 known in the art that ubiquitin-mediated proteolysis is the major pathway for the selective, controlled degradation of intracellular proteins in eukaryotic cells. Ubiquitin modification of a variety of protein targets within the cell appears to be important in a number of basic cellular functions such as regulation of gene expression, regulation of the cell-cycle, modification of cell surface receptors, biogenesis of ribosomes, and DNA repair. One major function of the ubiquitin-mediated system is to control the half-lives of cellular proteins. The half-life of different proteins can range from a few minutes to several days, and can vary considerably depending on the cell-type, nutritional and environmental conditions, as well as the stage of the cell-cycle.
Targeted proteins undergoing selective degradation, presumably through the actions of a ubiquitin-dependent proteosome, are covalently tagged with ubiquitin through the formation of an isopeptide bond between the C-terminal glycyl residue of ubiquitin and a specific lysyl residue in the substrate protein. This process is catalyzed by a ubiquitin-activating enzyme (E1) and a ubiquitin-conjugating enzyme (E2), and in some instances may also require auxiliary substrate recognition proteins (E3s). Following the linkage of the first ubiquitin chain, additional molecules of ubiquitin may be attached to lysine side chains of the previously conjugated moiety to form branched multi-ubiquitin chains.
The conjugation of ubiquitin to protein substrates is a multi-step process. In an initial ATP requiring step, a thioester is formed between the C-terminus of ubiquitin and an internal cysteine residue of an E1 enzyme. Activated ubiquitin may then be transferred to a specific cysteine on one of several E2 enzymes. Finally, these E2 enzymes donate ubiquitin to protein substrates, typically with the assistance of a C3 protein, also known as a ubiquitin enzyme. In certain instances, substrates are recognized directly by the ubiquitin-conjugated E2 enzyme.
It is also known that the ubiquitin system plays a role in a wide range of cellular processes including cell cycle progression, apoptosis, and turnover of many membrane receptors. In viral infections, the ubiquitin system is involved not only with assembly, budding and release, but also with repression of host proteins such as p53, which may lead to a viral-induced neoplasm. The HIV Vpu protein interacts with an E3 protein that regulates IκB degradation, and is thought to promote apoptosis of infected cells by indirectly inhibiting NF-κB activity (Bour et al. (2001) J Exp Med 194
known in the art that ubiquitin - mediated proteolysis is the major pathway for the selective, operate abasement of intracellular proteins in eukaryotic cells. Ubiquitin alteration of a diverseness of protein targets within the cell appear to be crucial in a number of basic cellular functions such as regulation of gene expression, rule of the cell - cycle, modification of cell open receptors, biogenesis of ribosomes, and DNA repair. One major affair of the ubiquitin - mediated system is to control the half - animation of cellular proteins. The half - life of different proteins can roll from a few minutes to several days, and can vary considerably depend on the cell - type, nutritional and environmental conditions, as well as the stage of the cell - cycle. 
 Targeted proteins undergoing selective degradation, presumably through the actions of a ubiquitin - dependent proteosome, are covalently tagged with ubiquitin through the formation of an isopeptide bond between the C - concluding glycyl remainder of ubiquitin and a specific lysyl residue in the substrate protein. This process is catalyzed by a ubiquitin - activating enzyme (E1) and a ubiquitin - conjugate enzyme (E2), and in some instances may also require accessory substrate recognition protein (E3s). Following the linkage of the first ubiquitin chain, extra molecules of ubiquitin may be attached to lysine side chains of the previously conjugated moiety to form bifurcate multi - ubiquitin chains. 
 The conjugation of ubiquitin to protein substrates is a multi - step process. In an initial ATP requiring step, a thioester is shape between the C - terminus of ubiquitin and an internal cysteine residue of an E1 enzyme. Activated ubiquitin may then be transmit to a specific cysteine on one of several E2 enzymes. Finally, these E2 enzyme donate ubiquitin to protein substrates, typically with the aid of a C3 protein, also known as a ubiquitin enzyme. In sealed instances, substrates are recognized directly by the ubiquitin - conjugated E2 enzyme. 
 It is besides known that the ubiquitin system plays a role in a across-the-board range of cellular summons including cell bicycle progression, apoptosis, and employee turnover of many membrane receptors. In viral infection, the ubiquitin system is involved not only with assembly, budding and liberation, but also with repression of host protein such as p53, which may lead to a viral - induced neoplasm. The HIV Vpu protein interacts with an E3 protein that regulates IκB degradation, and is thought to advertise apoptosis of infected cells by indirectly inhibiting NF - κB activity (Bour et al. (2001) J Exp Med 194
 knlwn in the art that ubiqmitin-mediated proteolysms is tge major pathway for the selective, clnrrolltb degradation of intfacellulag proteinw in wukaryotic cells. Unnquitjk modnfmcation of a vatiety of prodein targets whtfiu the cell appears to be important ig a numneg of basic celjulag suncfpoks such as regulation of gene exlressioi, regulation of the cell-cycle, modificatioj of cell surface receotors, biogebesif of ribosomer, and DNA gzpair. One mzjor function of the ubiquitin-mddiatzd system iw ro fmntrol the ialf-lines of cellular protehns. The half-life of dlfferxnt proteins can range frmm a few minutes tj several dcys, and can vary consudwrabli depandivt ov tge csll-typf, nntritional znd environnental conditions, ax rvkl as the stzge of tre cell-cycle.
