyunsaturated fatty acids.
Hua, Tian; Wu, Dong; Ding, Wei; Wang, Jiangyun; Shaw, Neil; Liu, Zhi-Jie (Nankai) [Nankai; (Chinese Aca. Sci.)
STAT5 activators modulate acyl CoA oxidase (AOX) expression in adipocytes and STAT5A binds to the AOX promoter in vitro
Growth hormone (GH) diminishes adipose tissue mass in vivo and prolactin (PRL) can also modulate adipocyte metabolism. Both GH and PRL are potent activators of STAT5 and exert a variety of effects on adipocyte gene expression. In this study, we have demonstrated that GH and PRL increase the mRNA of acyl CoA oxidase in 3T3-L1 adipocytes. We also identified seven
Ann A. Coulter; Jacqueline M. Stephens
C75 is converted to C75CoA in the hypothalamus, where it inhibits carnitine palmitoyltransferase 1 and decreases food intake and body weight
Central nervous system administration of C75 produces hypophagia and weight loss in rodents identifying C75 as a potential drug against obesity and type 2 diabetes. However, the mechanism underlying this effect is unknown. Here we show that C75-CoA is generated chemically, in vitro and in vivo from C75 and that it is a potent inhibitor of carnitine palmitoyltranferase 1 (CPT1),
Paula Mera; Assia Bentebibel; Eduardo Lopez-Vinasb; Antonio G. Cordente; Chandrashekaran Gurunathan; David Sebasti�n; Irene V�zquez; Laura Herrero; Xavier Ariza; Paulino G�mez-Puertas; Guillermina Asins; Dolors Serra; Jordi Garc�a; Fausto G. Hegardt
Valence-band structure of the skutterudite compounds CoAs3, CoSb3, and RhSb3 studied by x-ray photoelectron spectroscopy
The valence-band structure of CoAs3, CoSb3, and RhSb3 with a skutterudite-type crystal structure has been investigated by x-ray photoelectron spectroscopy. The photoemission spectra are compared with recent density-of-states calculations. Our photoemission spectra results and theoretical results are in good agreement for the energy positions in the metal d states and the pnicogen p states, but relatively large differences are found
H. Anno; K. Matsubara; T. Caillat; J.-P. Fleurial
Neuroprotection in rabbit retina with N-acetyl-aspartylglutamate and 2-phosphonyl-methyl pentanedioic acid
NASA Astrophysics Data System (ADS)
Retinal tissue is subject to ischemia from diabetic retinopathy and other conditions that affect the retinal vasculature such as lupus erythematosus and temporal arteritis. There is evidence in animal models of reversible ischemia that a therapeutic window exists during early recovery when agents that reduce glutamate activity at its receptor sites can rescue neurons from injury. To model ischemia, we used sodium cyanide (NaCN), to inhibit oxidative metabolism, and 2-deoxyglucose (2-DG) to inhibit glycolysis. Dissociated rabbit retina cells were studied to evaluate the potential neuroprotective effects of N-acetyl-aspartyl-glutamate (MAAG), which competes with glutamate as a low-potency agonist at the NMDA receptor complex. N-acetylated ?-linked acidic dipeptidase (NAALADase; the NAAG-hydrolyzing enzyme) is responsible for the hydrolysis of NAAG into glutamate, a neurotransmitter and potent excitotoxin, and N-acetylaspartate. 2-Phosphonyl-methyl pentanedioic acid (PMPA) and ?-linked NAAG (?-NAAG), inhibitors of NAALADase, were also tested, since inhibition of NAALADase could reduce synaptic glutamate and increase the concentration of NAAG. We found that metabolic inhibition with NaCN/2-DG for 1 hour caused 50% toxicity as assessed with the MTT assay. Co-treatment with NAAG resulted in dose-dependent protection of up to 55% (p<0.005). When the non-hydrolyzable, NAALADase inhibitor ?-NAAG was employed dose-dependent protection of up to 37% was observed (p<0.001). PMPA also showed 48% protection (p<.05-.001) against these insults. These data suggest that NAAG may antagonize the effect of glutamate at the NMDA receptor complex in retina. Inhibition of NAALADase by PMPA and ?-NAAG may increase the activity of endogenous NAAG.
Hacker, Henry D.; Yourick, Debra L.; Koenig, Michael K.; Slusher, Barbara S.; Meyerhoff, James L.
Down-regulation of hydroxycinnamoyl CoA: shikimate hydroxycinnamoyl transferase in transgenic alfalfa affects lignification, development and forage quality.
The recently discovered enzyme hydroxycinnamoyl CoA: shikimate hydroxycinnamoyl transferase (HCT) catalyzes the reactions both immediately preceding and following the insertion of the 3-hydroxyl group into monolignol precursors. A number of independent transgenic lines of alfalfa (Medicago sativa L.) were generated in which the levels of HCT were reduced through antisense HCT expression under control of the bean PAL2 promoter which is preferentially expressed in vascular tissue. Reduction of enzyme activity in these lines was from at least 15-50%. The most severely down-regulated lines exhibited significant stunting, reduction of biomass and delayed flowering. HCT down-regulation resulted in strongly reduced lignin content and striking changes in lignin monomer composition, with predominant deposition of 4-hydroxyphenyl units in the lignin. Vascular structure was impaired in the most strongly down-regulated lines. Analysis of forage quality parameters showed strong reductions of neutral- and acid-detergent fiber in the down-regulated lines, in parallel with large increases (up to 20%) in dry matter forage digestibility. Although manipulation of lignin biosynthesis can greatly improve forage digestibility, accompanying effects on plant development need to be better understood. PMID:17466347
Shadle, Gail; Chen, Fang; Srinivasa Reddy, M S; Jackson, Lisa; Nakashima, Jin; Dixon, Richard A
Downregulation of Caffeic Acid 3-O-Methyltransferase and Caffeoyl CoA 3-O-Methyltransferase in Transgenic Alfalfa
Transgenic alfalfa plants were generated harboring caffeic acid 3-O-methyltransferase (COMT) and caffeoyl CoA 3-O-methyltransferase (CCOMT) cDNA sequences under control of the bean phenylalanine ammonia-lyase PAL2 promoter. Strong downregulation of COMT resulted in decreased lignin content, a reduction in total guaiacyl (G) lignin units, a near total loss of syringyl (S) units in monomeric and dimeric lignin degradation products, and appearance of low levels of 5-hydroxy guaiacyl units and a novel dimer. No soluble monolignol precursors accumulated. In contrast, strong downregulation of CCOMT led to reduced lignin levels, a reduction in G units without reduction in S units, and increases in ?-5 linked dimers of G units. Accumulation of soluble caffeic acid ?-d-glucoside occurred only in CCOMT downregulated plants. The results suggest that CCOMT does not significantly contribute to the 3-O-methylation step in S lignin biosynthesis in alfalfa and that there is redundancy with respect to the 3-O-methylation reaction of G lignin biosynthesis. COMT is unlikely to catalyze the in vivo methylation of caffeic acid during lignin biosynthesis.
Guo, Dianjing; Chen, Fang; Inoue, Kentaro; Blount, Jack W.; Dixon, Richard A.
Continuous steam hydrolysis of tulip poplar
To produce ethanol from hardwood it is desirable to fractionate the hardwood in order to produce a relatively pure cellulosic pulp for dilute acid hydrolysis. An experimental investigation of continuous steam hydrolysis of tulip poplar wood chips indicates that over 90% of the lignin present can be extracted by 0.1N sodium hydroxide, resulting in a cellulose pulp containing over 90% hexosan. The study was performed using a Stake Technology, Ltd., continuous digester rated at one oven dry ton per hour of wood chips. The yields of hexosans, hexoses, xylan, xylose, lignin, furfural, acetic acid and methanol were determined as a function of residence time and steam pressure in the digester. The information provides a basis for establishing a material and energy balance for a hardwood to ethanol plant.
Fieber, C.; Colcord, A.R.; Faass, S.; Muzzy, J.D.; Roberts, R.S.
Alkaline hydrolysis of N-benzoylanthranilic acid
1.The hydrolysis rate of N-benzoylanthranilic acid was studied at a KOH concentration of 2.2�44.6% and a temperature of 25�95�.2.The limiting step of the process is the reaction of the singly ionized form of the reagent with a molecule of water.3.The equilibrium constant for the addition of hydroxyl ion to the carbonyl atom of the amido group of N-benzoylanthranilic acid is
A. Teies Gonsalo; A. I. Donskikh; V. N. Kulagin; Yu. V. Moiseev; G. M. Tseitlin
Mercerization and acid hydrolysis of bacterial cellulose
Structural changes in never- dried, disintegrated bacteria l cellulose by treatment with aqueous NaOH were examined by electron microscopy, X-ray diffractometry and acid hydrolysis behaviour and compared with those of cotton cellulose. The microfibril kept its fibrillar morphology after treatment with NaOH solutions of less than 9% (w\\/w), but changed into irregular aggregates when treated with NaOH above 12% (w\\/w),
HIDEKI Shibazaki; SHIGENORI Kuga; TAKESHI Okano
?-cyclodextrin assistant flavonoid glycosides enzymatic hydrolysis
Background: The content of icaritin and genistein in herba is very low, preparation with relatively large quantities is an important issue for extensive pharmacological studies. Objective: This study focuses on preparing and enzymic hydrolysis of flavonoid glycosides /?-cyclodextrin inclusion complex to increase the hydrolysis rate. Materials and Methods: The physical property of newly prepared inclusion complex was tested by differential scanning calorimetry (DSC). The conditions of enzymatic hydrolysis were optimized for the bioconversion of flavonoid glycosides /?-cyclodextrin inclusion complex by mono-factor experimental design. The experiments are using the icariin and genistein as the model drugs. Results: The solubility of icariin and genistein were increased almost 17 times from 29.2 ?g/ml to 513.5 ?g/ml at 60�C and 28 times from 7.78 ?g/ml to 221.46 ?g/ml at 50�C, respectively, demonstrating that the inclusion complex could significantly increase the solubility of flavonoid glycosides. Under the optimal conditions, the reaction time of icariin and genistin decreased by 68% and 145%, when compared with that without ?-CD inclusion. By using this enzymatic condition, 473 mg icaritin (with the purity of 99.34%) and 567 mg genistein(with the purity of 99.46%), which was finally determined by melt point, ESI-MS, UV, IR, 1H NMR and 13C NMR, was obtained eventually by transforming the inclusion complex(contains 1.0 g substrates). Conclusion: This study can clearly indicate a new attempt to improve the speed of enzyme-hydrolysis of poorly water-soluble flavonoid glycosides and find a more superior condition which is used to prepare icaritin and genistein.
Jin, Xin; Zhang, Zhen-hai; Sun, E.; Jia, Xiao-Bin
21 CFR 172.372 - N-Acetyl-L-methionine.
Code of Federal Regulations, 2012 CFR
...conditions: (a) N- Acetyl-L-methionine (Chemical Abstracts Service Registry No. 65-82-7) is the derivative of the...3) Adequate directions for use to provide a finished food meeting the limitations prescribed by paragraph (c) of this...
21 CFR 172.372 - N-Acetyl-L-methionine.
Code of Federal Regulations, 2011 CFR
...conditions: (a) N- Acetyl-L-methionine (Chemical Abstracts Service Registry No. 65-82-7) is the derivative of the...3) Adequate directions for use to provide a finished food meeting the limitations prescribed by paragraph (c) of this...
Acetylation of banana fibre to improve oil absorbency.
Oil spill leaves detrimental effects on the environment, living organisms and economy. In the present work, an attempt is made to provide an efficient, easily deployable method of cleaning up oil spills and recovering of the oil. The work reports the use of banana fibres which were acetylated for oil spill recovery. The product so formed was characterized by FT-IR, TG, SEM and its degree of acetylation was also evaluated. The extent of acetylation was measured by weight percent gain. The oil sorption capacity of the acetylated fibre was higher than that of the commercial synthetic oil sorbents such as polypropylene fibres as well as un-modified fibre. Therefore, these oil sorption-active materials which are also biodegradable can be used to substitute non-biodegradable synthetic materials in oil spill cleanup. PMID:23218302
Teli, M D; Valia, Sanket P
NMR and rheological study of Aloe barbadensis partially acetylated glucomannan.
The structural and rheological properties of the Aloe extract (AE) and the polysaccharidic fraction (PF) obtained from the leaves pulp of Aloe barbadensis Miller were investigated. Structural analyses carried out by composition, methylation analysis and NMR spectroscopy showed that PF is mainly constituted by a partially acetylated 4-linked ?-d-glucomannan. The acetyl groups are located at C-2, C-2 and C-3, C-3 and/or C-6. The acetylation pattern of this type of polysaccharide was for the first time established using bidimensional NMR analyses. AE and PF aqueous solutions at 25�C showed a non-Newtonian behavior (with pseudoplastic characteristics), however PF showed higher apparent viscosity than AE. Dynamic oscillatory analyses showed that both samples, at the same concentration, behaved as a concentrated solution. PF presented higher values of G' compared with those of AE and this behavior could be consequence of its higher content in partially acetylated glucomannan. PMID:23544569
Campestrini, L H; Silveira, J L M; Duarte, M E R; Koop, H S; Noseda, M D
21 CFR 172.372 - N-Acetyl-L-methionine.
Code of Federal Regulations, 2013 CFR
...conditions: (a) N- Acetyl-L-methionine (Chemical Abstracts Service Registry No. 65-82-7) is the derivative of the...3) Adequate directions for use to provide a finished food meeting the limitations prescribed by paragraph (c) of this...
Towards a Functional Understanding of Protein N-Terminal Acetylation
Protein N-terminal acetylation is a major modification of eukaryotic proteins. Its functional implications include regulation of protein�protein interactions and targeting to membranes, as demonstrated by studies of a handful of proteins. Fifty years after its discovery, a potential general function of the N-terminal acetyl group carried by thousands of unique proteins remains enigmatic. However, recent functional data suggest roles for N-terminal acetylation as a degradation signal and as a determining factor for preventing protein targeting to the secretory pathway, thus highlighting N-terminal acetylation as a major determinant for the life and death of proteins. These contributions represent new and intriguing hypotheses that will guide the research in the years to come.
Arnesen, Thomas
Partially Acetylated Sugarcane Bagasse For Wicking Oil From Contaminated Wetlands
Sugarcane bagasse was partially acetylated to enhance its oil-wicking ability in saturated environments while holding moisture for hydrocarbon biodegradation. The water sorption capacity of raw bagasse was reduced fourfold after treatment, which indicated considerably increased ...
Lysine Acetylation Targets Protein Complexes and Co-Regulates Major Cellular Functions
Lysine acetylation is a reversible posttranslational modification of proteins and plays a key role in regulating gene expression. Technological limitations have so far prevented a global analysis of lysine acetylation's cellular roles. We used high-resolution mass spectrometry to identify 3600 lysine acetylation sites on 1750 proteins and quantified acetylation changes in response to the deacetylase inhibitors suberoylanilide hydroxamic acid and
Chunaram Choudhary; Chanchal Kumar; Florian Gnad; Michael L. Nielsen; Michael Rehman; Tobias C. Walther; Jesper V. Olsen; Matthias Mann
Thiogalactopyranosides are resistant to hydrolysis by ?-galactosidases.
Fluorescently tagged glycosides containing terminal ?(1?3) and ?(1?4)-linked thiogalactopyranosides have been prepared and tested for resistance to hydrolysis by ?-galactosidases. Eight fluorescent glycosides containing either galactose or 5-thiogalactose as the terminal sugar were enzymatically synthesized using galactosyltransferases, with lactosyl glycosides as acceptors and UDP-galactose or UDP-5'-thiogalactose, respectively, as donors. The glycosides were incubated with human ?-galactosidase A (CAZy family GH27, a retaining glycosidase), Bacteroides fragilis ?-1,3-galactosidase (GH110, an inverting glycosidase), or homogenates of MCF-7 human breast cancer cells or NG108-15 rat glioma cells. Substrate hydrolysis was monitored by capillary electrophoresis with fluorescence detection. All compounds containing terminal O-galactose were readily degraded. Their 5-thiogalactose counterparts were resistant to hydrolysis by human ?-galactosidase A and the enzymes present in the cell extracts. B. fragilis ?-1,3-galactosidase hydrolyzed both thio- and O-galactoside substrates; however, the thiogalactosides were hydrolyzed at only 1-3 % of the rate of O-galactosides. The hydrolytic resistance of 5-thiogalactose was also confirmed by an in vivo study using cells in culture. The results suggest that 5-thiogalactosides may be useful tools for the study of anabolic pathways in cell extracts or in single cells. PMID:22740420
Adlercreutz, Dietlind; Yoshimura, Yayoi; Mannerstedt, Karin; Wakarchuk, Warren W; Bennett, Eric P; Dovichi, Norman J; Hindsgaul, Ole; Palcic, Monica M
Palm date fibers: analysis and enzymatic hydrolysis.
Waste palm dates were subjected to analysis for composition and enzymatic hydrolysis of their flesh fibers. The fruit contained 32% glucose and 30% fructose, while the water-insoluble fibers of its flesh consisted of 49.9% lignin and 20.9% polysaccharides. Water-insoluble fibers were settled to 55% of its initial volume in 12 h. The presence of skin and flesh colloidal fibers results in high viscosity and clogging problems during industrial processes. The settling velocity of the fibers was improved by enzymatic hydrolysis. Hydrolysis resulted in 84.3% conversion of the cellulosic part of the fibers as well as reducing the settling time to 10 minutes and the final settled volume to 4% of the initial volume. It implies easier separation of the fibers and facilitates fermentation processes in the corresponding industries. Two kinds of high- and low-lignin fibers were identified from the water-insoluble fibers. The high-lignin fibers (75% lignin) settled easily, while the low-lignin fibers (41.4% lignin) formed a slurry suspension which settled very slowly. The hydrophilicity of these low-lignin fibers is the major challenge of the industrial processes. PMID:21151438
Shafiei, Marzieh; Karimi, Keikhosro; Taherzadeh, Mohammad J
Structural basis unifying diverse GTP hydrolysis mechanisms.
