The present invention features molecules that relate to SYNGAP (Synaptic GTPase Activating Protein), an excitatory synapse protein that has been found to bind synaptic proteins and modulate signal transduction. In one aspect, the invention provides isolated polynucleotides that encode SYNGAP or fragments or derivatives thereof. Further provided are SYNGAP or SYNGAP-related polypeptides encoded by the polynucleotides. In another aspect, the invention provides immunological molocules that are capable of binding the polypetides. Additionally provided are methods for using the molocules of this invention, e.g., to treat or prevent a disorder mediated by SYNGAP. The invention also provides screening assays for detecting compounds useful in the diagnosis or treatment of disorders impacted by SYNGAP.

1. Field of the Invention
The present invention relates to signal transducing synaptic molecules and particularly to mammalian SYNGAP (Synaptic GTPase Activating Protein), including recombinant SYNGAPs and fragments and derivatives thereof. In one aspect, the invention provides molecules for detecting and analyzing SYNGAPs in vitro and in vivo. In another aspect, the invention provides assays for detecting compounds that modulate SYNGAP or SYNGAP-related activities. The invention has a variety of applications including use in screens to detect pharmacological agents useful in the diagnosis or treatment of disorders associated with SYNGAP.
2. Background
Neurons communicate by a variety of means including synaptic transmission. One form of synaptic transmission involves chemical signaling; a process generally involving neurotransmitter release from one neuron and modulation of a post-synaptic receptor in another neuron. The release and modulation is usually manifested by propagation of a chemical or electrical impulse. See Edelman, G. M. et al. (eds.) (1987) in Synaptic Function, New York: Wiley; and Goodman and Gilman (1996) in The Pharmacological Basis of Therapeutics, 9th ed. J. G. Hardman, et al. (eds) Pergamon Press, NY.
In many instances, chemical signaling requires specialized neuronal structures called chemical synapses. Analysis of chemical synapses has attracted substantial interest. For example, chemical synapses have been reported to be involved in many, if not all, nervous system functions including neuronal plasticity. In particular, neuronal plasticity is believed to impact critical functions such as cognition, e.g., memory and learning, as well as certain neurodegenerative disorders. See e.g., Kandel, E. R. et al; (1991) in Principles of Neuroscience, Appleton and Lange, Norwalk, Conn. and references cited therein.
A variety of approaches have been used in attempts to understand chemical synapses. For example, certain molecular and biochemical approaches have suggested that chemical synapses are structured and include molecules such as receptors, cytoskeletal proteins, and signal transduction molecules. See e.g., Ehlers, M. D., et al. (1996) Curr. Opin. Cell Biol. 8: 484; Sheng, M. (1996) Neuron 17:575; and Huganir, R. L. and Greengard, M. (1990) neuron 5: 555.
More particularly, it has been reported that appropriate chemical synapse structure requires presence of a protein termed PSD-95/SAP90. The PSD-95/SAP90 protein is representative of a family of molecules including SAP120. Specific members of the PSD-95/SAP90 family are localized at or near chemical synapses. See e.g., Ehlers, M. D., et al. supra; Sheng, M supra; Lau, L. F., et al. (1996) J. Biol. Chem. 271:21622; Muller, B. M., et al. (1996) Neuron 17:255; and references cited therein.
There has much effort towards understanding PSD-95/SAP90 and related proteins. For example, it has been reported that most members of the protein family exhibit the same or a closely related structure, i.e., three tandem PDZ (PSD-95, DLG, ZO-1) domains, a SH3 (src homology 3) domain, and an inactive yeast guanylate kinase domain (GK). See e.g., Kim, E., et al. (1995) Nature 378:85; Kornau, H. C., et al. (1995) Science 269:1737.
There has been recognition that PDZ domains exist in proteins associated with cell membranes. For example, it has been reported that the PDZ domains of PSD-95 bind to specific subunits of the (N-methyl-D-aspartate) NMDA receptor. It has also been reported that the last three amino acids of the NMDA receptor subunits define a consensus sequence (T/SXV; X is any amino acid). The subunit sequence has been reported to facilitate the binding. See e.g., Kennedy, M. B. (1995) Trends Biochem. Sci. 20; Gomperts, S. N. (1996) Cell 84:659; Sheng, M. supra.
There has also been recognition that chemical synapses can employ signal transduction to modulate pre- and post-synaptic activity. For example, certain members of the PSD-95/SAP90 protein family are believed to be associated with signal transduction. More particularly, the PSD-95 protein has been proposed to interact with certain signal transduction kinases. That interaction has been proposed to be important to synaptic function. See Gomperts, supra; Saras, J., and Heldin, C. H. (1996) Trends Biochem. Sci. 21:455; Huganir, R. L. and Greengard, M. supra.
There has also been recognition that the PSD-95/SAP90 protein may be capable of serving as an adaptor molecule. More specifically, there have been reports that the PSD-95/SAP90 protein may be able to relate synaptic activity and signal transduction in some instances. That property is believed to important to appropriate synapse function. See e.g., Brenman, J. E. et al. (1996) Cell 84: 757; and Saras, J. and Heldin, Carl-Henrik (1996), supra.