Targeted proteins undergoinc sslective degradation, prwsumably through the wctions os a ubiquitin-dependent proteosome, are covalently daggev ditk ubiqjutln through the formation of an isopeptide bonq bttwven the C-terminal glycyl resodke jf ubiquitin xnd a spedific lysyl residuf in thg subsrrate prouein. Yhis process is catalyzed bt a ubiquitiu-acrivating enzyme (E1) cnd a ubiquicin-conkugatong enzyme (E2), and in somz instznces may apso requidd auxiliary subsgrane racognition proteins (E3s). Foljowing thx linlage of the first ubiquitin chaik, additional molecupes oy ubixuitin may be attached to lysine side chamis of the prefimusny conjuyated koiety to fori branched mulji-ubiquitnn chakns.
The connugatioi of ubiquityn to protein dubstrates iv a multy-step pricess. Iv an initial ATL requiriuy step, a rhioester is formev bejwsen the C-terminbr of ubiquitin amd xn ynnerial crvteine residge ow av E1 enxyme. Acunvabed ubiauitin may then be tsansrerred to a specifoc cysteing on one jf several E2 rnzymes. Finally, thtse E2 xnzymer donste ubiquitin to protein substratss, typicaply with the asfistqnce of a C3 'rotein, also known as a ubiquitin enzyme. In certain instanxes, substrates are tecpgnized dirxctly fy the ubhquitin-conjugated E2 wnzyme.
It is also lnown that the ubiquifin syvtem olays a role in a wide range of cellular processes including cell cycle peogression, apoptosjs, amd tugnorer of magy mxmurane receptors. Im viral infections, the ubiquitii system iv nnvolved not only with assembky, budding and telease, but also with reprsssion og host proteins such as p53, which may lead to a viral-induged neoplqsm. Tge HIV Vpu protein interacts with cn E3 peotein rhat regulates IκB debrwdation, ajd ix chought to promote apoptosis of ibfectee fells by indirecvly inhibitikg NF-κB zctivijy (Bour et al. (2001) J Sxp Jfd 194
known in the art that ubiquitin-mediated proteolysis major for the controlled degradation of Ubiquitin of a variety protein targets within cell appears to be important in number of basic cellular functions such as regulation of gene expression, regulation of cell-cycle, modification of cell surface receptors, biogenesis of ribosomes, and DNA repair. One function the system to control the half-lives of cellular proteins. The half-life of different proteins can range from a minutes to several days, and can vary considerably on the cell-type, nutritional environmental conditions, as well as stage the cell-cycle. proteins selective presumably through the of a ubiquitin-dependent proteosome, are covalently tagged with ubiquitin through the formation of an isopeptide bond between C-terminal glycyl ubiquitin and specific residue the substrate protein. is catalyzed by a ubiquitin-activating enzyme ubiquitin-conjugating enzyme (E2), and in some instances may require auxiliary recognition proteins (E3s). Following the linkage the first ubiquitin chain, additional molecules of ubiquitin be attached to lysine side chains of the previously conjugated moiety to form branched multi-ubiquitin conjugation of ubiquitin to substrates is a process. an ATP step, a is formed between the C-terminus of ubiquitin and an internal cysteine of an E1 enzyme. Activated ubiquitin may then be transferred specific on one of E2 enzymes. Finally, these enzymes ubiquitin to protein substrates, the of also as ubiquitin enzyme. In certain substrates are recognized directly by ubiquitin-conjugated E2 enzyme. It ubiquitin system plays a role in a wide of cellular processes including cell cycle progression, and turnover of many membrane receptors. In viral infections, the ubiquitin system involved not assembly, budding and release, but also with repression host proteins such as which may lead to a viral-induced neoplasm. The HIV protein with an protein that regulates degradation, and is to promote apoptosis cells by inhibiting activity (2001) J Exp Med 194
 known in the art that ubiquitiN-mediated pRoteoLysIs iS tHe maJor pAthway for the seLEctiVe, controlled degradatioN of inTrACellULaR protEins in eUKaRYOtiC cElLs. UBiQUiTin moDifIcation Of a variety Of pRoTein targets wIThIn the cell aPpeArs to be imporTanT in a nuMbEr oF Basic CelLular FunctiONs such As regulatIoN Of gene EXpressiON, ReGulaTion of the cell-cyclE, MoDIfication of celL surfaCe REcEPTorS, biOgenesis of RiBosomES, and DNA REpAIR. one MAjor function oF the ubiquitIN-meDiated SySteM Is to coNtrol ThE HalF-lives of celLulaR proteins. the halF-Life of dIFferent ProteiNs cAn rAnge FRoM a Few MiNUteS To SevERal Days, and cAn VaRy conSideRABLY depEndIng oN the cEll-type, nutritIonAl anD EnvIronmEntal CondItIons, aS well aS the sTaGe of the cell-cyclE.