Central to biological processes is the regulation rendered by GTPases. Until recently, the GTP hydrolysis mechanism, exemplified by Ras-family (and G-?) GTPases, was thought to be universal. This mechanism utilizes a conserved catalytic Gln supplied "in cis" from the GTPase and an arginine finger "in trans" from a GAP (GTPase activating protein) to stabilize the transition state. However, intriguingly different mechanisms are operative in structurally similar GTPases. MnmE and dynamin like cation-dependent GTPases lack the catalytic Gln and instead employ a Glu/Asp/Ser situated elsewhere and in place of the arginine finger use a K(+) or Na(+) ion. In contrast, Rab33 possesses the Gln but does not utilize it for catalysis; instead, the GAP supplies both a catalytic Gln and an arginine finger in trans. Deciphering the underlying principles that unify seemingly unrelated mechanisms is central to understanding how diverse mechanisms evolve. Here, we recognize that steric hindrance between active site residues is a criterion governing the mechanism employed by a given GTPase. The Arf-ArfGAP structure is testimony to this concept of spatial (in)compatibility of active site residues. This understanding allows us to predict an as yet unreported hydrolysis mechanism and clarifies unexplained observations about catalysis by Rab11 and the need for HAS-GTPases to employ a different mechanism. This understanding would be valuable for experiments in which abolishing GTP hydrolysis or generating constitutively active forms of a GTPase is important. PMID:23293872
Anand, Baskaran; Majumdar, Soneya; Prakash, Balaji
Sugarcane bagasse hydrolysis using yeast cellulolytic enzymes.
Ethanol fuel production from lignocellulosic biomass is emerging as one of the most important technologies for sustainable development. To use this biomass, it is necessary to circumvent the physical and chemical barriers presented by the cohesive combination of the main biomass components, which hinders the hydrolysis of cellulose and hemicellulose into fermentable sugars. This study evaluated the hydrolytic capacity of enzymes produced by yeasts, isolated from the soils of the Brazilian Cerrado biome (savannah) and the Amazon region, on sugarcane bagasse pre-treated with H2SO4. Among the 103 and 214 yeast isolates from the Minas Gerais Cerrado and the Amazon regions, 18 (17.47%) and 11 (5.14%) isolates, respectively, were cellulase-producing. Cryptococcus laurentii was prevalent and produced significant ?- glucosidase levels, which were higher than the endo- and exoglucanase activities. In natura sugarcane bagasse was pre-treated with 2% H2SO4 for 30 min at 150oC. Subsequently, the obtained fibrous residue was subjected to hydrolysis using the Cryptococcus laurentii yeast enzyme extract for 72 h. This enzyme extract promoted the conversion of approximately 32% of the cellulose, of which 2.4% was glucose, after the enzymatic hydrolysis reaction, suggesting that C. laurentii is a good ?-glucosidase producer. The results presented in this study highlight the importance of isolating microbial strains that produce enzymes of biotechnological interest, given their extensive application in biofuel production. PMID:23851270
Souza, Angelica Cristina de; Carvalho, Fernanda Paula; Silva e Batista, Cristina Ferreira; Schwan, Rosane Freitas; Dias, Disney Ribeiro
Palm Date Fibers: Analysis and Enzymatic Hydrolysis
Waste palm dates were subjected to analysis for composition and enzymatic hydrolysis of their flesh fibers. The fruit contained 32% glucose and 30% fructose, while the water-insoluble fibers of its flesh consisted of 49.9% lignin and 20.9% polysaccharides. Water-insoluble fibers were settled to 55% of its initial volume in 12 h. The presence of skin and flesh colloidal fibers results in high viscosity and clogging problems during industrial processes. The settling velocity of the fibers was improved by enzymatic hydrolysis. Hydrolysis resulted in 84.3% conversion of the cellulosic part of the fibers as well as reducing the settling time to 10 minutes and the final settled volume to 4% of the initial volume. It implies easier separation of the fibers and facilitates fermentation processes in the corresponding industries. Two kinds of high- and low-lignin fibers were identified from the water-insoluble fibers. The high-lignin fibers (75% lignin) settled easily, while the low-lignin fibers (41.4% lignin) formed a slurry suspension which settled very slowly. The hydrophilicity of these low-lignin fibers is the major challenge of the industrial processes.
Shafiei, Marzieh; Karimi, Keikhosro; Taherzadeh, Mohammad J.
Fungal secretomes enhance sugar beet pulp hydrolysis.
The recalcitrance of lignocellulose makes enzymatic hydrolysis of plant biomass for the production of second generation biofuels a major challenge. This work investigates an efficient and economic approach for the enzymatic hydrolysis of sugar beet pulp (SBP), which is a difficult to degrade, hemicellulose-rich by-product of the table sugar industry. Three fungal strains were grown on different substrates and the production of various extracellular hydrolytic and oxidative enzymes involved in pectin, hemicellulose, and cellulose breakdown were monitored. In a second step, the ability of the culture supernatants to hydrolyze thermally pretreated SBP was tested in batch experiments. The supernatant of Sclerotium rolfsii, a soil-borne facultative plant pathogen, was found to have the highest hydrolytic activity on SBP and was selected for further hydrolyzation experiments. A low enzyme load of 0.2 mg g(-1) protein from the culture supernatant was sufficient to hydrolyze a large fraction of the pectin and hemicelluloses present in SBP. The addition of Trichoderma reesei cellulase (1-17.5 mg g(-1) SBP) resulted in almost complete hydrolyzation of cellulose. It was found that the combination of pectinolytic, hemicellulolytic, and cellulolytic activities works synergistically on the complex SBP composite, and a combination of these hydrolytic enzymes is required to achieve a high degree of enzymatic SBP hydrolysis with a low enzyme load. PMID:24677771
Kracher, Daniel; Oros, Damir; Yao, Wanying; Preims, Marita; Rezic, Iva; Haltrich, Dietmar; Rezic, Tonci; Ludwig, Roland
Acetylation Polymorphism and Prevalence of Colorectal Adenomas1
Polymorphic W-acetyltransferase (NAT2), an enzyme present in the colon, may affect incidence of colon cancer. Individuals with NAT2 fast acetylator genotypes may have higher colon cancer risks due to faster conversion of certain carcinogens to mutagens. We determined NAT2 genotypes in 447 subjects with distal colon adenomas and in 487 controls. No significant increase in adenoma prevalence among fast acetylators
Nicole M. Probst-Hensch; Robert W. Haile; Sue Ann Ingles; Matthew P. Longnecker; Chun-Ya Han; Bruce K. Lin; David B. Lee; Gordon T. Sakamoto; Harold D. Frank; Eric R. Lee; Henry J. Lin
Nitric oxide and histone acetylation-shaping craniofacial development.
Previous work linked nitric oxide (NO) signaling to histone deacetelyases (HDACs) in the control of tissue homeostasis and suggested that deregulation of this signaling contributes to human diseases. In the previous issue of Chemistry & Biology, Kong and colleagues showed that coordinated NO signaling and histone acetylation are required for proper cranial neural crest development and craniofacial morphogenesis and suggested that alterations of NO/acetylation network can contribute to the pathogenesis of craniofacial malformations. PMID:24856136
Berghella, Libera; Puri, Pier Lorenzo
Acetylation of rhamnogalacturonan I and homogalacturonan: Theoretical calculations
The possible confirmation of experimental data through theoretical calculations on the exact position of acetyl groups at the galacturonic acid residues in pectin backbones was investigated. With MM3(92) it was calculated that acetyl groups at both O2 and O3 of galacturonic acid in the backbone of rhamnogalacturonan I (RG-I) and homogalacturonan are energetically favourable, where the most important contribution comes
Milou Kouwijzer; Henk Schols; Serge P�rez
Ultra high pressure (UHP)-assisted acetylation of corn starch
Potential roles of ultra high pressure (UHP) in starch granule reactivity and properties of acetylated starch were investigated. Corn starch was substituted with acetic anhydride at pressure range of 0.1�400MPa for 15min; also, conventional reaction (30�C, 60min) was conducted as reaction control. Native and acetylated corn starches were assessed with respect to degree of substitution (DS), X-ray diffraction pattern\\/relative crystallinity,
Hyun-Shik Choi; Hyun-Seok Kim; Cheon-Seok Park; Byung-Yong Kim; Moo-Yeol Baik
Predicting N-terminal acetylation based on feature selection method
Methionine aminopeptidase and N-terminal acetyltransferase are two enzymes that contribute most to the N-terminal acetylation, which has long been recognized as a frequent and important kind of co-translational modifications [R.A. Bradshaw, W.W. Brickey, K.W. Walker, N-terminal processing: the methionine aminopeptidase and N alpha-acetyl transferase families, Trends Biochem. Sci. 23 (1998) 263�267]. The combined action of these two enzymes leads to
Yu-Dong Cai; Lin Lu
Investigation of the Acetylation Mechanism by GCN5 Histone Acetyltransferase
The histone acetylation of post-translational modification can be highly dynamic and play a crucial role in regulating cellular proliferation, survival, differentiation and motility. Of the enzymes that mediate post-translation modifications, the GCN5 of the histone acetyltransferase (HAT) proteins family that add acetyl groups to target lysine residues within histones, has been most extensively studied. According to the mechanism studies of
Junfeng Jiang; Junyan Lu; Dan Lu; Zhongjie Liang; Lianchun Li; Sisheng Ouyang; Xiangqian Kong; Hualiang Jiang; Bairong Shen; Cheng Luo
Reduced microtubule acetylation in cystic fibrosis epithelial cells.
Dysfunctional cystic fibrosis transmembrane conductance regulator (CFTR) leads to many cellular consequences, including perinuclear accumulation of free cholesterol due to impaired endosomal transport. The hypothesis being tested is that CF-related perinuclear cholesterol accumulation due to disrupted endocytic trafficking occurs as a result of reduced microtubule (MT) acetylation. Here, it is identified that acetylated-?-tubulin (Ac-tub) content is reduced by ?40% compared with respective wild-type controls in both cultured CF cell models (IB3) and primary Cftr-/- mouse nasal epithelial tissue. Histone deacetylase 6 (HDAC6) has been shown to regulate MT acetylation, which provides reasonable grounds to test its impact on reduced Ac-tub content on CF cellular phenotypes. Inhibition of HDAC6, either through tubastatin treatment or HDAC6 knockdown in CF cells, increases Ac-tub content and results in redistributed free cholesterol and reduced stimulation of NF-?B activity. Mechanistically, endoplasmic reticulum stress, which is widely reported in CF and leads to aggresome formation, is identified as a regulator of MT acetylation. F508del CFTR correction with C18 in primary airway epithelial cells restores MT acetylation and cholesterol transport. A significant role for phosphatidyl inositol-3 kinase p110? is also identified as a regulator of MT acetylation. PMID:23873844
Rymut, Sharon M; Harker, Alyssa; Corey, Deborah A; Burgess, James D; Sun, Hongtao; Clancy, John P; Kelley, Thomas J
We review recent evidence that acetylation and deacetylation of cellular proteins, including transcription factors and nuclear cofactors, may be involved in the regulation of muscle mass. The level of protein acetylation is balanced by histone acetyltransferases (HATs) and histone deacetylases (HDACs) and studies suggest that this balance is perturbed in muscle wasting. Hyperacetylation of transcription factors and nuclear cofactors regulating gene transcription in muscle wasting may influence muscle mass. In addition, hyperacetylation may render proteins susceptible to degradation by different mechanisms, including intrinsic ubiquitin ligase activity exerted by HATs and by dissociation of proteins from cellular chaperones. In recent studies, inhibition of p300/HAT expression and activity and stimulation of SIRT1-dependent HDAC activity reduced glucocorticoid-induced catabolic response in skeletal muscle, providing further evidence that hyperacetylation plays a role in muscle wasting. It should be noted, however, that although several studies advocate a role of hyperacetylation in muscle wasting, apparently contradictory results have also been reported. For example, muscle atrophy caused by denervation or immobilization may be associated with reduced, rather than increased, protein acetylation. In addition, whereas hyperacetylation results in increased degradation of certain proteins, other proteins may be stabilized by increased acetylation. Thus, the role of acetylation and deacetylation in the regulation of muscle mass may be both condition- and protein-specific. The influence of HATs and HDACs on the regulation of muscle mass as well as methods to modulate protein acetylation are important areas for continued research aimed at preventing and treating muscle wasting.
Alamdari, Nima; Aversa, Zaira; Castillero, Estibaliz; Hasselgren, Per-Olof
Acetylation of p53 activates transcription through recruitment of coactivators/histone acetyltransferases.
Cellular DNA damage causes stabilization and activation of the tumor suppressor and transcription factor p53, in part by promoting multiple covalent modifications of the p53 protein, including acetylation. We investigated the importance of acetylation in p53 function and the mechanism by which acetylation influences p53 activity. Acetylation site substitutions reduced p53-dependent transcriptional induction and G1 cell cycle arrest. Chromatin immunoprecipitation analysis of the endogenous p21 promoter showed increased association of p53, coactivators (CBP and TRRAP), and acetylated histones following cell irradiation. Results with acetylation-defective p53 demonstrate that the critical function of acetylation is not to increase the DNA binding affinity of p53 but rather to promote coactivator recruitment and histone acetylation. Therefore, we propose that an acetylation cascade consisting of p53 acetylation-dependent recruitment of coactivators/HATs is crucial for p53 function. PMID:11779500
Barlev, N A; Liu, L; Chehab, N H; Mansfield, K; Harris, K G; Halazonetis, T D; Berger, S L
EPR of Gamma irradiated N?-acetyl L-glutamic acid and N?-acetyl L-glutamine
NASA Astrophysics Data System (ADS)
Single crystals of N?-acetyl L-glutamic acid and N?-acetyl L-glutamine were ?-irradiated and the paramagnetic species formed were investigated at room temperature by electron spin resonance technique. The observed species in N?-acetyl L-glutamic acid single crystal were attributed to the, HOOCCH 2CH 2C(NHCOCH 3)COOH radical; and those in N?-acetyl L-glutamine single crystal to the NH2COCH2CH2 C( NHCOCH3) COOH and NH2COCH2CH2CH( NHCOCH3) C overlineOOH radicals. The g values and the hyperfine coupling constants of the unpaired electron with the environmental methylene, methine protons and 14N nucleus were determined.
K�ksal, F.; Osmano?lu, ?.; Kartal, �.; Ucun, F.
Characteristics of acetylated starches prepared using starches separated from different rice cultivars
The physico-chemical properties of acetylated starches from different rice cultivars were evaluated. The acetylation of starches from different cultivars showed different acetyl (%) and degrees of substitution (DS). The FT-IR spectral analysis confirmed the introduction of acetyl moiety in the acetylated starches through a band at 1730.8cm?1. The scanning electron microscopy did not show any significant difference between the external
Navdeep Singh Sodhi; Narpinder Singh
Acetylation of acetyl-CoA synthetase from Mycobacterium tuberculosis leads to specific inactivation of the adenylation reaction.
Acetyl-CoA synthetase (ACS) catalyzes the formation of AcCoA from acetate, ATP and Coenzyme A, allowing the organism to grow on acetate as the sole carbon source. ACS was the first enzyme in Mycobacterium tuberculosis shown to be regulated by posttranslational acetylation by the cAMP-dependent protein acetyltransferase. This modification results in the inactivation of the enzyme and can be reversed in the presence of NAD(+) and a mycobacterial sirtuin-like deacetylase. In this study we characterize the kinetic mechanism of MtACS, where the overall reaction can be divided into two half-reactions: the acetyl-adenylate forming reaction and the thiol-ligation reaction. We also provide evidence for the existence of the acetyl-adenylate intermediate via (31)P NMR spectroscopy. Furthermore, we dissect the regulatory role of K617 acetylation and show that acetylation inhibits only the first, adenylation half-reaction while leaving the second half reaction unchanged. Finally, we demonstrate that the chemical mechanism of the enzyme relies on a conformational change which is controlled by the protonation state of aspartate 525. Together with our earlier results, this suggests a degree of regulation of enzyme activity that is appropriate for the role of the enzyme in central carbon metabolism. PMID:24751484
Noy, Tahel; Xu, Hua; Blanchard, John S
Organosolv pretreatment for enzymatic hydrolysis of poplars: I. enzyme hydrolysis of cellulosic residues
Aspen (Populus tremuloides) and black cottonwood (Populus trichocarpa) organosolv pulps produced in a wide range of solvent composition (between 30 and 70% by volume of methanol) and catalysts (H/sub 2/SO/sub 4/ and H/sub 3/PO/sub 4/) such that the cooking liquor pH less than or equal to 3 are easily digested by enzymes. The total yields of hydrolysis residues (pulps) are in the 40-60% range; the acid-catalyzed delignification followed by enzyme hydrolysis can generate 70-88% of the original six-carbon sugars contained in the wood. Glucomannan and arabinogalactan are dissolved in to the pulping liquor in the pH range of 2-4.5. Lower pH (less than or equal to 3) leads to additional solubilization of six-carbon sugars. These sugars may be fermented directly. From the insoluble hydrolysis residues, 36-41% conversions of wood into fermentable sugars were obtained after enzyme hydrolysis; the starting feedstocks contain 50.8 and 46.6% hexosans, respectively, for aspen and black cottonwood. The kinetics of enzymatic hydrolysis of cellulose can be formally treated as two simultaneous pseudo-first-order reactions in which fast and slow hydrolysis of cellulose occur. Correlations between the glucan digestibility and the effect of the pretreatment have been made. The higher residual xylan content reduces the amount of the rapidly hydrolyzable glucan fraction and lowers the glucan digestibility. The proposed simple kinetic treatment is very helpful in assessing the effect of the pretreatment on pulp enzyme hydrolyzability.
Chum, H.L.; Johnson, D.K.; Black, S.; Baker, J.; Grohmann, K.; Sarkanen, K.V.; Wallace, K.; Schroeder, H.A.
Determination of the individual rate and association constants of the hydrolysis catalysed by serine proteinases by means of added nucleophiles in Eisenthal and Cornish-Bowden coordinates.
It is shown that in the three-step enzyme-catalysed hydrolysis the addition of nucleophile shifts the common intersection point in Eisenthal and Cornish-Bowden coordinates when registering the second product P2. The different points obtained at different nucleophile concentrations are situated on a straight line with intercepts Ks on the Km axis and k3 [E]o on the V axis. Since the Eisenthal and Cornish-Bowden method is considered as the best graphic method for determination of the kinetic parameters Km and V of enzyme reactions, the graphic procedure proposed here for determination of k2, k3, k4 and Ks by the method of added nucleophile to be preferred. This procedure was applied for determination of individual constants on the hydrolysis of N-acetylated L-amino acid methyl esters catalysed by alkaline mesentericopeptidase. PMID:6998498
Dorovska-Taran, V; Raykova, D
Purification and characterization of an extracellular muramidase of Clostridium acetobutylicum ATCC 824 that acts on non-N-acetylated peptidoglycan.
An extracellular enzyme showing lytic activity on non-N-acetylated peptidoglycan has been isolated from Clostridium acetobutylicum ATCC 824. The lytic enzyme was purified to homogeneity by anion-exchange chromatography and gel filtration, with a recovery of 24%. The enzyme was monomeric and had an estimated molecular weight of 41,000 and an isoelectric point of 3.8. It has been characterized as a muramidase whose 23-amino-acid N terminus displayed 39% homology with the N,O-diacetyl muramidase of the fungus Chalaropsis sp. The muramidase hydrolyzed purified cell walls at an optimum pH of 3, with a maximum velocity of 9.1 mumol of reducing sugars released min-1 mg of muramidase-1 and a concentration of cell walls giving a half-maximum rate of 0.01 mg ml-1. Its activity was inhibited by glucosamine, N-acetylglucosamine, Hg2+, Fe3+, and Ag+ but not by choline. The muramidase-peptidoglycan complex rapidly dissociated before total hydrolysis of the chain and randomly reassociated on another peptidoglycan chain. The affinity of the muramidase was affected by the protein content and the acetylation of the cell wall. Images
Croux, C; Canard, B; Goma, G; Soucaille, P
Synergy between cellulases and pectinases in the hydrolysis of hemp.
The impact of pectinases in the hydrolysis of fresh, steam-exploded and ensiled hemp was investigated and the synergy between cellulases, pectinases and xylanase in the hydrolysis was evaluated. About half; 59.3% and 46.1% of pectin in the steam-exploded and ensiled hemp, respectively, could be removed by a low dosage of pectinases used. Pectinases were more efficient than xylanase in the hydrolysis of fresh and ensiled hemp whereas xylanase showed higher hydrolytic efficiency than the pectinase preparation used in the hydrolysis of steam-exploded hemp. Clear synergistic action between cellulases and xylanase could be observed in the hydrolysis of steam-exploded hemp. Supplementation of pectinase resulted in clear synergism with cellulases in the hydrolysis of all hemp substrates. Highest hydrolysis yield of steam-exploded hemp was obtained in the hydrolysis with cellulases and xylanase. In the hydrolysis of ensiled hemp, the synergistic action between cellulases and pectinases was more obvious for efficient hydrolysis. PMID:23262004
Zhang, Junhua; Pakarinen, Annukka; Viikari, Liisa
Expression of Brassica juncea 3-hydroxy-3-methylglutaryl CoA synthase is developmentally regulated and stress-responsive.
3-Hydroxy-3-methylglutaryl-coenzyme A synthase (HMGS) is an enzyme in mevalonate biosynthesis. In plants, investigations have focused on HMG CoA reductase (HMGR) and less is known of the preceding enzyme, HMGS. To understand the regulation of HMGS, we have isolated a Brassica juncea cDNA encoding HMGS, BjHMGS1, for use as a hybridization probe in Northern blot analyses. BjHMGS is expressed in all plant organs and shows developmental regulation in flower, seed and seedling, with highest expression in early development. In seedlings, expression is highest in young hypocotyls and is induced during the greening of etiolated cotyledons. BjHMGS is down-regulated by abscisic acid, osmotic stress and dehydration, the effects of which arrested seedling growth. Thus BjHMGS expression shows correlation with rapid cell division and growth, like HMGR. This is not unexpected, as mevalonate is the precursor to many essential isoprenoid compounds, including sterols for membrane biogenesis. Wounding, methyl jasmonate or salicylic acid induce BjHMGS expression, suggesting that, like HMGR, HMGS is involved in defence. As in animals, coordinated regulation of HMGS with HMGR occurred in B. juncea upon germination and in response to salicylic acid. HMGS assays confirmed that Escherichia coli-expressed recombinant BjHMGS1 shows HMGS activity that is inhibited by F244, a specific inhibitor of HMGS. Southern blot analysis revealed gene families encoding HMGS in Brassica species and a summation of homologous genes in the fusion amphidiploid genome of B. juncea, a bi-parental species derived from diploids B. nigra and B. campestris. PMID:10849357
Alex, D; Bach, T J; Chye, M L
Hydrolysis of ferric chloride in solution
The Detox{trademark} process uses concentrated ferric chloride and small amounts of catalysts to oxidize organic compounds. It is under consideration for oxidizing transuranic organic wastes. Although the solution is reused extensively, at some point it will reach the acceptable limit of radioactivity or maximum solubility of the radioisotopes. This solution could be cemented, but the volume would be increased substantially because of the poor compatibility of chlorides and cement. A process has been developed that recovers the chloride ions as HCl and either minimizes the volume of radioactive waste or permits recycling of the radioactive chlorides. The process involves a two-step hydrolysis at atmospheric pressure, or preferably under a slight vacuum, and relatively low temperature, about 200{degrees}C. During the first step of the process, hydrolysis occurs according to the reaction below: FeCl{sub 3 liquid} + H{sub 2}O {r_arrow} FeOCl{sub solid} + 2 HCl{sub gas} During the second step, the hot, solid, iron oxychloride is sprayed with water or placed in contact with steam, and hydrolysis proceeds to the iron oxide according to the following reaction: 2 FeOCl{sub solid} + H{sub 2}O {r_arrow} Fe{sub 2}O{sub 3 solid} + 2 HCl{sub gas}. The iron oxide, which contains radioisotopes, can then be disposed of by cementation or encapsulation. Alternately, these chlorides can be washed off of the solids and can then either be recycled or disposed of in some other way.
Lussiez, G.; Beckstead, L.
[Immobilized glucoamylase: A biocatalyst of dextrin hydrolysis].
Heterogeneous biocatalysts of starch conversion based on glucoamylase and carbon-containing carriers were obtained, and their biocatalytic properties in enzymatic hydrolysis of corn dextrins were studied. It was shown that the morphology of the surface carbon layer of carriers markedly affected the properties of biocatalysts. Glucoamylase that was immobilized by adsorption on the surface of carriers covered with a layer of catalytic fibrous or pyrolytic carbon had the maximum enzymatic activity and stability, whereas the biocatalysts prepared on the basis of carriers that had no carbon layer or were covered with graphite-like surface carbon had a low activity and stability. PMID:16761568
Kovalenko, G A; Perminova, L V; Plaksin, G V; Chuenko, T V; Komova, O V; Rudina, N A
Improved method for detection of starch hydrolysis
A new starch hydrolysis detection method which does not rely on iodine staining or the use of color-complexed starch is described. A linear relationship was obtained with agar-starch plates when net clearing zones around colonies of yeasts were plotted against enzyme levels (semilogarithm scale) produced by the same yeast strains in liquid medium. A similar relationship between starch clearing zones and alpha-amylase levels from three different sources was observed. These observations suggest that the method is useful in mutant isolations, strain improvement programs, and the prediction of alpha-amylase activities in culture filtrates or column effluents. (Refs. 18).
Ohawale, M.R.; Wilson, J.J.; Khachatourians, G.G.; Ingledew, W.M.
Hydrolysis of olefin oxides to glycols
A process is described for making 1,2-diols by the catalytic hydrolysis of an olefin oxide or a cyclo-olefin oxide, in the presence of water. The process comprises contacting the olefin oxide or a cyclo-olefin oxide of 2 to 20 carbon atoms with a catalyst comprising a silicate or zeolite. The silicate or zeolite, has a contraint index of about 1 to about 12, wherein the silicate or zeolite has an alpha value ranging from 1 to 500; and wherein the silicate or zeolite contains acidic hydrogen atoms.
Chang, C.D.; Hellring, S.D.
Urea hydrolysis and calcium carbonate reaction fronts
NASA Astrophysics Data System (ADS)
The mobility of toxic or radioactive metal contaminants in subsurface environments can be reduced by the formation of mineral precipitates that form co-precipitates with the contaminants or that isolate them from the mobile fluid phase. An engineering challenge is to control the spatial distribution of precipitation reactions with respect to: 1) the location of a contaminant, and 2) where reactants are introduced into the subsurface. One strategy being explored for immobilizing contaminants, such as Sr-90, involves stimulating mineral precipitation by forming carbonate ions and hydroxide via the in situ, microbially mediated hydrolysis of urea. A series of column experiments have been conducted to explore how the construction or design of such an in situ reactant production strategy can affect the temporal and spatial distribution of calcium carbonate precipitation, and how the distribution is coupled to changes in permeability. The columns were constructed with silica gel as the porous media. An interval midway through the column contained an adsorbed urease enzyme in order to simulate a biologically active zone. A series of influent solutions were injected to characterize hydraulic properties of the column (e.g., bromide tracer), profiles of chemical conditions and reaction products as the enzyme catalyzes urea hydrolysis (e.g., pH, ammonia, urea), and changes that occur due to CaCO3 precipitation with the introduction of a calcium+urea solutions. In one experiment, hydraulic conductivity was reduced as precipitate accumulated in a layer within the column that had a higher fraction of fine grained silica gel. Subsequent reduction of permeability and flow (for a constant head condition) resulted in displacement of the hydrolysis and precipitation reaction profiles upstream. In another experiment, which lacked the physical heterogeneity (fine grained layer), the precipitation reaction did not result in loss of permeability or flow velocity and the reaction profile, characterized by the pH profile and hydrolysis reaction species, was extended downstream of the enzyme zone. Downstream extension of the reaction profile was due partially to the partial mobility of the enzyme in the column. The experiments are helping to illustrate the complexity of transient reaction fronts as well as the needs and challenges for advanced modeling approaches. A modeling platform developed at the Idaho National Laboratory, which is capable of simulating tightly coupled physical-chemical processes (the Reactive Transport simulator), is being applied to pre-experimental simulations and post-experimental interpretation of results.
Fox, D. T.; Redden, G. D.; Henriksen, J.; Fujita, Y.; Guo, L.; Huang, H.
Metal ion-carbonyl oxygen recognition in complexes of acetyl phosphate.
Studies on acetyl phosphate (AcP2-), one of the so-called 'energy-rich' mixed-acid anhydrides, are summarized. Based on stability constants determined by potentiometric pH titrations in aqueous solution, it is shown that the M(AcP) complexes of Ca2+, Mg2+, Mn2+, Cu2+, and Zn2+ are more stable than is expected from the basicity of the phosphate group of AcP2-. This observed stability increase is attributed to an additional interaction of the already phosphate-coordinated metal ion (M2+) with the carbonyl oxygen of the anhydride unit. These conclusions are corroborated by the properties of the complexes of the hydrolysis-stable acetonylphosphonate (AnP2-). The formation degrees of the various six-membered chelates occurring in the M(AcP) and M(AnP) systems are presented and evidence is given that these chelates persist in mixed ligand complexes and that their formation degree is promoted by a low solvent polarity. The biological relevance of these results regarding carbonyl oxygen-metal ion interactions is briefly indicated. PMID:10830874
Sigel, H; Da Costa, C P
Xylan as limiting factor in enzymatic hydrolysis of nanocellulose.
The role of xylan as a limiting factor in the enzymatic hydrolysis of cellulose was studied by hydrolysing nanocellulose samples prepared by mechanical fibrillation of birch pulp with varying xylan content. Analyzing the nanocelluloses and their hydrolysis residues with dynamic FT-IR spectroscopy revealed that a certain fraction of xylan remained tightly attached to cellulose fibrils despite partial hydrolysis of xylan with xylanase prior to pulp fibrillation and that this fraction remained in the structure during the hydrolysis of nanocellulose with cellulase mixture as well. Thus, a loosely bound fraction of xylan was predicted to have been more likely removed by purified xylanase. The presence of loosely bound xylan seemed to limit the hydrolysis of crystalline cellulose, indicated by an increase in cellulose crystallinity and by preserved crystal width measured with wide-angle X-ray scattering. Removing loosely bound xylan led to a proportional hydrolysis of xylan and cellulose with the cellulase mixture. PMID:23238342
Penttil�, Paavo A; V�rnai, Anik�; Pere, Jaakko; Tammelin, Tekla; Salm�n, Lennart; Siika-aho, Matti; Viikari, Liisa; Serimaa, Ritva
The Hydrolysis of Carbonyl Sulfide at Low Temperature: A Review
Catalytic hydrolysis technology of carbonyl sulfide (COS) at low temperature was reviewed, including the development of catalysts, reaction kinetics, and reaction mechanism of COS hydrolysis. It was indicated that the catalysts are mainly involved metal oxide and activated carbon. The active ingredients which can load on COS hydrolysis catalyst include alkali metal, alkaline earth metal, transition metal oxides, rare earth metal oxides, mixed metal oxides, and nanometal oxides. The catalytic hydrolysis of COS is a first-order reaction with respect to carbonyl sulfide, while the reaction order of water changes as the reaction conditions change. The controlling steps are also different because the reaction conditions such as concentration of carbonyl sulfide, reaction temperature, water-air ratio, and reaction atmosphere are different. The hydrolysis of carbonyl sulfide is base-catalyzed reaction, and the force of the base site has an important effect on the hydrolysis of carbonyl sulfide.
Zhao, Shunzheng; Yi, Honghong; Tang, Xiaolong; Jiang, Shanxue; Gao, Fengyu; Zhang, Bowen; Zuo, Yanran; Wang, Zhixiang
Coordinate Regulation of the Nuclear and Plastidic Genes Coding for the Subunits of the Heteromeric Acetyl-Coenzyme A Carboxylase1
Plastidic acetyl-coenzyme A (CoA) carboxylase (ACCase) catalyzes the first committed reaction of de novo fatty acid biosynthesis. This heteromeric enzyme is composed of one plastid-coded subunit (?-carboxyltransferase) and three nuclear-coded subunits (biotin carboxy-carrier, biotin carboxylase, and ?-carboxyltransferase). We report the primary structure of the Arabidopsis ?-carboxyltransferase and ?-carboxyltransferase subunits deduced from nucleotide sequences of the respective genes and/or cDNA. Co-immunoprecipitation experiments confirm that the ?-carboxyltransferase and ?-carboxyltransferase subunits are physically associated. The plant ?-carboxyltransferases have gained a C-terminal domain relative to eubacteria, possibly via the evolutionary acquisition of a single exon. This C-terminal domain is divergent among plants and may have a structural function rather than being essential for catalysis. The four ACCase subunit mRNAs accumulate to the highest levels in tissues and cells that are actively synthesizing fatty acids, which are used either for membrane biogenesis in rapidly growing tissues or for oil accumulation in developing embryos. Development coordinately affects changes in the accumulation of the ACCase subunit mRNAs so that these four mRNAs maintain a constant molar stoichiometric ratio. These data indicate that the long-term, developmentally regulated expression of the heteromeric ACCase is in part controlled by a mechanism(s) that coordinately affects the steady-state concentrations of each subunit mRNA.
Ke, Jinshan; Wen, Tuan-Nan; Nikolau, Basil J.; Wurtele, Eve Syrkin
Lysine acetylation: enzymes, bromodomains and links to different diseases.
Lysine acetylation refers to transfer of the acetyl moiety from acetyl-CoA to the ?-amino group of a lysine residue on a protein. This has recently emerged as a major covalent modification and interplays with other modifications, such as phosphorylation, methylation, ubiquitination (addition of a small protein called ubiquitin) and SUMOylation [addition of a ubiquitin-like protein known as SUMO (small ubiquitin-related modifier)], to form multisite modification programmes for cellular regulation in diverse organisms. This modification is post-translational (i.e. after synthesis of a protein) and reversible, with its level being dynamically balanced by two groups of enzymes known as lysine acetyltransferases and deacetylases. The acetyltransferases belong to three major families, whereas deacetylases have been divided into the classical and sirtuin [Sir-tu-in, for Sir2 (silent information regulator 2)-like protein; named after the yeast protein Sir2] families. In addition to these enzymes, proteins containing the bromodomain, a protein module named after the fly protein Brahma (God of creation in Hindu), are relevant to lysine acetylation biology due to their ability to recognize acetyl-lysine-containing peptides. Importantly, recent studies have made intimate links between these three different groups of proteins to different pathological conditions. In this chapter, we provide a brief overview of these proteins and emphasize their direct links to related human diseases. PMID:22708559
You, Linya; Nie, Jianyun; Sun, Wei-Jian; Zheng, Zhi-Qiang; Yang, Xiang-Jiao
Counteraction on experimentally induced diabetic neuropathy by levocarnitine acetyl.
The effect of levocarnitine acetyl on diabetic peripheral neuropathy induced by a single injection of streptozotocin or alloxan was studied. Levocarnitine acetyl was administered intraperitoneally one week after induction of diabetes at the dose of 50 mg/kg/day for five and ten weeks. At the end of treatment, neuromuscular conduction velocity (m/sec) was evaluated by stimulating the sciatic nerve and recording the soleus muscle potentials evoked, and the muscle contraction force (mm) by measuring the isometric muscular tension. Motor coordination was evaluated on the Rota-rod apparatus. Treatment with levocarnitine acetyl fully prevented the reduction (20%) in the neuromuscular conduction velocity observed in both experimental models of diabetes. The decrease (30-33%) in muscle contraction force was prevented partially in streptozotocin-induced diabetes and fully in alloxan-induced diabetes. Levocarnitine acetyl also improved the concomitantly reduced motor performance. The results of the present study suggest a beneficial effect of levocarnitine acetyl on peripheral neuropathy and muscle performance. PMID:1301403
Pacifici, L; Bellucci, A; Piovesan, P; Maccari, F; Gorio, A; Ramacci, M T
Reversible acetylation regulates vascular endothelial growth factor receptor-2 activity.