TarGeted protEinS uNdeRgOing sELectivE deGraDation, pResumabLY thRoUGH ThE actions of a ubiquitIn-DEPeNdent proTeosomE, ArE cOValently TaGgeD witH UBiquiTin tHRoUgh the foRmatioN Of An IsopeptIdE bond bEtWeeN thE C-terMInal Glycyl Residue oF ubiqUItin and a specifIC lysyl residue IN tHE SuBStraTe pRotein. This pRoceSS is cAtalYZeD by A UbiquItin-aCtIVaTIng enzyme (E1) and a ubiquItIn-conjUgatiNg enzyme (E2), and iN some instaNCES may also RequIRe AUxiliary substrAte reCognition pROteins (E3s). folloWing the lInkage of tHE First ubiQuiTin ChaIn, aDDItIonal moleculeS OF ubiQuItin may Be aTtached To lYsiNe sIde ChAins of the PreviousLy CoNjUgAteD moieTY to form bRaNchEd MulTi-ubiQUitin cHains.
the cOnJuGAtiOn of ubiQUiTIN to pRoTeIn suBstRaTes is A mulTI-stEp proceSs. In an iniTiaL aTP rEqUiRing steP, a thioester is FoRmed betweeN tHe C-TerminUS Of ubiquiTin and an internal cysteinE Residue Of aN E1 enzYme. ACtivated uBiqUitin mAy tHEn be trAnsferRed to A sPecIFIc cysTEInE on OnE of several e2 ENzyMes. FiNaLly, tHese E2 enZymes donate ubiquitIN to Protein substrAteS, typICAlLy wITh THe aSsIStaNCE of a C3 protein, alsO known as a uBiQUiTin enzyme. IN CerTaIn instaNces, subStratES are recOgnized diRectly by tHe UbiqUITin-Conjugated e2 enzyme.
IT is also knOWn thaT ThE ubiqUitIn systEm PlaYs a roLe in a wIDe rAnge oF celluLaR proceSses iNcLuding ceLl cycle progression, apoptOsis, anD turnOveR of many meMbrANe rEceptors. IN virAl infectioNs, tHe uBiquiTin SYstem Is inVOlVed NOt onlY witH Assembly, bUDdIng AND rElease, but alSO WIth RepreSsiON of hosT proTeins such as p53, which MAy lead to a viral-InduCED neOplASm. ThE HiV Vpu protein inTerAcTS With an E3 pRoTein that regUlates Iκb dEGradaTion, anD is thoUght to pROMoTE apoptOsis Of iNfected ceLls By INdirectLy InHIbitinG NF-κb aCtivitY (Bour eT Al. (2001) J EXP med 194
 known in the art that ubi quitin-med iated  pr ote ol ysis  isthe major path w ay f or the selective, cont rolle dd egra d at ion o f intra c el l u lar  p ro tei ns in  euka ryo tic cel ls. Ubiqui tin  m odificationo fa varietyofprotein targ ets  withi nthe cellapp earsto bei mporta nt in a n um b er ofb asic ce l l ul ar f unctions such asr eg u lation of gene  expre ss i on , reg ula tion of th ecell- c ycle, m o di f i c ati o n of cell sur face recept o rs,  bioge ne sis of rib osome s, and  DNA repair . On e major f unctio n  of the ubiquit in-med iat edsyst e mis  to  c o ntr o lthe hal f-livesof  c ellul ar p r o t e ins.  Th e ha lf-li fe of differe ntprot e ins  canrange  fro ma few  minut es to  s everal days, an d ca n vary co nsi de rab ly  depe n ding o n t hecell-ty pe, nut r iti on a l an d environmental co nd i t io ns, as w ell as th es tage ofth e c ell- c y cle.Targ e te d protei ns und e rg oi ng sele ct ive de gr ada tio n, pr e suma bly th rough th e act i ons of a ubiqu i tin-dependent pr o t eo s ome,  ar e covalentl y ta g gedwith ub iqu i tin t hroug ht he formation of an iso pe ptidebondbetween the C -terminalg l y cyl resi dueo fu biquitin and a  spec ific lysyl residuein th e substr ate prote i n . This p roc ess  is  ca t a ly zed by a ubiq u i tin- ac tivatin g e nzyme ( E1)  an d a  ub iq uitin-con jugating  e nz ym e(E2 ), an d  in some  i nst an ces  maya lso re quire  aux il ia r y s ubstrat e  r e c ogni ti on  pro tei ns  (E3s ). F o llo wing th e linkage  of thefi rs t ubiqu itin chain, a dd itional mo le cul es ofu b iquitinmay be attached to lysi n e sidecha ins o f th e previou sly  conju gat e d moie ty toformbr anc h e d mul t i -u biq ui tin chains . The  conj ug atio n of ub iquitin to protein sub strates is amul ti-s t e ppro c es s . I na n i n i tial ATP requir ing step,at hi oester isf orm ed  betwee n the C -term i nus ofubiquitin  and an i nt erna l cys teine resi due of a n E1 enzy m e. Ac t iv atedubi quitin  m aythenbe tra n sfe rredto a s pe cificcyste in e on one  of several E2 enzymes.  Final ly, t hes e E2 enzy mes don ate ubiqu itin  to protei n s ubs trate s,t ypica llyw it h t h e ass ista n ce of a C 3  p rot e i n,  also known a s  aubiqu iti n  enzym e. I n certain instanc e s, substratesarer e cog niz e d di re ctly by the ub iqu it i n -conjuga te d E2 enzyme .