The tyrosine kinase receptor vascular endothelial growth factor receptor 2 (VEGFR2) is a key regulator of angiogenesis. Here we show that VEGFR2 is acetylated in endothelial cells both at four lysine residues forming a dense cluster in the kinase insert domain and at a single lysine located in the receptor activation loop. These modifications are under dynamic control of the acetyltransferase p300 and two deacetylases HDAC5 and HDAC6. We demonstrate that VEGFR2 acetylation essentially regulates receptor phosphorylation. In particular, VEGFR2 acetylation significantly alters the kinetics of receptor phosphorylation after ligand binding, allowing receptor phosphorylation and intracellular signaling upon prolonged stimulation with VEGF. Molecular dynamics simulations indicate that acetylation of the lysine in the activation loop contributes to the transition to an open active state, in which tyrosine phosphorylation is favored by better exposure of the kinase target residues. These findings indicate that post-translational modification by acetylation is a critical mechanism that directly affects VEGFR2 function. PMID:24620033
Zecchin, Annalisa; Pattarini, Lucia; Gutierrez, Maria Ines; Mano, Miguel; Mai, Antonello; Valente, Sergio; Myers, Mike P; Pantano, Sergio; Giacca, Mauro
Distribution of acetylated alpha-tubulin in Physarum polycephalum
The expression and cytological distribution of acetylated alpha-tubulin was investigated in Physarum polycephalum. A monoclonal antibody specific for acetylated alpha-tubulin, 6-11B-1 (Piperno, G., and M. T. Fuller, 1985, J. Cell Biol., 101:2085-2094), was used to screen for this protein during three different stages of the Physarum life cycle-- the amoeba, the flagellate, and the plasmodium. Western blots of two- dimensional gels of amoebal and flagellate proteins reveal that this antibody recognizes the alpha 3 tubulin isotype, which was previously shown to be formed by posttranslational modification (Green, L. L., and W. F. Dove, 1984, Mol. Cell. Biol., 4:1706-1711). Double-label immunofluorescence demonstrates that, in the flagellate, acetylated alpha-tubulin is localized in the flagella and flagellar cone. Similar experiments with amoebae interestingly reveal that only within the microtubule organizing center (MTOC) are there detectable amounts of acetylated alpha-tubulin. In contrast, the plasmodial stage gives no evidence for acetylated alpha-tubulin by Western blotting or by immunofluorescence.
Acetylation-Dependent Regulation of Skp2 Function
SUMMARY Aberrant Skp2 signaling has been implicated as a driving event in tumorigenesis. Although the underlying molecular mechanisms remain elusive, cytoplasmic Skp2 correlates with more aggressive forms of breast and prostate cancers. Here, we report that Skp2 is acetylated by p300 at K68 and K71, which is a process that can be antagonized by the SIRT3 deacetylase. Inactivation of SIRT3 leads to elevated Skp2 acetylation, which leads to increased Skp2 stability through impairment of the Cdh1-mediated proteolysis pathway. As a result, Skp2 oncogenic function is increased, whereby cells expressing an acetylation-mimetic mutant display enhanced cellular proliferation and tumorigenesis in vivo. Moreover, acetylation of Skp2 in the nuclear localization signal (NLS) promotes its cytoplasmic retention, and cytoplasmic Skp2 enhances cellular migration through ubiquitination and destruction of E-cadherin. Thus, our study identifies an acetylation-dependent regulatory mechanism governing Skp2 oncogenic function and provides insight into how cytoplasmic Skp2 controls cellular migration.
Inuzuka, Hiroyuki; Gao, Daming; Finley, Lydia W.S.; Yang, Wen; Wan, Lixin; Fukushima, Hidefumi; Chin, Y. Rebecca; Zhai, Bo; Shaik, Shavali; Lau, Alan W.; Wang, Zhiwei; Gygi, Steven P.; Nakayama, Keiko; Teruya-Feldstein, Julie; Toker, Alex; Haigis, Marcia C.; Pandolfi, Pier Paolo; Wei, Wenyi
Starch hydrolysis modeling: application to fuel ethanol production
Efficiency of the starch hydrolysis in the dry grind corn process is a determining factor for overall conversion of starch\\u000a to ethanol. A model, based on a molecular approach, was developed to simulate structure and hydrolysis of starch. Starch structure\\u000a was modeled based on a cluster model of amylopectin. Enzymatic hydrolysis of amylose and amylopectin was modeled using a Monte
Ganti S. MurthyDavid; David B. Johnston; Kent D. Rausch; M. E. Tumbleson; Vijay Singh
Influence of relativistic effects on hydrolysis of Ra 2+
Summary��Using 224Ra radiotracer the first hydrolysis constant (pK1h) of Ra2+ cations has been determined. The pK1h value of Ra2+ was compared with the pK1h values of other Group 2 cations. It has been shown that the electrostatic hydrolysis model based on assumption that pK1h is a linear function of reciprocal ionic radii (1\\/ri) does not describe well the hydrolysis of
B. Zieli?ska; A. Bilewicz
Technical bases for precipitate hydrolysis process operating parameters
This report provides the experimental data and rationale in support of the operating parameters for tetraphenylborate precipitate hydrolysis specified in WSRC-RP-92-737. The report is divided into two sections, the first dealing with lab-scale precipitate hydrolysis experimentation while the second part addresses large-scale runs conducted to demonstrate the revised operating parameters in the Precipitate Hydrolysis Experimental Facility (PHEF). The program was in conjunction with reducing the nitrite ion level in DWPF feed.
Bannochie, C.J.; Lambert, D.P.
Technical bases for precipitate hydrolysis process operating parameters. Revision 1
This report provides the experimental data and rationale in support of the operating parameters for tetraphenylborate precipitate hydrolysis specified in WSRC-RP-92-737. The report is divided into two sections, the first dealing with lab-scale precipitate hydrolysis experimentation while the second part addresses large-scale runs conducted to demonstrate the revised operating parameters in the Precipitate Hydrolysis Experimental Facility (PHEF). The program was in conjunction with reducing the nitrite ion level in DWPF feed.
Bannochie, C.J.; Lambert, D.P.
Enhanced hydrolysis resistance of biodegradable polymers and bio-composites
This study investigated the hydrolysis of biodegradable polymers and bio-composites at 50�C and 90% relative humidity (RH). With increasing hydrolysis time, the mechanical properties of the biodegradable polymers and bio-composites significantly decreased due to the easy hydrolytic degradation of the ester linkage of the biodegradable polymers. With increasing hydrolysis time, the tensile strength of the polybutylene succinate (PBS) treated with
Hee-Soo Kim; Hyun-Joong Kim
Identification and preliminary characterization of acsF, a Putative Ni-insertase used in the biosynthesis of acetyl-CoA synthase from Clostridium thermoaceticum
OAK-B135 The acsABCDE genes in the Clostridium thermoaceticum genome are used for autotrophic acetyl-CoA synthesis using the Wood/Ljungdahl pathway. A 2.8 kb region between acsC and acsD was cloned and sequenced. Two open reading frames, orf7 ({approx} 1.9 kb) and acsF ({approx} 0.7 kb) were identified. orf7 appears to encode an Fe-S protein, in that it contains 5 conserved cysteine residues, 3 of which are present in a motif (CXXXXXCXXC) commonly used to coordinate Fe-S clusters. However, Orf7 is probably not involved in autotrophic acetyl-CoA synthesis, as homologous genes are present in organisms that do not utilize this pathway and are absent in many that do. In contrast, acsF is probably involved in this pathway. Sequence alignment of AcsF and 11 homologs reveals a number of conserved regions, including a P-loop that binds nucleoside triphosphates and catalyzes their hydrolysis. One homolog is CooC, an ATPase/GTPase that inserts Ni into a precursor form of the C-cluster of the carbon monoxide dehydrogenase (CODH) from Rhodospirillum rubrum. Purified AcsF lacked Ni and Fe, and slowly catalyzed the hydrolysis of ATP. Such similarities to CooC suggest that AcsF may function to insert Ni into a Ni-deficient form of the bifunctional acetyl-CoA synthase/CODH from C. thermoaceticum (ACSCt). However, this could not be established, as expression of acsF did not effect activation of recombinant AcsAB expressed in E. coli. Also, E. coli cells defective in hypB retained the ability to synthesize active recombinant AcsAB. Rather, the concentration of extracellular Ni2+ ions was critical to activation.
Huay-Keng Loke; Paul A. Lindahl
Studies on Cellulose Hydrolysis by Acetivibrio cellulolyticus�
Acetivibrio cellulolyticus extracellular cellulase extensively hydrolyzed crystalline celluloses such as Avicel (FMC Corp., Food and Pharmaceutical Products Div., Philadelphia, Pa.) but only if it was desalted and supplemented with Ca2+. The Ca2+ effect was one of increased enzyme stability in the presence of the ion. Although preincubation of the cellulase complex at 40�C for 5 h without added Ca2+ had a negligible effect on endoglucanase activity or on the subseqent hydrolysis of amorphous cellulose, the capacity of the enzyme to hydrolyze crystalline cellulose was almost completely lost. Adsorption studies showed that 90% of the Avicel-solubilizing component of the total enzyme preparation bound to 2% Avicel at 40�C. Under these conditions, only 15% of the endoglucanase and 25% of the protein present in the enzyme preparation adsorbed to the substrate. The protein profile of the bound enzyme, as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was complex and distinctly different from the profile observed for total cellulase preparations. The specific activity of A. cellulolyticus cellulase with respect to Avicel hydrolysis was compared with that of commercially available Trichoderma reesei cellulase. Images
MacKenzie, C. Roger; Bilous, Doris; Patel, Girishchandra B.
Acetylation is indispensable for p53 antiviral activity
Tumor suppressor p53 is known to be a direct transcriptional target of type I interferons (IFNs), contributing to virus-induced apoptosis, and in turn activating itself the interferon pathway. Acetylation, among many other post-translational modifications of p53, is thought to exert a crucial role regulating p53 activity. Here, we examined the contribution of this modification on the antiviral activity mediated by p53. Our results show that virus infection induces p53 acetylation at lysine 379, and that this modification is absolutely required for p53-dependent transcriptional transactivation of both, pro-apoptotic and IFN-stimulated genes induced by virus infection, and for p53-mediated control of virus replication. Thus, our study identifies p53 acetylation as an indispensable event that enables the p53-mediated antiviral response.
Munoz-Fontela, Cesar; Gonzalez, Dolores; Marcos-Villar, Laura; Campagna, Michela; Gallego, Pedro; Gonzalez-Santamaria, Jose; Herranz, Daniel; Gu, Wei; Serrano, Manuel; Aaronson, Stuart A.; Rivas, Carmen
Histone acetylation and its role in embryonic stem cell differentiation
The understanding of mechanisms leading to cellular differentiation is the main aim of numerous studies. Accessibility of DNA to transcription factors depends on local chromatin structure and chromatin compaction inhibits gene transcription. Histone acetylation correlates with an open chromatin structure and increased gene expression. Gene transcription levels are changed in early embryonic stem cells differentiation in a tissue-specific manner and epigenetic marks are modified, including increased global acetylation levels. Manipulation of histone deacetylases activity might be an interesting tool to generate populations of specific cell types for transplantation purposes. Thus, this review aims to show recent findings on histone acetylation, a post translational modification and its manipulation in embryonic stem cells differentiation.
Saraiva, Naiara Z; Oliveira, Clara S; Garcia, Joaquim M
Steady State Fluorescence Studies of Wild Type Recombinant Cinnamoyl CoA Reductase (Ll-CCRH1) and its Active Site Mutants.
Fluorescence quenching and time resolved fluorescence studies of wild type recombinant cinnamoyl CoA reductase (Ll-CCRH1), a multitryptophan protein from Leucaena leucocephala and 10 different active site mutants were carried out to investigate tryptophan environment. The enzyme showed highest affinity for feruloyl CoA (K a ?=?3.72?�?10(5)�M(-1)) over other CoA esters and cinnamaldehydes, as determined by fluorescence spectroscopy. Quenching of the fluorescence by acrylamide for wild type and active site mutants was collisional with almost 100�% of the tryptophan fluorescence accessible under native condition and remained same after denaturation of protein with 6�M GdnHCl. In wild type Ll-CCRH1, the extent of quenching achieved with iodide (f a?=?1.0) was significantly higher than cesium ions (f a?=?0.33) suggesting more density of positive charge around surface of trp conformers under native conditions. Denaturation of wild type protein with 6�M GdnHCl led to significant increase in the quenching with cesium (f a?=?0.54), whereas quenching with iodide ion was decreased (f a?=?0.78), indicating reorientation of charge density around trp from positive to negative and heterogeneity in trp environment. The Stern-Volmer plots for wild type and mutants Ll-CCRH1 under native and denatured conditions, with cesium ion yielded biphasic quenching profiles. The extent of quenching for cesium and iodide ions under native and denatured conditions observed in active site mutants was significantly different from wild type Ll-CCRH1 under the same conditions. Thus, single substitution type mutations of active site residues showed heterogeneity in tryptophan microenvironment and differential degree of conformation of protein under native or denatured conditions. PMID:24322526
Sonawane, Prashant; Vishwakarma, Rishi Kishore; Singh, Somesh; Gaikwad, Sushama; Khan, Bashir M
The influence of the primary and secondary xanthan structure on the enzymatic hydrolysis of the xanthan backbone.
Differently modified xanthans, varying in degree of acetylation and/or pyruvylation were incubated with the experimental cellulase mixture C1-G1 from Myceliophthora thermophila C1. The ionic strength and/or temperature of the xanthan solutions were varied, to obtain different xanthan conformations. The exact conformation at the selected incubation conditions was determined by circular dichroism. The xanthan degradation was analyzed by size exclusion chromatography. It was shown that at a fixed xanthan conformation, the backbone degradation by cellulases is equal for each type of xanthan. Complete backbone degradation is only obtained at a fully disordered conformation, indicating that only the secondary xanthan structure influences the final degree of hydrolysis by cellulases. It is thereby shown that, independently on the degree of substitution, xanthan can be completely hydrolyzed to oligosaccharides. These oligosaccharides can be used to further investigate the primary structure of different xanthans and to correlate the molecular structure to the xanthan functionalities. PMID:23911459
Kool, Marijn M; Schols, Henk A; Delahaije, Roy J B M; Sworn, Graham; Wierenga, Peter A; Gruppen, Harry
Wax Crystal-Sparse Leaf1 encodes a ?�ketoacyl CoA synthase involved in biosynthesis of cuticular waxes on rice leaf
Cuticular waxes, forming the plant\\/atmosphere interface of plants colonizing the terrestrial environment, are complex mixtures\\u000a of very-long chain fatty acids (VLCFAs) and their derivatives. In VLCFAs biosynthesis, ?-ketoacyl CoA synthase (E.C.220.127.116.11,\\u000a KCS) is the key enzyme. Using T-DNA insertional mutagenesis, we identified a cuticle-deficient rice mutant, which displayed\\u000a a pleiotropic phenotype including reduced growth, leaf fusion, sparse wax crystals, enhanced
Dongmei Yu; Kosala Ranathunge; Huasun Huang; Zhongyou Pei; Rochus Franke; Lukas Schreiber; Chaozu He
Specificity of antibodies to O-acetyl-positive and O-acetyl-negative group C meningococcal polysaccharides in sera from vaccinees and carriers.
Most group C Neisseria meningitidis strains produce an O-acetyl-positive polysaccharide, a homopolymer of alpha-2----9-linked N-acetylneuraminic acid with O-acetyl groups at the C-7 and C-8 of its sialic acid residues. The majority of disease isolates have been reported to contain this polysaccharide. Some strains produce group C polysaccharide lacking O-acetyl groups. The licensed vaccine contains the O-acetyl-positive polysaccharide. We have measured the antibody specificities to the two polysaccharides in sera from asymptomatic group C meningococcal carriers and vaccinated adults by a new enzyme-linked immunosorbent assay (ELISA) procedure using methylated human serum albumin for coating the group C polysaccharide onto microtiter plates. Inhibition of binding of serum antibodies to polysaccharide-coated plates was measured by ELISA after incubation with O-acetyl-positive and O-acetyl-negative group C polysaccharides. Greater inhibition of binding of carrier sera was observed with the homologous polysaccharide. There was substantial inhibition of binding of vaccinee sera to the O-acetyl-positive polysaccharide-coated plate following preincubation with O-acetyl-positive polysaccharide, but homologous inhibition on plates coated with the O-acetyl-negative polysaccharide required much higher concentrations of polysaccharide. Carrier sera may have a higher proportion of antibodies with greater specificity for the O-acetyl-negative polysaccharide, while vaccinee sera contain antibodies with greater affinity for the O-acetyl-positive polysaccharide. Studies with monoclonal antibodies specific for O-acetyl-positive and O-acetyl-negative polysaccharides reveal that the percentage of group C meningococcal disease caused by O-acetyl-negative strains remains about 15%, as found over 15 years ago.
Arakere, G; Frasch, C E
Predicting N-terminal acetylation based on feature selection method.
Methionine aminopeptidase and N-terminal acetyltransferase are two enzymes that contribute most to the N-terminal acetylation, which has long been recognized as a frequent and important kind of co-translational modifications [R.A. Bradshaw, W.W. Brickey, K.W. Walker, N-terminal processing: the methionine aminopeptidase and N alpha-acetyl transferase families, Trends Biochem. Sci. 23 (1998) 263-267]. The combined action of these two enzymes leads to two types of N-terminal acetylated proteins that are with/without the initiator methionine after the N-terminal acetylation. To accurately predict these two types of N-terminal acetylation, a new method based on feature selection has been developed. 1047 N-terminal acetylated and non-acetylated decapeptides retrieved from Swiss-Prot database (http://cn.expasy.org) are encoded into feature vectors by amino acid properties collected in Amino Acid Index database (http://www.genome.jp/aaindex). The Maximum Relevance Minimum Redundancy method (mRMR) combining with Incremental Feature Selection (IFS) and Feature Forward Selection (FFS) is then applied to extract informative features. Nearest Neighbor Algorithm (NNA) is used to build prediction models. Tested by Jackknife Cross-Validation, the correct rate of predictors reach 91.34% and 75.49% for each type, which are both better than that of 84.41% and 62.99% acquired by using motif methods [S. Huang, R.C. Elliott, P.S. Liu, R.K. Koduri, J.L. Weickmann, J.H. Lee, L.C. Blair, P. Ghosh-Dastidar, R.A. Bradshaw, K.M. Bryan, et al., Specificity of cotranslational amino-terminal processing of proteins in yeast, Biochemistry 26 (1987) 8242-8246; R. Yamada, R.A. Bradshaw, Rat liver polysome N alpha-acetyltransferase: substrate specificity, Biochemistry 30 (1991) 1017-1021]. Furthermore, the analysis of the informative features indicates that at least six downstream residues might have effect on the rules that guide the N-terminal acetylation, besides the penultimate residue. The software is available upon request. PMID:18533108
Cai, Yu-Dong; Lu, Lin
Interaction of RNA polymerase II with acetylated nucleosomal core particles
Chemical acetylation of nucleosomal cores is accompanied by an increase in their efficiency as in vitro transcription templates. Low amounts of acetic anhydride cause preferential modification of the amino-terminal tails of core histones. Modification of these domains, which causes moderate structural effects, is apparently correlated with the observed stimulation of RNA synthesis. In contrast, extensive modification of the globular regions of core histones, which is accompanied by a large structural relaxation of the particle, causes little additional effect on transcription. Acetylation of the amino-terminal domains of histones might stimulate transcription by changing the interaction of the histone tails with components of the transcriptional machinery.
Pineiro, M.; Gonzalez, P.J.; Hernandez, F.; Palacian, E. (Centro de Biologia Molecular, Universidad Autonoma de Madrid (Spain))
The Department of Energy (DOE) has one of the largest and most complete collections of information on crude oil composition that is available to the public. The computer program that manages this database of crude oil analyses has recently been rewritten to allow easier access to this information. This report describes how the new system can be accessed and how the information contained in the Crude Oil Analysis Data Bank can be obtained.
Fox, B.; Pautz, J.; Sellers, C.
Hydrolysis of polyesters by serine proteases.
The substrate specificity of alpha-chymotrypsin and other serine proteases, trypsin, elastase, proteinase K and subtilisin, towards hydrolysis of various polyesters was examined using poly(L-lactide) (PLA), poly(beta-hydroxybutyrate) (PHB), poly(ethylene succinate) (PES), poly(ethylene adipate) (PEA), poly(butylene succinate) (PBS), poly(butylene succinate-co-adipate) (PBS/A), poly[oligo(tetramethylene succinate)-co-(tetramethylane carbonate)] (PBS/C), and poly(epsilon-caprolactone) (PCL). alpha-Chymotrypsin could degrade PLA and PEA with a lower activity on PBS/A. Proteinase K and subtilisin degraded almost all substrates other than PHB. Trypsin and elastase had similar substrate specificities to alpha-chymotrypsin. PMID:15928850
Lim, Hyun-A; Raku, Takao; Tokiwa, Yutaka
nanoparticles fabricated by modified hydrolysis technique
NASA Astrophysics Data System (ADS)
We have tested modified hydrolysis method for the preparation of ?-Fe2O3 nanoparticles. The particles after synthesis were applied for a series of physicochemical techniques. Iron chloride was used as a precursor material. The particle size distribution was determined using zeta sizer and scanning electron microscopy. The surface area and the morphology of the particles vary by changing the concentration of the precursor material. The size of nanoparticles varies from 10 to 90 nm. The particles having size of 23 � 1 nm were separated out from the solution and their size remains almost the same even after one month. Energy dispersive X-ray analysis (EDX) of Fe2O3 nanoparticles confirms the purity of the desired material. The weight loss of the particles with respect to the temperature was studied by thermogravimetric and differential thermogravimetric (TG/DTG) analysis. X-ray diffraction (XRD) has been employed to study the crystallinity of the particles.
Waseem, Muhammad; Munsif, Sajida; Rashid, Umer; Imad-ud-Din
Transport and hydrolysis of peptides by microorganisms.
The structural specificities of the dipeptide and oligopeptide permeases of E. coli are briefly reviewed and related to the requirements found for other microorganisms. New, quick, sensitive methods for studying peptide transport are described, based on the following: (i) peptide-dependent incorporation of free radioactive amino acid into newly synthesized protein by a double amino acid auxotroph, (ii) colorimetric assay of peptide-dependent enzyme synthesis by an amino acid auxotroph, (iii) dansyl fingerprint technique. These approaches provide information on peptide binding affinity to a permease and rates of peptide uptake and amino acid efflux. Among current and future research areas considered are: the influence of the pKb of the N-terminal amino group on transport, generality of peptide transport in microorganisms, energy coupling and regulation, involvement of binding proteins, and the 'smugglin' concept. Peptide hydrolysis, and nutritional ultilization of peptides, by microorganisms are briefly discussed. PMID:340177
Payne, J W
Effect of sodium polyacrylate on the hydrolysis of octacalcium phosphate
Octacalcium phosphate (OCP) hydrolysis into hydroxyapatite (HA) has been investigated in aqueous solutions at different concentrations of sodium polyacrylate (NaPA). In the absence of the polyelectrolyte, OCP undergoes a complete transformation into HA in 48 h . The hydrolysis is inhibited by the polymer, which is significantly adsorbed on the crystals, up to about 22 wt.%. A polymer concentration of
Adriana Bigi; Elisa Boanini; Giuseppe Falini; Silvia Panzavolta; Norberto Roveri
Hydrolysis of oligosaccharides by cation exchanger silica gels
Sulfonic acid cation-exchange resin usually synthesized from styrene divinylbenzene copolymers are not well effective as catalysts of oligosaccharides hydrolysis. The present paper deals with the rate of hydrolysis of oligosaccharides on a new cation exchanger obtained by grafting sulfonic groups on porous silica. With the large pore sizes, the sieving effect of large solutes is suppressed and the catalytic activity
Alain Heyraud; Marguerite Rinaudo
Kinetic study of the acid hydrolysis of sugar cane bagasse
Economic interest in xylitol production can be enhanced if the needed xylose solutions can be obtained from the hydrolysis of low-cost lignocellulosic wastes. Sugar cane bagasse is a renewable, cheap and widely available waste in tropical countries. The hydrolysis of sugar cane bagasse to obtain xylose solutions has a double consequence, the elimination of a waste and the generation of
R Aguilar; J. A Ram??rez; G Garrote; M V�zquez
Continuous acid hydrolysis of waste cellulose for ethanol production
The continuous dilute acid hydrolysis of waste cellulose materials (paper pulp and wood sawdust at 10 and 95% solids, respectively) to glucose in a twin screw reactor gave high conversion yields and good energy efficiencies. A scheme for a scaled-up acid hydrolysis-fermentation distillation facility based on a waste cellulose feedstock to produce fuel-grade ethanol is proposed.
B. Rugg; W. Brenner
The laccase-catalyzed modification of lignin for enzymatic hydrolysis
The efficient use of cellulases in the hydrolysis of pretreated lignocellulosic biomass is limited due to the presence of lignin. Lignin is known to bind hydrolytic enzymes nonspecifically, thereby reducing their action on carbohydrate substrates. The composition and location of residual lignin therefore seem to be important for optimizing the enzymatic hydrolysis of lignocellulosic substrates. The use of lignin-modifying enzymes
Ulla Moilanen; Miriam Kellock; Sari Galkin; Liisa Viikari
Protein recovery from mechanically deboned turkey residue by enzymic hydrolysis
Enzymic hydrolysis was used to recover a potentially edible high protein hydrolysate from mechanically deboned turkey residue (MDTR), a turkey processing waste. The hydrolysis process reduced the weight of the original MDTR by 51% on a dry weight basis, and recovered 46% of the MDTR proteins. The hydrolysate contained 78% protein, 4�6% ash and only 5�7% fat. The proteins consisted
L. G. Fonkwe; R. K. Singh
Transcript assisted phosphodiester bond hydrolysis by eukaryotic RNA polymerase II
Hydrolysis of the phosphodiester bonds of the transcript by bacterial RNA polymerase is assisted by 3?NMP of the RNA. Here we provide evidence that this mechanism is also involved in RNA cleavage by eukaryotic RNA polymerase II, suggesting that transcript assisted hydrolysis has emerged before divergence of bacteria and archaea/eukaryotes.
Nielsen, Soren; Zenkin, Nikolay
Class Projects in Physical Organic Chemistry: The Hydrolysis of Aspirin
ERIC Educational Resources Information Center
An exercise that provides a hands-on demonstration of the hydrolysis of aspirin is presented. The key to understanding the hydrolysis is recognizing that all six process may occur simultaneously and that the observed rate constant is the sum of the rate constants that one rate constant dominates the overall process.
Marrs, Peter S.
Kinetic study of sphingomyelin hydrolysis for ceramide production
Kinetic study of sphingomyelin hydrolysis catalyzed by Clostridium perfringens phospholipase C was, at the first time, conducted for ceramide production. Ceramide has the major role in maintaining the water-retaining properties of the epidermis. Hence, it is of great commercial potential in cosmetic and pharmaceutical industries such as in hair and skin care products. The enzymatic hydrolysis of sphingomyelin has been
Long Zhang; Lars I. Hellgren; Xuebing Xu
In an attempt to elucidate the effect of microwave pretreatment (MP) on enzymatic hydrolysis of rice straw (RS), three distinct MP procedures, microwave treatment (MT) alone, MT and alkali treatment (AT) together, and MT after AT, were investigated and compared with conventional alkali pretreatment (AP) by measurement of reducing sugar during its enzymatic hydrolysis. The combination of MT and AT
Zhu Shengdong; Yu Ziniu; Wu Yuanxin; Zhang Xia; Li Hui; Gao Ming
A colorimetric assay for the determination of acetyl xylan esterase or cephalosporin C acetyl esterase activities using 7-amino cephalosporanic acid, cephalosporin C, or acetylated xylan as substrate.
A bromothymol blue-based colorimetric assay has been devised to screen for acetyl xylan esterase or cephalosporin C (CPC) deacetylase activities using 7-amino cephalosporanic acid (7-ACA), CPC, or acetylated xylan as substrate. These enzymes are not screened with their natural substrates because of the tedious procedures available previously. Acetyl xylan esterase from Bacillus pumilus CECT 5072 was cloned, expressed in Escherichia coli Rosetta (DE3), and characterized using this assay. Similar K(M) values for 7-ACA and CPC were obtained when compared with those described using HPLC methods. The assay is easy to perform and can be carried out in robotic high-throughput colorimetric devices normally used in directed evolution experiments. The assay allowed us to detect improvements in activity at a minimum of twofold with a very low coefficient of variance in 96-well plates. This method is significantly faster and more convenient to use than are known HPLC and pH-stat procedures. PMID:17651681
Mart�nez-Mart�nez, Irene; Montoro-Garc�a, Silvia; Lozada-Ram�rez, Jos� Daniel; S�nchez-Ferrer, Alvaro; Garc�a-Carmona, Francisco
Xylans inhibit enzymatic hydrolysis of lignocellulosic materials by cellulases.
Hemicelluloses have been found to be physical barriers in the hydrolysis of cellulose, and prevent the access of enzymes to cellulose surface. In addition, soluble hemicelluloses may strongly inhibit the cellulase activity. In this work, birchwood xylan clearly inhibited the enzymatic hydrolysis of wheat straw, Avicel and nanocellulose by cellulases. Hydrolysis efficiencies of cellobiohydrolase I (CBHI, from Thermoascus aurantiacus), cellobiohydrolase II (CBHII, from Trichoderma reesei) and endoglucanase II (from T. aurantiacus) were clearly inhibited by birchwood xylan, respectively. The strongest inhibitory effect of birchwood xylan was observed on the hydrolysis of Avicel by CBHI and CBHII, as a dramatically decreased formation of the main product, cellobiose. After additions of soluble and insoluble oat spelt xylan, cleaved cellobiose units by CBHI from cellulose chain decreased from 8 to 4 and 6, respectively. The results in this work demonstrated that xylans clearly inhibited the hydrolysis efficiencies of both endoglucanase and cellobiohydrolase. PMID:22858461
Zhang, Junhua; Tang, Ming; Viikari, Liisa
Improvement of thermal hydrolysis rate of dichloroacetic acid using alcohol.
Dichloroacetic acid (DCAA) is produced during the oxidation of trichloroethylene. It is also produced in drinking water treatment as a disinfection by-product. DCAA is a problem material, because of its toxicity. The objective of this research is to find the final products and the reaction pathway of the DCAA decomposition by hydrolysis, and to increase the hydrolysis rate. The removal of both chlorine atoms in DCAA structure was achieved with hydrolysis at around 75 degrees C, and the final products were oxalic acid and glycolic acid. The reaction pathway was the production of oxalic acid and glycolic acid from two glyoxylic acid molecules by Cannizzaro reaction after the glyoxylic acid production from dechlorination of DCAA with hydrolysis. The hydrolysis rate of DCAA was increased with the use of 90% ethanol solution as solvent. The activation energy of DCAA was about 80 kJ/mol in it, while it was around 105 kJ/mol in water. PMID:12892671
Okuda, Tetsuji; Nam, Se Yong; Lim, Jae Lim; Shin, Hang Sik
Modeling of autocatalytic hydrolysis of adefovir dipivoxil in solid formulations.
The stability and hydrolysis kinetics of a phosphate prodrug, adefovir dipivoxil, in solid formulations were studied. The stability relationship between five solid formulations was explored. An autocatalytic mechanism for hydrolysis could be proposed according to the kinetic behavior which fits the Prout-Tompkins model well. For the classical kinetic models could hardly describe and predict the hydrolysis kinetics of adefovir dipivoxil in solid formulations accurately when the temperature is high, a feedforward multilayer perceptron (MLP) neural network was constructed to model the hydrolysis kinetics. The build-in approaches in Weka, such as lazy classifiers and rule-based learners (IBk, KStar, DecisionTable and M5Rules), were used to verify the performance of MLP. The predictability of the models was evaluated by 10-fold cross-validation and an external test set. It reveals that MLP should be of general applicability proposing an alternative efficient way to model and predict autocatalytic hydrolysis kinetics for phosphate prodrugs. PMID:21467805
Dong, Ying; Zhang, Yan; Xiang, Bingren; Deng, Haishan; Wu, Jingfang
Factors affecting cellulose hydrolysis based on inactivation of adsorbed enzymes.
The rate of enzymatic hydrolysis of cellulose reaction is known to decrease significantly as the reaction proceeds. Factors such as reaction temperature, time, and surface area of substrate that affect cellulose conversion were analyzed relative to their role in a mechanistic model based on first order inactivation of adsorbed cellulases. The activation energies for the hydrolytic step and inactivation step were very close in magnitude: 16.3kcalmol(-1) for hydrolysis and 18.0kcalmol(-1) for inactivation, respectively. Therefore, increasing reaction temperature would cause a significant increase in the inactivation rate in addition to the catalytic reaction rate. Vmax,app was only 20% or less of the value at 72h compared to at 2h as a result of inactivation of adsorbed cellulases, suggesting prolonged hydrolysis is not an efficient way to improve cellulose hydrolysis. Hydrolysis rate increased with corresponding increases in available substrate surface binding area. PMID:25027809
Ye, Zhuoliang; Berson, R Eric
Sesame cake protein hydrolysis by alcalase: Effects of process parameters on hydrolysis, solubilisation, and enzyme inactivation
We investigated the effects of process parameters (substrate concentration, enzyme concentration, temperature and pH) on the\\u000a hydrolysis and solubilization of sesame cake protein as well as enzyme stability. The sesame cake protein was hydrolyzed by\\u000a Alcalase enzyme (a bacterial protease produced by a selected strain of Bacillus Licheniformis) that was chosen among five commercial enzymes examined. The optimum process conditions
El�in Demirhan; Dilek K?l?� Apar; Belma �zbek
Fully acetylated carbamate and hypotensive thiocarbamate glycosides from Moringa oleifera
Six new and three synthetically known glycosides have been isolated from the leaves of Moringa oleifera, employing a bioassay-directed isolation method on the ethanolic extract. Most of these compounds, bearing thiocarbamate, carbamate or nitrile groups, are fully acetylated glycosides, which are very rare in nature. Elucidation of the structures was made using chemical and spectroscopic methods, including 2D NMR techniques.
Shaheen Faizi; Bina Shaheen Siddiqui; Rubeena Saleem; Salimuzzaman Siddiqui; Khalid Aftab; Anwar-Ul-Hassan Gilani
Prebiotically plausible oligoribonucleotide ligation facilitated by chemoselective acetylation
NASA Astrophysics Data System (ADS)
The recent synthesis of pyrimidine ribonucleoside-2?,3?-cyclic phosphates under prebiotically plausible conditions has strengthened the case for the involvement of ribonucleic acid (RNA) at an early stage in the origin of life. However, a prebiotic conversion of these weakly activated monomers, and their purine counterparts, to the 3?,5?-linked RNA polymers of extant biochemistry has been lacking (previous attempts led only to short oligomers with mixed linkages). Here we show that the 2?-hydroxyl group of oligoribonucleotide-3?-phosphates can be chemoselectively acetylated in water under prebiotically credible conditions, which allows rapid and efficient template-directed ligation. The 2?-O-acetyl group at the ligation junction of the product RNA strand can be removed under conditions that leave the internucleotide bonds intact. Remarkably, acetylation of mixed oligomers that possess either 2?- or 3?-terminal phosphates is selective for the 2?-hydroxyl group of the latter. This newly discovered chemistry thus suggests a prebiotic route from ribonucleoside-2?,3?-cyclic phosphates to predominantly 3?,5?-linked RNA via partially 2?-O-acetylated RNA.