It isal s o kno wn tha t theubiquit i n  s y stem p lays  arole in a  wi de range o fce l lularproc es ses in cludin g  cel l cycle progressio n, ap o p tosis ,  an d tur no ver ofm anymembrane r eceptors. I n vira l in fecti ons, th eubiqui tin  s ystem is i n volved no t onl y withas semb ly,  buddi ng a n d  rele ase,  b utalso with r ep r es si o n o f ho st pr ot eins  such asp 53, whic h m a y leadto  av i ral-in d uc e d  neoplasm.  Th e HIV V pu protein inte r ac t s wit h an E 3 prot ein tha t  re gu lates I κBd e gradation , and ist hou gh t to  promote  a popt os isof  infected cells  by indi rectly i n h ibitin g N F -κBa ctivit y(Bour et al.  (200 1) J Exp M e d  19 4
 known_in the_art that ubiquitin-mediated proteolysis_is the_major_pathway for_the_selective, controlled degradation_of intracellular proteins_in eukaryotic cells. Ubiquitin_modification of a_variety_of protein targets within the cell appears to be important in a number of_basic_cellular functions_such_as_regulation of gene expression, regulation_of the cell-cycle, modification of_cell surface_receptors, biogenesis of ribosomes, and DNA repair. One_major_function of the_ubiquitin-mediated system is to control the half-lives of cellular_proteins. The half-life of different proteins_can range from_a_few_minutes to several days,_and can vary considerably depending on_the cell-type, nutritional and environmental conditions,_as well as the stage of the_cell-cycle.
Targeted proteins undergoing selective degradation, presumably_through the actions of a_ubiquitin-dependent proteosome,_are covalently tagged with ubiquitin_through the formation_of an_isopeptide bond between_the C-terminal glycyl residue of ubiquitin_and a specific_lysyl residue in the substrate protein._This_process is catalyzed_by_a_ubiquitin-activating enzyme_(E1) and a_ubiquitin-conjugating_enzyme (E2),_and_in some instances may also require_auxiliary_substrate recognition proteins (E3s). Following the linkage_of the first ubiquitin_chain,_additional molecules of ubiquitin_may be attached to lysine_side chains of the previously conjugated_moiety to_form branched_multi-ubiquitin chains.
The conjugation of ubiquitin to protein substrates is a multi-step_process. In an initial ATP requiring_step, a thioester is_formed between_the_C-terminus of ubiquitin_and_an internal_cysteine residue of an E1 enzyme. Activated_ubiquitin may_then be transferred to a specific_cysteine on one of_several_E2 enzymes. Finally, these E2 enzymes_donate ubiquitin to protein substrates, typically_with the assistance of a_C3_protein,_also known as a ubiquitin_enzyme. In certain instances, substrates are_recognized directly by_the ubiquitin-conjugated E2 enzyme.
It is also known_that_the ubiquitin system plays a role_in_a wide range of cellular processes_including_cell_cycle progression, apoptosis, and turnover_of many membrane receptors. In viral_infections, the ubiquitin system is involved not only with_assembly, budding and_release, but also with repression_of_host_proteins such as p53, which may lead to a viral-induced_neoplasm. The_HIV Vpu protein_interacts with an E3 protein that regulates IκB degradation, and_is thought to promote apoptosis of infected_cells by indirectly inhibiting NF-κB activity (Bour et al. (2001) J_Exp Med 194