Bowler, Frank R.; Chan, Christopher K. W.; Duffy, Colm D.; Gerland, B�atrice; Islam, Saidul; Powner, Matthew W.; Sutherland, John D.; Xu, Jianfeng
Biodegradation of Acetylated Southern Pine and Aspen Composition Boards
The objective of this study was to investigate the influence of the acetylation treated wood fiber, Phenol-formaldehyde resin content level, two wood fiber species, three fungi species on the dimensional stability and decay resistance of high density composition boards. A standard ASTM method was used to evaluate weight loss and thickness change. Thc linear shrinkage and expansion of each species
Poo Chow; T. Harp; R. Meimban; J. A. Youngquist; R M. Rowell
Characterization of an acetylated heteroxylan from Eucalyptus globulus Labill.
A heteroxylan was isolated from Eucalyptus globulus wood by extraction of peracetic acid delignified holocellulose with dimethyl sulfoxide. Besides (1-->4)-linked beta-D-xylopyranosyl units of the backbone and short side chains of terminal (1-->2)-linked 4-O-methyl-alpha-D-glucuronosyl residues (MeGlcA) in a 1:10 molar ratio, this hemicellulose contained galactosyl and glucosyl units attached at O-2 of MeGlcA originating from rhamnoarabinogalactan and glucan backbones, respectively. About 30% of MeGlcA units were branched at O-2. The O-acetyl-(4-O-methylglucurono)xylan showed an acetylation degree of 0.61, as determined by 1H NMR spectroscopy, and a weight-average molecular weight (M(w)) of about 36 kDa (P=1.05) as revealed from size-exclusion chromatography (SEC) analysis. About half of the beta-D-xylopyranosyl units of the backbone were found as acetylated moieties at O-3 (34 mol%), O-2 (15 mol%) or O-2,3 (6 mol%). Practically, all beta-D-xylopyranosyl units linked at O-2 with MeGlcA residues were 3-O-acetylated (10 mol%). PMID:12644372
Evtuguin, Dmitry V; Tom�s, Jorge L; Silva, Artur M S; Neto, Carlos Pascoal
Prebiotically plausible oligoribonucleotide ligation facilitated by chemoselective acetylation
The recent synthesis of pyrimidine ribonucleoside-2?,3?-cyclic phosphates under prebiotically plausible conditions has strengthened the case for the involvement of RNA at an early stage in the origin of life. However, a prebiotic conversion of these weakly activated monomers, and their purine counterparts, to the 3?,5?-linked RNA polymers of extant biochemistry has been lacking � previous attempts leading only to short oligomers with mixed linkages. Here we show that the 2?-hydroxyl group of oligoribonucleotide-3?-phosphates can be chemoselectively acetylated in water under prebiotically credible conditions, allowing rapid and efficient template-directed ligation. The 2?-O-acetyl group at the ligation junction of the product RNA strand can be removed under conditions that leave the internucleotide bonds intact. Remarkably, acetylation of mixed oligomers possessing either 2?- or 3?-terminal phosphates is selective for the 2?-hydroxyl group of the latter. This newly discovered chemistry thus suggests a prebiotic route from ribonucleoside-2?,3?-cyclic phosphates to predominantly 3?,5?-linked RNA via partially 2?-O-acetylated-RNA.
Bowler, Frank R.; Chan, Christopher K. W.; Duffy, Colm D.; Gerland, Beatrice; Islam, Saidul; Powner, Matthew W.; Sutherland, John D.; Xu, Jianfeng
Genetic control of differential acetylation in diabetic rats.
Post-translational protein modifications such as acetylation have significant regulatory roles in metabolic processes, but their relationship to both variation in gene expression and DNA sequence is unclear. We address this question in the Goto-Kakizaki (GK) rat inbred strain, a model of polygenic type 2 diabetes. Expression of the NAD-dependent deacetylase Sirtuin-3 is down-regulated in GK rats compared to normoglycemic Brown Norway (BN) rats. We show first that a promoter SNP causes down-regulation of Sirtuin-3 expression in GK rats. We then use mass-spectrometry to identify proteome-wide differential lysine acetylation of putative Sirtuin-3 protein targets in livers of GK and BN rats. These include many proteins in pathways connected to diabetes and metabolic syndrome. We finally sequence GK and BN liver transcriptomes and find that mRNA expression of these targets does not differ significantly between GK and BN rats, in contrast to other components of the same pathways. We conclude that physiological differences between GK and BN rats are mediated by a combination of differential protein acetylation and gene transcription and that genetic variation can modulate acetylation independently of expression. PMID:24743600
Kaisaki, Pamela J; Otto, Georg W; McGouran, Joanna F; Toubal, Amine; Argoud, Kar�ne; Waller-Evans, Helen; Finlay, Clare; Cald�rari, Sophie; Bihoreau, Marie-Th�r�se; Kessler, Benedikt M; Gauguier, Dominique; Mott, Richard
MICROBIOLOGY: Bacteria Seize Control by Acetylating Host Proteins
NSDL National Science Digital Library
Access to the article is free, however registration and sign-in are required. The plague-causing bacterium Yersinia pestis injects toxic proteins into its hosts' cells. One of these interferes with the host's secretion of a protective factor by adding acetyl groups to a signaling kinase, blocking its activation.
Carolyn A. Worby (University of California ;Departments of Pharmacology, Cellular and Molecular Medicine, and Chemistry and Biochemistry); Jack E. Dixon (University of California ;Departments of Pharmacology, Cellular and Molecular Medicine, and Chemistry and Biochemistry)
Characterization of two long-chain fatty acid CoA ligases in the Gram-positive bacterium Geobacillus thermodenitrificans NG80-2.
The functions of two long-chain fatty acid CoA ligase genes (facl) in crude oil-degrading Geobacillus thermodenitrificans NG80-2 were characterized. Facl1 and Facl2 encoded by GTNG_0892 and GTNG_1447 were expressed in Escherichia coli and purified as His-tagged fusion proteins. Both enzymes utilized a broad range of fatty acids ranging from acetic acid (C(2)) to melissic acid (C(30)). The most preferred substrates were capric acid (C(10)) for Facl1 and palmitic acid (C(16)) for Facl2, respectively. Both enzymes had an optimal temperature of 60�C, an optimal pH of 7.5, and required ATP as a cofactor. Thermostability of the enzymes and effects of metal ions, EDTA, SDS and Triton X-100 on the enzyme activity were also investigated. When NG80-2 was cultured with crude oil rather than sucrose as the sole carbon source, upregulation of facl1 and facl2 mRNA was observed by real time RT-PCR. This is the first time that the activity of fatty acid CoA ligases toward long-chain fatty acids up to at least C(30) has been demonstrated in bacteria. PMID:22694860
Dong, Yanpeng; Du, Huiqian; Gao, Chunxu; Ma, Ting; Feng, Lu
Antidiabetic and antihyperlipidemic activity of asiatic acid in diabetic rats, role of HMG CoA: in vivo and in silico approaches.
Hyperlipidemia is an associated complication of diabetes and also a major risk factor for cardiovascular diseases. The present study was designed to examine the antihyperlipidemic effect of asiatic acid (AA) in streptozotocin (STZ) induced diabetic rats. Diabetes was induced in male Wistar rats by a single intraperitoneal injection of STZ (40 mg/kg b.w.). Diabetic rats show increased plasma glucose, total cholesterol, triglycerides, free fatty acids, phospholipids, low density lipoprotein, very low density liprotein, atherogenic index and decreased insulin and high density lipoprotein in diabetic rats. The activity of 3-hydroxy 3-methylglutaryl coenzyme A (HMG CoA) reductase increased significantly in contrast to the activities of lipoprotein lipase and lecithin cholesterol acyltransferase. In addition, the molecular docking of AA against HMG CoA reductase involved in cholesterol biosynthesis using Argus software. Diabetic rats were treated with AA shifted all these parameters towards normalcy. AA has shown best ligand binding energy 11.8122 kcal/mol. The antihyperlipidemic effect of AA was compared with glibenclamide; a well-known antihyperglycemic drug. In conclusion, this study indicates that AA showed an antihyperlipidemic effect in addition to its antidiabetic effect in experimental diabetes. PMID:24075211
Ramachandran, Vinayagam; Saravanan, Ramalingam; Senthilraja, Poomalai
Regioselective enzymatic acetylation of the aglycone moiety of a secoiridoid glucoside. Two new secoiridoid glucoside acetates.
Candida antarctica lipase (CAL) catalyses the regioselective acetylation of the 10-hydroxyl group of 10-hydroxyoleoside dimethyl ester, a secoiridoid glucoside, using THF as a solvent and ethyl acetate or vinyl acetate as acetyl group suppliers. Two acetyl derivatives at 3'- and 6'-sites of the glucosidic ring of 10-acetoxyoleoside dimethylester, not previously described, were obtained by acetylation in the same conditions. PMID:15340202
P�rez, Jos� Andr�s; Boluda, Carlos; L�pez, Hermelo; Trujillo, Juan Manvel; Hern�ndez, Jos� Manuel
Biosynthetic preparation of the NO-glucosiduronic acid of N-acetyl-N-phenylhydroxylamine
1. Sodium (N-acetyl-N-phenylhydroxylamine ?-d-glucosid)uronate was isolated from the urine of rabbits receiving N-acetyl-N-phenylhydroxylamine. 2. Its chemical structure was confirmed by the correspondence of the infrared spectrum of its tri-O-acetyl methyl ester derivative with the tri-O-acetyl methyl ester derivative of an authentic specimen prepared by the Koenigs�Knorr synthesis.
Kato, Keitaro; Ide, Hiroyuki; Hirohata, Itsuyo; Fishman, William H.
Cohesin Acetylation Promotes Sister Chromatid Cohesion Only in Association with the Replication Machinery*
Acetylation of the Smc3 subunit of cohesin is essential to establish functional cohesion between sister chromatids. Smc3 acetylation is catalyzed by members of the Eco family of acetyltransferases, although the mechanism by which acetylation is regulated and how it promotes cohesion are largely unknown. In vertebrates, the cohesin complex binds to chromatin during mitotic exit and is converted to a functional form during or shortly after DNA replication. The conserved proliferating cell nuclear antigen-interacting protein box motif in yeast Eco1 is required for function, and cohesin is acetylated during the S phase. This has led to the notion that acetylation of cohesin is stimulated by interaction of Eco1 with the replication machinery. Here we show that in vertebrates Smc3 acetylation occurs independently of DNA replication. Smc3 is readily acetylated before replication is initiated and after DNA replication is complete. However, we also show that functional acetylation occurs only in association with the replication machinery: disruption of the interaction between XEco2 and proliferating cell nuclear antigen prevents cohesion establishment while having little impact on the overall levels of Smc3 acetylation. These results demonstrate that Smc3 acetylation can occur throughout interphase but that only acetylation in association with the replication fork promotes sister chromatid cohesion. These data reveal how the generation of cohesion is limited to the appropriate time and place during the cell cycle and provide insight into the mechanism by which acetylation ensures cohesion.
Song, Jianhua; Lafont, Andrea; Chen, Jingrong; Wu, Frank M.; Shirahige, Katsuhiko; Rankin, Susannah
Specific Acetylation of Chromosomal Protein HMG-17 by PCAF Alters Its Interaction with Nucleosomes
Nonhistone chromosomal proteins HMG-14 and HMG-17 are closely related nucleosomal binding proteins that unfold the higher-order chromatin structure, thereby enhancing the transcription and replication potential of chromatin. Here we report that PCAF, a transcription coactivator with intrinsic histone acetyltransferase activity, specifically acetylates HMG-17 but not HMG-14. Using mass spectrum sequence analysis, we identified the lysine at position 2 as the predominant site acetylated by PCAF. Lysine 2 is a prominent acetylation site in vivo, suggesting that this PCAF-mediated acetylation is physiologically relevant. Experiments with HMG-17 deletion mutants and competition studies with various protein fragments indicate that the specific acetylation of HMG-17 is not determined solely by the primary sequence near the acetylation site. By equilibrium dialysis we demonstrated that acetylation reduces the affinity of HMG-17 to nucleosome cores. In addition, we found that the binding of HMG-14 and HMG-17 to nucleosome cores inhibits the PCAF-mediated acetylation of histone H3. Thus, the presence of HMG-14 and HMG-17 affects the ability of PCAF to acetylate chromatin, while the acetylation of HMG-17 reduces its binding affinity to chromatin. Conceivably, in HMG-17-containing chromatin, acetylation of HMG-17 precedes the acetylation of histones.
Herrera, Julio E.; Sakaguchi, Kazuyasu; Bergel, Michael; Trieschmann, Lothar; Nakatani, Yoshihiro; Bustin, Michael
Human acetylator polymorphism: estimate of allele frequency in Libya and details of global distribution.
Acetylator phenotyping by means of a sulphadimidine tests revealed 65% of Libyan Arabs to be slow acetylators. Hence the frequency of the allele controlling slow acetylation (As) is estimated as q = 0.81 +/- 0.05. This estimate is similar to those previously recorded in European and adjacent Middle Eastern populations.
Karim, A K; Elfellah, M S; Evans, D A
Combined deacetylation and PFI refining pretreatment of corn cob for the improvement of a two-stage enzymatic hydrolysis.
A combined deacetylation and PFI refining pretreatment was applied to corn cob for the improvement of a two-stage enzymatic hydrolysis. In stage 1, the pretreated corn cob was first hydrolyzed by xylanase to produce xylo-oligosaccharides (XOS). In stage 2, the solid residue isolated from stage 1 was further hydrolyzed by cellulase and ?-glucosidase. NaOH, Na2CO3, and Ca(OH)2 were tested to remove acetyl groups in the process of deacetylation, and it was found that Ca(OH)2 could be the most suitable alkali for deacetylation in this work. After deacetylation using 0.8 mmol of Ca(OH)2/g of substrate and PFI refining, 50.5% xylan in the raw material could be hydrolyzed into XOS. The corresponding xylan yield of stage 1, the glucan yield of stage 2, and the total sugar yield (all sugars released in the hydrolyzate) after the two-stage enzymatic hydrolysis were 0.306, 0.305, and 0.661 g/g of corn cob, respectively. PMID:24810587
Zhang, Yuedong; Mu, Xindong; Wang, Haisong; Li, Bin; Peng, Hui
Relativistic Density Functional Theory Calculations of the Electron Paramagnetic Resonance Parameters for Vanadyl Acetyl Acetonate and Copper Acetyl Acetonate
NASA Astrophysics Data System (ADS)
Relativistic density functional theory calculations of electron paramagnetic resonance (EPR) parameters using a variety of basis sets have been computed for the model systems Vanadyl acetyl acetonate and Copper acetyl acetonate using the ORCA program. The basis set dependence of g and A tensor calculations for Vanadyl acetyl acetonate and Copper acetyl acetonate were studied using Pople Style and Ahlrichs basis sets in Local and gradient corrected functionals (BP86 and PWP) and Hybrid functionals (B3LYP and PW1PW). The PW1PW hybrid functional gives the best values for VO(acac)2 using the TZV basis set and for Cu(acac)2 using the 6-311G(d) basis set. The calculated A values with PW1PW hybrid functional for VO(acac)2 and Local and gradient corrected functional (BP86) for Cu(acac)2 with same basis set (DZ) give better results than previously reported values using the Amsterdam Density Functional Theory (ADF) Software. Our calculated g and A tensor values are in good agreement with the values determined from experiment.
Mainali, Laxman; Sahu, Indra; Earle, Keith
Protein release from galactoglucomannan hydrogels: influence of substitutions and enzymatic hydrolysis by beta-mannanase.
O-Acetyl-galactoglucomannan (AcGGM) is the major soft-wood hemicellulose. Structurally modified AcGGM and hydrogels of AcGGM were prepared. The degree of substitution (DS) of AcGGM was modified enzymatically with alpha-galactosidase, and chemically with an acrylate derivative, 2-hydroxyethylmethacrylate (HEMA). The hydrolysis of AcGGM with beta-mannanase was shown to increase with decreasing DS. AcGGM hydrogels were prepared from chemically modified AcGGM with varying DS of HEMA. Bovine serum albumin (BSA) was encapsulated in hydrogels. A spontaneous burst release of BSA was decreased with increased DS of HEMA. The addition of beta-mannanase significantly enhanced the BSA release from hydrogels with a DS of 0.36, reaching a maximum of 95% released BSA after eight hours compared to 60% without enzyme. Thus, both the pendant group composition and the enzyme action are valuable tools in the tailoring of hydrogel release profiles of potential interest for intestine drug delivery. PMID:18590309
Roos, Alexandra Andersson; Edlund, Ulrica; Sj�berg, John; Albertsson, Ann-Christine; St�lbrand, Henrik
Cladosporium fulvum Avr4 protects fungal cell walls against hydrolysis by plant chitinases accumulating during infection.
Resistance against the leaf mold fungus Cladosporium fulvum is mediated by the tomato Cf proteins which belong to the class of receptor-like proteins and indirectly recognize extracellular avirulence proteins (Avrs) of the fungus. Apart from triggering disease resistance, Avrs are believed to play a role in pathogenicity or virulence of C. fulvum. Here, we report on the avirulence protein Avr4, which is a chitin-binding lectin containing an invertebrate chitin-binding domain (CBM14). This domain is found in many eukaryotes, but has not yet been described in fungal or plant genomes. We found that interaction of Avr4 with chitin is specific, because it does not interact with other cell wall polysaccharides. Avr4 binds to chitin oligomers with a minimal length of three N-acetyl glucosamine residues. In vitro, Avr4 protects chitin against hydrolysis by plant chitinases. Avr4 also binds to chitin in cell walls of the fungi Trichoderma viride and Fusarium solani f. sp. phaseoli and protects these fungi against normally deleterious concentrations of plant chitinases. In situ fluorescence studies showed that Avr4 also binds to cell walls of C. fulvum during infection of tomato, where it most likely protects the fungus against tomato chitinases, suggesting that Avr4 is a counter-defensive virulence factor. PMID:17153926
van den Burg, Harrold A; Harrison, Stuart J; Joosten, Matthieu H A J; Vervoort, Jacques; de Wit, Pierre J G M
Effect of hydrolysis on identifying prenatal cannabis exposure
Identification of prenatal cannabis exposure is important due to potential cognitive and behavioral consequences. A two-dimensional gas chromatography�mass spectrometry method for cannabinol, ?9-tetrahydrocannabinol (THC), 11-hydroxy-THC (11-OH-THC), 8?,11-dihydroxy-THC, and 11-nor-9-carboxy-THC (THCCOOH) quantification in human meconium was developed and validated. Alkaline, enzymatic, and enzyme�alkaline tandem hydrolysis conditions were optimized with THC- and THCCOOH-glucuronide reference standards. Limits of quantification ranged from 10 to 15 ng/g, and calibration curves were linear to 500 ng/g. Bias and intra-day and inter-day imprecision were <12.3%. Hydrolysis efficiencies were analyte-dependent; THC-glucuronide was effectively cleaved by enzyme, but not base. Conversely, THCCOOH-glucuronide was most sensitive to alkaline hydrolysis. Enzyme�alkaline tandem hydrolysis maximized efficiency for both glucuronides. Identification of cannabinoid-positive meconium specimens nearly doubled following alkaline and enzyme�alkaline hydrolysis. Although no 11-OH-THC glucuronide standard is available, enzymatic hydrolysis improved 11-OH-THC detection in authentic specimens. Maximal identification of cannabis-exposed neonates and the widest range of cannabis biomarkers are achieved with enzyme�alkaline tandem hydrolysis.
Gray, Teresa R.; Barnes, Allan J.
Factors impeding enzymatic wheat gluten hydrolysis at high solid concentrations.
Enzymatic wheat gluten hydrolysis at high solid concentrations is advantageous from an environmental and economic point of view. However, increased wheat gluten concentrations result in a concentration effect with a decreased hydrolysis rate at constant enzyme-to-substrate ratios and a decreased maximum attainable degree of hydrolysis (DH%). We here identified the underlying factors causing the concentration effect. Wheat gluten was hydrolyzed at solid concentrations from 4.4% to 70%. The decreased hydrolysis rate was present at all solid concentrations and at any time of the reaction. Mass transfer limitations, enzyme inhibition and water activity were shown to not cause this hydrolysis rate limitation up to 50% solids. However, the hydrolysis rate limitation can be, at least partly, explained by a second-order enzyme inactivation process. Furthermore, mass transfer impeded the hydrolysis above 60% solids. Addition of enzyme after 24?h at high solid concentrations scarcely increased the DH%, suggesting that the maximum attainable DH% decreases at high solid concentrations. Reduced enzyme activities caused by low water activities can explain this DH% limitation. Finally, a possible influence of the plastein reaction on the DH% limitation is discussed. Biotechnol. Bioeng. 2014;111: 1304-1312. � 2014 Wiley Periodicals, Inc. PMID:24474643
Hardt, N A; Janssen, A E M; Boom, R M; van der Goot, A J
Acetyl Coenzyme A Synthetase Is Acetylated on Multiple Lysine Residues by a Protein Acetyltransferase with a Single Gcn5-Type N-Acetyltransferase (GNAT) Domain in Saccharopolyspora erythraea.
Reversible lysine acetylation (RLA) is used by cells of all domains of life to modulate protein function. To date, bacterial acetylation/deacetylation systems have been studied in a few bacteria (e.g., Salmonella enterica, Bacillus subtilis, Escherichia coli, Erwinia amylovora, Mycobacterium tuberculosis, and Geobacillus kaustophilus), but little is known about RLA in antibiotic-producing actinomycetes. Here, we identify the Gcn5-like protein acetyltransferase AcuA of Saccharopolyspora erythraea (SacAcuA, SACE_5148) as the enzyme responsible for the acetylation of the AMP-forming acetyl coenzyme A synthetase (SacAcsA, SACE_2375). Acetylated SacAcsA was deacetylated by a sirtuin-type NAD(+)-dependent consuming deacetylase (SacSrtN, SACE_3798). In vitro acetylation/deacetylation of SacAcsA enzyme was studied by Western blotting, and acetylation of lysine residues Lys(237), Lys(380), Lys(611), and Lys(628) was confirmed by mass spectrometry. In a strain devoid of SacAcuA, none of the above-mentioned Lys residues of SacAcsA was acetylated. To our knowledge, the ability of SacAcuA to acetylate multiple Lys residues is unique among AcuA-type acetyltransferases. Results from site-specific mutagenesis experiments showed that the activity of SacAcsA was controlled by lysine acetylation. Lastly, immunoprecipitation data showed that in vivo acetylation of SacAcsA was influenced by glucose and acetate availability. These results suggested that reversible acetylation may also be a conserved regulatory posttranslational modification strategy in antibiotic-producing actinomycetes. PMID:24957627
You, Di; Yao, Li-Li; Huang, Dan; Escalante-Semerena, Jorge C; Ye, Bang-Ce
Alkaline phosphatase revisited: hydrolysis of alkyl phosphates.
Escherichia coli alkaline phosphatase (AP) is the prototypical two metal ion catalyst with two divalent zinc ions bound approximately 4 A apart in the active site. Studies spanning half a century have elucidated many structural and mechanistic features of this enzyme, rendering it an attractive model for investigating the potent catalytic power of bimetallic centers. Unfortunately, fundamental mechanistic features have been obscured by limitations with the standard assays. These assays generate concentrations of inorganic phosphate (P(i)) in excess of its inhibition constant (K(i) approximately 1 muM). This tight binding by P(i) has affected the majority of published kinetic constants. Furthermore, binding limits k(cat)/K(m) for reaction of p-nitrophenyl phosphate, the most commonly employed substrate. We describe a sensitive (32)P-based assay for hydrolysis of alkyl phosphates that avoids the complication of product inhibition. We have revisited basic mechanistic features of AP with these alkyl phosphate substrates. The results suggest that the chemical step for phosphorylation of the enzyme limits k(cat)/K(m). The pH-rate profile and additional results suggest that the serine nucleophile is active in its anionic form and has a pK(a) of < or = 5.5 in the free enzyme. An inactivating pK(a) of 8.0 is observed for binding of both substrates and inhibitors, and we suggest that this corresponds to ionization of a zinc-coordinated water molecule. Counter to previous suggestions, inorganic phosphate dianion appears to bind to the highly charged AP active site at least as strongly as the trianion. The dependence of k(cat)/K(m) on the pK(a) of the leaving group follows a Br�nsted correlation with a slope of beta(lg) = -0.85 +/- 0.1, differing substantially from the previously reported value of -0.2 obtained from data with a less sensitive assay. This steep leaving group dependence is consistent with a largely dissociative transition state for AP-catalyzed hydrolysis of phosphate monoesters. The new (32)P-based assay employed herein will facilitate continued dissection of the AP reaction by providing a means to readily follow the chemical step for phosphorylation of the enzyme. PMID:11863460
O'Brien, Patrick J; Herschlag, Daniel
Kinetics of iodine hydrolysis in unbuffered solutions
The kinetics of hydrolysis or disproportionation of hypoiodite were studied spectrophotometrically in basic solution at an ionic strength of 0.2 M as a function of pH, iodide and total iodine concentration, and temperature. The existence of three independent pathways for this second-order process was confirmed. The pH-stat method was used to monitor the corresponding reaction of hypoiodous acid in weakly alkaline solution. The generalized rate law for the disproportionation is: /minus/d((HOI) + (OI/sup /minus//))dt = k /sub a/(HOI)/sup 2/ + k/sub b/(HOI) (OI/sup /minus//) + k/sub c/(OI/sup /minus//)/sup 2/ + k/sub d/(I/sub 2/OH/sup /minus//) (OI/sup /minus//). The values of k/sub a/ and k/sub b/ are substantially smaller than previously reported. However, an unexplained contribution to the rate law resulting from the pH-stat measurements was also obtained. The rapid recombination of iodide and iodate in HClO/sub 4/ solutions was followed by stopped-flow spectrophotometry at three ionic strengths, and over a range of iodide and hydrogen ion concentrations, and at eight temperatures. Fifth-order kinetics were observed with no detectable induction period. 14 refs., 4 figs., 1 tab.
Palmer, D.A.; Lyons, L.J.
Microbial hydrolysis of organic nitriles and amides.
Nitrile-hydrating enzymes produced by bacteria and fungi catalyse the conversion of a large number of chemically diverse nitriles, including many economically important compounds used industrially for chemical synthesis of amides and acids. This paper presents data on two new, highly different nitrile-hydrolysing enzymes which were isolated in connection with our studies on enzymic nitrile transformations. Particular attention was paid to the enzymes' substrate specificities and sensitivity to substrate/product inhibition. One of our microbial isolates was a Rhodococcus sp. (strain CH5). This strain produces a constitutive hydratase that has a broad substrate spectrum, including aliphatic and aromatic nitriles, mononitriles and dinitriles, hydroxynitriles and amino-nitriles. It also produces a constitutive amidase of equally low substrate specificity. The hydratase/amidase system catalysed the hydrolysis of D,L-aminonitriles into racemic mixtures of amino acids. Strain CH5 is able to produce high concentrations of malonic acid monoamide from malononitrile and malonamide. The other isolate, Alcaligenes sp. (strain I4), can convert high concentrations of cyanoacetate into malonic acid, presumably by means of an aliphatic nitrilase that is specific for cyanoacetate. Enzyme kinetic experiments have shown that this enzyme is very resistant to both substrate and product inhibition. PMID:3073055
Ingvorsen, K; Yde, B; Godtfredsen, S E; Tsuchiya, R T
Synthesis, hydrolysis and stability of psilocin glucuronide.
A two-step synthesis of psilocin glucuronide (PCG), the main metabolite of psilocin, with methyl 2,3,4-tri-O-isobutyryl-1-O-trichloroacetimidoyl-?-d-glucopyranuronate is reported. With the synthesized PCG, hydrolysis conditions in serum and urine were optimized. Escherichia coli proved to be a better enzyme source for ?-glucuronidase than Helix pomatia. It was essential to add ascorbic acid to serum samples to protect psilocin during incubation. Furthermore the stability of PCG and psilocin was compared as stability data are the basis for forensic interpretation of measurements. PCG showed a greater long-term stability after six months in deep frozen serum and urine samples than psilocin. The short-term stability of PCG for one week in whole blood at room temperature and in deep frozen samples was also better than that of psilocin. Therefore, PCG can be considered to be more stable than the labile psilocin and should always be included if psilocin is analyzed in samples. PMID:24513688
Martin, Rafaela; Sch�renkamp, Jennifer; Pfeiffer, Heidi; Lehr, Matthias; K�hler, Helga
Acetylation of malate dehydrogenase 1 promotes adipogenic differentiation via activating its enzymatic activity
Acetylation is one of the most crucial post-translational modifications that affect protein function. Protein lysine acetylation is catalyzed by acetyltransferases, and acetyl-CoA functions as the source of the acetyl group. Additionally, acetyl-CoA plays critical roles in maintaining the balance between carbohydrate metabolism and fatty acid synthesis. Here, we sought to determine whether lysine acetylation is an important process for adipocyte differentiation. Based on an analysis of the acetylome during adipogenesis, various proteins displaying significant quantitative changes were identified by LC-MS/MS. Of these identified proteins, we focused on malate dehydrogenase 1 (MDH1). The acetylation level of MDH1 was increased up to 6-fold at the late stage of adipogenesis. Moreover, overexpression of MDH1 in 3T3-L1 preadipocytes induced a significant increase in the number of cells undergoing adipogenesis. The introduction of mutations to putative lysine acetylation sites showed a significant loss of the ability of cells to undergo adipogenic differentiation. Furthermore, the acetylation of MDH1 dramatically enhanced its enzymatic activity and subsequently increased the intracellular levels of NADPH. These results clearly suggest that adipogenic differentiation may be regulated by the acetylation of MDH1 and that the acetylation of MDH1 is one of the cross-talk mechanisms between adipogenesis and the intracellular energy level.
Kim, Eun Young; Kim, Won Kon; Kang, Hyo Jin; Kim, Jeong-Hoon; Chung, Sang J.; Seo, Yeon Soo; Park, Sung Goo; Lee, Sang Chul; Bae, Kwang-Hee
Acetylation of malate dehydrogenase 1 promotes adipogenic differentiation via activating its enzymatic activity.
Acetylation is one of the most crucial post-translational modifications that affect protein function. Protein lysine acetylation is catalyzed by acetyltransferases, and acetyl-CoA functions as the source of the acetyl group. Additionally, acetyl-CoA plays critical roles in maintaining the balance between carbohydrate metabolism and fatty acid synthesis. Here, we sought to determine whether lysine acetylation is an important process for adipocyte differentiation. Based on an analysis of the acetylome during adipogenesis, various proteins displaying significant quantitative changes were identified by LC-MS/MS. Of these identified proteins, we focused on malate dehydrogenase 1 (MDH1). The acetylation level of MDH1 was increased up to 6-fold at the late stage of adipogenesis. Moreover, overexpression of MDH1 in 3T3-L1 preadipocytes induced a significant increase in the number of cells undergoing adipogenesis. The introduction of mutations to putative lysine acetylation sites showed a significant loss of the ability of cells to undergo adipogenic differentiation. Furthermore, the acetylation of MDH1 dramatically enhanced its enzymatic activity and subsequently increased the intracellular levels of NADPH. These results clearly suggest that adipogenic differentiation may be regulated by the acetylation of MDH1 and that the acetylation of MDH1 is one of the cross-talk mechanisms between adipogenesis and the intracellular energy level. PMID:22693256
Kim, Eun Young; Kim, Won Kon; Kang, Hyo Jin; Kim, Jeong-Hoon; Chung, Sang J; Seo, Yeon Soo; Park, Sung Goo; Lee, Sang Chul; Bae, Kwang-Hee
Chaperone-mediated acetylation of histones by Rtt109 identified by quantitative proteomics
Rtt109 is a fungal-specific histone acetyltransferase (HAT) that associates with either Vps75 or Asf1 to acetylate histone H3. Recent biochemical and structural studies suggest that site-specific acetylation of H3 by Rtt109 is dictated by the binding chaperone where Rtt109-Asf1 acetylates K56, while Rtt109-Vps75 acetylates K9 and K27. To gain further insights into the roles of Vps75 and Asf1 in directing site-specific acetylation of H3, we used quantitative proteomics to profile the global and site-specific changes in H3 and H4 during in vitro acetylation assays with Rtt109 and its chaperones. Our analyses showed that Rtt109-Vps75 preferentially acetylates H3 K9 and K23, the former residue being the major acetylation site. At high enzyme to substrate ratio, Rtt109 also acetylated K14, K18, K27 and to a lower extent K56 of histone H3. Importantly, this study revealed that in contrast to Rtt109-Vps75, Rtt109-Asf1 displayed a far greater site-specificity, with K56 being the primary site of acetylation. For the first time, we also report the acetylation of histone H4 K12 by Rtt109-Vps75, whereas Rtt109-Asf1 showed no detectable activity toward H4.
Abshiru, Nebiyu; Ippersiel, Kevin; Tang, Yong; Yuan, Hua; Marmorstein, Ronen; Verreault, Alain; Thibault, Pierre
Characterization of an acetyl esterase from Myceliophthora thermophila C1 able to deacetylate xanthan.
Screening of eight carbohydrate acetyl esterases for their activity towards xanthan resulted in the recognition of one active esterase. AXE3, a CAZy family CE1 acetyl xylan esterase originating from Myceliophthora thermophila C1, removed 31% of all acetyl groups present in xanthan after a 48h incubation. AXE3 activity towards xanthan was only observed when xanthan molecules were in the disordered conformation. Optimal performance towards xanthan was observed at 53�C in the complete absence of salt, a condition favouring the disordered conformation. AXE3-deacetylated xanthan was hydrolyzed using cellulases and analyzed for its repeating units using UPLC-HILIC-ELSD/ESI-MS. This showed that AXE3 specifically removes the acetyl groups positioned on the inner mannose and that acetyl groups positioned on the outer mannose are not removed at all. After a prolonged incubation at optimal conditions, 57% of all acetyl groups, representing 70% of all acetyl groups on the inner mannose units, were hydrolyzed. PMID:25037346
Kool, Marijn M; Schols, Henk A; Wagenknecht, Martin; Hinz, Sandra W A; Moerschbacher, Bruno M; Gruppen, Harry
Facile microencapsulation of curcumin in acetylated starch microparticles.
Abstract Highly acetylated starch was prepared by reacting corn starch with acetic anhydride. Acetylated starch (AS) is soluble in acetone and can be precipitated out from AS-acetone solution when deionised (DI) water is added as a non-solvent. The curcumin dissolved in AS-acetone solution could be effectively encapsulated in AS as spherical microparticles (1-3??m) when mixed with the DI water. The encapsulation yield of curcumin could reach 96.3%. AS encapsulation not only resulted in five-fold lower UV degradation rate of curcumin, the free radical scavenging activity of curcumin still could be well maintained. Approximately, 40% of the 1,1-diphenyl-2-picryl-hydrazyl free radical (DPPH�) was consumed by curcumin encapsulated in AS microparticles within 2?min. PMID:24697176
Nata, Iryanti Fatyasari; Chen, Kuan-Jung; Lee, Cheng-Kang
An acetylated galactoglucomannan from Picea abies L. Karst.
A water-soluble galactoglucomannan composed of D-galactose, D-glucose, and D-mannose in 1:3:17 mole proportion has been isolated from the secondary cell walls of Picea abies L. Karst. About 33% of the polysaccharide units were substituted by acetyl groups. Structural studies of the polymer indicated a beta-(1-->4)-linked glucomannopyranosyl backbone with a low content of branch points at O-6 of mannosyl and glucosyl residues. A preference for mannosyl groups indicates the presence of a single D-galactosyl unit side-chain. About half of the mannose residues were O-acetylated at C-2 and C-3 in 1.7:1 mole proportion. PMID:12039544
Capek, Peter; Alf�ldi, Juraj; Liskov�, Desana
Structures of aminoacylase 3 in complex with acetylated substrates
Trichloroethylene (TCE) is one of the most widespread environmental contaminants, which is metabolized to N-acetyl-S-1,2-dichlorovinyl-l-cysteine (NA-DCVC) before being excreted in the urine. Alternatively, NA-DCVC can be deacetylated by aminoacylase 3 (AA3), an enzyme that is highly expressed in the kidney, liver, and brain. NA-DCVC deacetylation initiates the transformation into toxic products that ultimately causes acute renal failure. AA3 inhibition is therefore a target of interest to prevent TCE induced nephrotoxicity. Here we report the crystal structure of recombinant mouse AA3 (mAA3) in the presence of its acetate byproduct and two substrates: N?-acetyl-l-tyrosine and NA-DCVC. These structures, in conjunction with biochemical data, indicated that AA3 mediates substrate specificity through van der Waals interactions providing a dynamic interaction interface, which facilitates a diverse range of substrates.
Hsieh, Jennifer M.; Tsirulnikov, Kirill; Sawaya, Michael R.; Magilnick, Nathaniel; Abuladze, Natalia; Kurtz, Ira; Abramson, Jeff; Pushkin, Alexander
Hydrolysis of the amorphous cellulose in cotton-based paper.
Hydrolysis of cellulose in Whatman no. 42 cotton-based paper was studied using gel permeation chromatography (GPC), electrospray ionization-mass spectrometry (ESI-MS), and uniaxial tensile testing to understand the course and kinetics of the reaction. GPC results suggested that scission reactions passed through three stages. Additionally, the evolution of soluble oligomers in the ESI-MS data and the steady course of strength loss showed that the hydrolysis reaction occurred at a constant rate. These findings are explained with a more detailed description of the cellulose hydrolysis, which includes multiple chain scissions on amorphous segments. The breaks occur with increasing frequency near the ends of amorphous segments, where chains protrude from crystalline domains. Oligomers unattached to crystalline domains are eventually created. Late-stage reactions near the ends of amorphous segments produce a kinetic behavior that falsely suggests that hydrolysis had ceased. Monte Carlo simulations of cellulose degradation corroborated the experimental findings. PMID:18324778
Stephens, Catherine H; Whitmore, Paul M; Morris, Hannah R; Bier, Mark E
Hydrolysis of whey lactose using CTAB-permeabilized yeast cells.
Disposal of lactose in whey and whey permeates is one of the most significant problems with regard to economics and environmental impact faced by the dairy industries. The enzymatic hydrolysis of whey lactose to glucose and galactose by beta-galactosidase constitutes the basis of the most biotechnological processes currently developed to exploit the sugar content of whey. Keeping this in view, lactose hydrolysis in whey was performed using CTAB permeabilized Kluyveromyces marxianus cells. Permeabilization of K. marxianus cells in relation to beta-galactosidase activity was carried out using cetyltrimethyl ammonium bromide (CTAB) to avoid the problem of enzyme extraction. Different process parameters (biomass load, pH, temperature, and incubation time) were optimized to enhance the lactose hydrolysis in whey. Maximum hydrolysis (90.5%) of whey lactose was observed with 200 mg DW yeast biomass after 90 min of incubation period at optimum pH of 6.5 and temperature of 40 degrees C. PMID:18431601
Kaur, Gurpreet; Panesar, Parmjit S; Bera, Manav B; Kumar, Harish
Hydrolysis of Al3+ from constrained molecular dynamics
NASA Astrophysics Data System (ADS)
We investigated the hydrolysis reactions of Al3+ in AlCl3 aqueous solution using the constrained molecular dynamics based on the Car-Parrinello molecular-dynamics method. By employing the proton-aluminum coordination number as a reaction coordinate in the constrained molecular dynamics the deprotonation as well as dehydration processes are successfully realized. From our free-energy difference of ?G0~=8.0 kcal mol-1 the hydrolysis constant pKa1 is roughly estimated as 5.8, comparable to the literature value of 5.07. We show that the free-energy difference for the hydrolysis of Al3+ in acidic conditions is at least 4 kcal mol-1 higher than that in neutral condition, indicating that the hydrolysis reaction is inhibited by the presence of excess protons located around the hydrated ion, in agreement with the change of the predominant species by pH.
Ikeda, Takashi; Hirata, Masaru; Kimura, Takaumi
Mechanisms of lactone hydrolysis in neutral and alkaline conditions.
The neutral and base-catalyzed hydrolysis of nine carboxylic acid esters was studied using a hybrid supermolecule-PCM approach including six explicit water molecules. The molecules studied included two linear esters, four ?-lactones, two ?-lactones, and one ?-lactone: ethyl acetate and methyl formate, ?-propiolactone, ?-butyrolactone, ?-isovalerolactone, diketene (4-methyleneoxetan-2-one), ?-butyrolactone, 2(5H)-furanone, and ?-valerolactone. DFT and ab initio methods were used to analyze the features of the various possible hydrolysis mechanisms. For all compounds, reasonable to very good qualitative and quantitative agreement with experimental work was found, and evidence is provided to support long-standing hypotheses regarding the role of solvent molecule as a base catalyst. In addition, novel evidence is presented for the existence of an elimination-addition mechanism in the basic hydrolysis of diketene. A parallel work addresses the acid-catalyzed hydrolysis of lactones. PMID:23758295
G�mez-Bombarelli, Rafael; Calle, Emilio; Casado, Julio
Hydrolysis of Cellulose by Purified Cellulase Components: Synergistic Effects.
National Technical Information Service (NTIS)
The hydrolysis of cellulose by purified cellulase components is reported. The adsorption of purified cellobiohydrolases (CBH I and II) and endoglucanases (EG I and II) from Trichoderma reesei strain L27 to microcrystalline cellulose (Avicel) has been stud...
J. Woodward N. E. Lee
Enzymatic hydrolysis of steryl glycosides for their analysis in foods.
Steryl glycosides (SG) contribute significantly to the total intake of phytosterols. The standard analytical procedure involving acid hydrolysis fails to reflect the correct sterol profile of SG due to isomerization of some of the labile sterols. Therefore, various glycosylases were evaluated for their ability to hydrolyse SG under milder conditions. Using a pure SG mixture in aqueous solution, the highest glycolytic activity, as demonstrated by the decrease in SG and increase in free sterols was achieved using inulinase preparations (decrease of >95%). High glycolytic activity was also demonstrated using hemicellulase (63%). The applicability of enzymatic hydrolysis using inulinase preparations was further verified on SG extracted from foods. For example in potato peel ?(5)-avenasteryl glucoside, a labile SG, was well preserved and contributed 26.9% of the total SG. Therefore, enzymatic hydrolysis is suitable for replacing acid hydrolysis of SG in food lipid extracts to accurately determine the sterol profile of SG. PMID:24912717
M�nger, Linda H; Nystr�m, Laura
Sucrose Hydrolysis-Temperature Dependence of the Activation Energy.
National Technical Information Service (NTIS)
Sucrose hydrolysis is often cited as one of the classic examples of a chemical reaction with a temperature-dependent aactivation enery. Two different investigators, Moelwyn-Hughes (polarimetry) and Leininger and Kilpatrick (dilatometry), both claimed that...
J. R. Ward
Enzymatic characterizations and activity regulations of N-acetyl-?-D-glucosaminidase from the spermary of Nile tilapia (Oreochromis niloticus).
N-Acetyl-?-D-glucosaminidase (NAGase) is proved to be correlated with reproduction of male animals. In this study, enzymatic characterizations of NAGase from spermary of Nile tilapia (Oreochromis niloticus) were investigated in order to further study its reproductive function in fish. Tilapia NAGase was purified to be PAGE homogeneous by the following techniques: (NH4)2SO4 fractionation (40-55%), DEAE-cellulose (DE-32) ion exchange chromatography, Sephadex G-200 gel filtration and DEAE-Sephadex (A-50). The specific activity of the purified enzyme was 4100 U/mg. The enzyme molecular weight was estimated as 118.0 kD. Kinetic studies showed that the hydrolysis of p-nitrophenyl-N-acetyl-?-D-glucosaminide (pNP-NAG) by the enzyme followed Michaelis-Menten kinetics. The Michaelis-Menten constant (Km) and maximum velocity (Vm) were determined to be 0.67 mM and 23.26 ?M/min, respectively. The optimum pH and optimum temperature of the enzyme for hydrolysis of pNP-NAG was to be at pH 5.7 and 55�C, respectively. The enzyme was stable in a pH range from 3.3 to 8.1 at 37�C, and inactive at temperature above 45�C. The enzyme activity was regulated by the following ions in decreasing order: Hg(2+) > Zn(2+) > Cu(2+) > Pb(2+) > Mn(2+). The IC50 of Cu(2+), Zn(2+) and Hg(2+) was 1.23, 0.28, and 0.0027 mM, respectively. However, the ions Li(+), Na(+), K(+), Mg(2+) and Ca(2+) had almost no influence on enzyme activity. In conclusion, the enzymatic characterizations of NAGase from tilapia were special to the other animals, which were correlated with its living habit; besides, CuSO4 and ZnSO4 should used very carefully as insecticides in tilapia cultivation since they both had strong regulations on the enzyme. PMID:24012383
Zhang, Wei-Ni; Bai, Ding-Ping; Huang, Yi-Fan; Hu, Chong-Wei; Chen, Qing-Xi; Huang, Xiao-Hong
Variability in Starch Acetylation Efficiency from Commercial Waxy Corn Hybrids
Cereal Chem. 80(1):68-71 Raw material variability is common for starch processors and is responsible for increased processing costs. In this study, variability of starch acetylation due to hybrid influence was quantified. Six waxy corn (maize) hybrids from 1998 and five waxy corn hybrids from 1999 were wet-milled in the laboratory. Starch obtained from each hybrid was modified according to a
M. R. Wilkins; P. Wang; L. Xu; Y. Niu; M. E. Tumbleson; K. D. Rausch
Role of Polyamines in the Regulation of Chromatin Acetylation
Changes in chromatin structure can affect gene transcription, cell proliferation, and differentiation (1). The structural remodeling of chromatin associated with gene expression is mediated in part by the coordinated targeting\\u000a of various chromatin modifying enzymes to gene regulatory regions via recruitment by transcription factors and accessory proteins\\u000a (2). The dynamic interplay between various classes of enzymes that acetylate\\/deacetylate, phosphorylate\\/dephosphorylate, and
Cheryl A. Hobbs; Susan K. Gilmour
An acetylated galactoglucomannan from Picea abies L. Karst
A water-soluble galactoglucomannan composed of d-galactose, d-glucose, and d-mannose in 1:3:17 mole proportion has been isolated from the secondary cell walls of Picea abies L. Karst. About 33% of the polysaccharide units were substituted by acetyl groups. Structural studies of the polymer indicated a ?-(1?4)-linked glucomannopyranosyl backbone with a low content of branch points at O-6 of mannosyl and glucosyl
Peter Capek; Juraj Alf�ldi; Desana Li�kov�
Deciphering the Transcriptional Histone Acetylation Code for a Human Gene
We report the results of experiments designed to test the histone code hypothesis. We found that only a small subset of lysines in histones H4 and H3 are acetylated in vivo by the GCN5 acetyltransferase during activation of the IFN-? gene. Reconstitution of recombinant nucleosomes bearing mutations in these lysine residues revealed the cascade of gene activation via a point-by-point
Theodora Agalioti; Guoying Chen; Dimitris Thanos
Inhibition of acetyl cholinesterase by solanaceous glycoalkaloids and alkaloids
Seven solanaceous glycoalkaloids (?-chaconine, ?2-chaconine, ?-solanine, dehydrocommersonine, commersonine, demissine and tomatine) and three alkaloids (solanidine, tomatidine\\u000a and demissidine) were tested for their ability to inhibit acetyl cholinesterase in anin vitro system. Glycoalkaloids at concentrations of 33�41 parts per million (ppm) gave cholinesterase inhibition ranging from 4.2\\u000a to 26.8%. All three alkaloids had lower anticholinesterase (4.2 to 15.4%) than the seven
Rodney J. Bushway; Sharon A. Savage; Bruce S. Ferguson
Traceless Staudinger acetylation of azides in aqueous buffers.
In this paper, we demonstrate the applicability of water-soluble p-dimethylaminoethyl substituted phosphinomethanethiol in acetyl transfer reactions by the traceless Staudinger ligation with unprotected ?-azido lysine containing peptides in aqueous buffer systems. Additionally, we present an improved synthesis pathway for the water-soluble phosphinothiol linkers requiring less steps in a comparable overall yield in comparison to previously published protocols. PMID:23545137
Sowa, Sylwia; M�hlberg, Michaela; Pietrusiewicz, K Michal; Hackenberger, Christian P R
?-Tricalcium phosphate hydrolysis to octacalcium phosphate: effect of sodium polyacrylate
?-Tricalcium phosphate (?-TCP) hydrolysis into octacalcium phosphate (OCP) has been investigated in phosphoric acid solution at different concentrations of sodium polyacrylate (NaPA). The hydrolysis process has been followed by powder X-ray diffraction, infrared absorption and scanning electron microscopy analyses. In the absence of the polyelectrolyte, ?-TCP undergoes a complete transformation into OCP in 24h. The presence of polyacrylate in solution
A. Bigi; E. Boanini; R. Botter; S. Panzavolta; K. Rubini
The Use of Lewis Acids During Rapid Steam Hydrolysis
Mixed hardwood chips were treated with various concentrations of aluminum chloride hexahydrate, aluminum sulfate hydrate, and ferric chloride and were subjected to rapid steam hydrolysis pretreatment (RASH). The three levels included 0.01, 0.03, and 0.05 moles of catalyst per 1000 grams of wood. Rapid steam hydrolysis (RASH) was done from 180� to 260�C at 20�C intervals for one minute. The
Jagdish Rughani; Louis Wasson; Gary McGinnis
Dilute-acid hydrolysis of sugarcane bagasse at varying conditions
Sugarcane bagasse, a byproduct of the cane sugar industry, is an abundant source of hemicellulose that could be hydrolyzed\\u000a to yield a fermentation feedstock for the production of fuel ethanol and chemicals. The effects of sulfuric acid concentration,\\u000a temperature, time, and dry matter concentration on hemicellulose hydrolysis were studied with a 20-L batch hydrolysis reactor\\u000a using a statistical experimental design.
Markus Neureiter; Herbert Danner; Christiane Thomasser; Bamusi Saidi; Rudolf Braun
Mild Nonoxidative Procedure for the Hydrolysis of Dimethylhydrazones
The past decade has seen amsiderable utilization of dkthylhyldrazones in organic synthesis. Their use as carbonyl equivalents depends on the ability of the practitioner to achieve hydrolylsis under mild conditions. Raagenta previously reported to acanplish this end include acid hydrolysis, methylation and hydrolysis, O3 , CH3C03H, NaI04 , Cu(OAc)2 , CoF3 , NOBF4 , MoF6 , MoOCl3 , UF6 ,
Robert E. Gawley; Enrico J. Termine
On the Br�nsted acid-catalyzed homogeneous hydrolysis of furans.
Furan affairs: Electronic structure calculations of the homogeneous Br�nsted acid-catalyzed hydrolysis of 2,5-dimethylfuran show that proton transfer to the ?-position is rate-limiting and provides support that the hydrolysis follows general acid catalysis. By means of projected Fukui indices, we show this to be the case for unsubstituted, 2-, and 2,5-substituted furans with electron-donating groups. PMID:24006230
Nikbin, Nima; Caratzoulas, Stavros; Vlachos, Dionisios G
Enzymatic hydrolysis of esterified diarrhetic shellfish poisoning toxins and pectenotoxins
Okadaic acid (OA) and dinophysistoxins-1 and -2 (DTX1, DTX2), the toxins responsible for incidents of diarrhetic shellfish\\u000a poisoning (DSP), can occur as complex mixtures of ester derivatives in both plankton and shellfish. Alkaline hydrolysis is\\u000a usually employed to release parent OA\\/DTX toxins, and analyses are conducted before and after hydrolysis to determine the\\u000a concentrations of nonesterified and esterified toxins. Recent
Erin Doucet; Neil N. Ross; Michael A. Quilliam
Acetyl and butyryl cholinesterase inhibitory sesquiterpene lactones from Amberboa ramosa
Background Alzheimer�s disease (AD) is characterized by a progressive memory loss that leads to a profound emotional disturbance in later stages. As no safe and effective drug is yet available for the treatment of AD, secondary metabolites from plants may be instrumental in meeting this challenge. Keeping in view this point we evaluated sesquiterpenes of medicinal plant Amberboa ramosa for their cholinesterase inhibitory activity. Results Four sesquiterpene lactones have been isolated from the ethyl acetate soluble fraction of Amberboa ramosa. In which one compound Amberbin C (1) was found to be new while other three Amberin (2), Amberbin A (3), and Amberbin B (4) were previously reported ones. The structures of the isolated compounds were elucidated using different spectroscopic techniques. Isolated compounds were tested for their inhibitory potential against acetyl cholinesterase and butyryl cholinesterase enzymes. All compounds showed excellent inhibitory activities against acetyl cholinesterase and butyryl cholinesterase. Conclusions A new sesquiterpene lactone has been isolated and fully characterized, the sesquiterpene lactones from Amberboa ramosa showed good inhibitory activities against acetyl cholinesterase and butyryl cholinesterase enzymes, this study indicated that sesquiterpene lactone can become interesting lead molecules in drug development against Alzheimer�s disease (AD).