A variety of signal transduction molecules are known including specified kinases and proto-oncogenes. For example, the proto-oncogene Ras is recognized as a G-protein that is apparently involved in signal transduction pathways affecting, e.g., cell growth, cell differentiation, neuronal plasticity and cell survival. In particular, Ras appears to have a substantial role in kinase activation. In addition, biological activity manifested by a variety of neurotrophic factors (i.e. neurotrophins) may be derived through Ras-associated signaling pathways. See e.g., Bokoch, G. M., and Der, C. J. (1993) FASEB. J. 7:750; Marshall, C. J. (1996) Cell Biol. 8:197; Finkbeiner, S. and Greenberg, M. E. (1996) Neuron 16:233; Kang, H., and Schuman, E. M. (1996) Science 273:1402.
Certain neuronal functions have been proposed to be affected by Ras-mediated signal transduction. See e.g., Seger, R., and Krebs, E. G. (1995) FASEB J. 9:726; and Finkbeiner and Greenberg, supra.
Additional signal transduction pathways are known. For example, the inositol triphosphate signaling pathway has been reported to couple modulation of certain receptors to a variety of functions, many of which relate to calcium. See e.g., Berridge, M. J. (1988) Pro. R. Soc. Lond. (Biol) 234: 359.
It would be desirable to identify molecules that impact chemical synapse function and particularly interact with the PSD-95/SAP90 protein. It would be further desirable to identify molecules that can bind the PSD-95/SAP90 protein and affect signal transduction. It would also be desirable to have effective assays for identifying compounds and especially pharmaceutical agents with capacity to modulate the function of these molecules.
The present invention features molecules that relate to SYNGAP (Synaptic GTPase Activating Protein); an excitatory synapse protein that has been found to bind synaptic proteins and modulate signal transduction. In one aspect, the invention provides isolated polynucleotides that encode SYNGAP or fragments or derivatives thereof Further provided are SYNGAP or SYNGAP-related polypeptide encoded by the polynucleotides. In another aspect, the invention provides immunological molecules that are capable of binding the polypeptides. Additionally provided are methods for using the molecules of this invention, e.g., to treat or prevent at least one disorder mediated by SYNGAP. The invention also provides screening assays for detecting compounds useful in the diagnosis or treatment of disorders impacted by SYNGAP.
We have discovered mammalian SYNGAP: a novel protein that binds synaptic proteins important for chemical synapse function. Additionally, we have found that SYNGAP is capable of modulating certain signal transduction molecules and particularly the Ras proto-oncogene (Ras). We have particularly found that SYNGAP is positioned to relate synaptic activity and signal transduction, thereby indicating a significant role in many aspects of nervous system function.
As will be discussed below, SYNGAP is represented by a family of alternatively spliced variants including SYNGAP-a, SYNGAP-b, and SYNGAP-c.
The present invention provides a number of significant uses and advantages. For example, the invention relates, for the first time, recognized synaptic proteins such as PSD95/SAP90 and SAP120 to signal transduction. More particularly, the invention provides SYNGAP and SYNGAP-related molecules that are believed to serve as adaptors between key synaptic proteins and specific transduction pathways. Accordingly, the present invention is expected to facilitate attempts to more fully understand relations between synapses and signal transduction, particularly relationships between synaptic function and signaling molecules such as Ras, inositol triphosphate and certain other transduction molecules.
The present invention has a wide spectrum of important applications. For example, specific molecules of this invention can be used as a diagnostic tool to detect excitatory synapses, i.e., those that include mammalian SYNGAP. Illustrative of such excitatory synapses are those impacting neuronal plasticity, particularly habituation, sensitization, leaning and memory; as well as certain neurological disorders. In addition, the invention can be employed in the diagnosis, treatment or prophylaxis of certain neurological disorders impacted by SYNGAP. As will be discussed in more detail below, exemplary neurological conditions include those specifically affecting awareness and cognition as well as neuronal growth and survival.
Further, the present invention provides a variety of highly useful molecular markers including polynucleotides, polypeptides, and immune system molecules that can be used in many commercial, medical, home or research settings. For example, certain molecules of this invention can be used in screening assays to detect compounds and particularly pharmaceutical agents useful in the diagnosis or treatment of neurological disorders impacted by SYNGAP. Of particular interest are recognized manipulations that can be employed for the identification of small molecules, e.g., synthetic peptides, peptide mimetics, drugs, etc., that can modulate interaction between SYNGAP and the synaptic proteins to which it associates; interaction between SYNGAP and signal transduction molecules or both. Related techniques can be used to screen for small molecules that potentially block or enhance one or all of these interactions. In particular, in vitro screens are provided that can detect SYNGAP antagonists or agonists.
Additional uses include use of SYNGAP polynucleotides to detect SYNGAP expression in desired cells or groups of cells such as tissue or an organ by conventional in situ hybridization methods.
Particularly useful are SYNGAP-binding immune system molecules of this invention such as the antibodies and antigen-binding antibody fragments provided below. For example, the antibodies (monoclonal and polyclonal) can be used alone or in combination with other agents to facilitate identification of excitatory chemical synapses that include SYNGAP. In some instances, it may be desirable to include the antibodies wholly or as part of a therapeutic strategy to monitor or in some cases modulate the excitatory chemical synapses. Preferred molecules of the invention are flexible and can be modified, if desired, to deliver a desired molecule such as a drug, toxin, enzyme or radionuclide optionally through a linker sequence such as a peptide linker sequence. As an illustration, a SYNGAP binding antibody of this invention can be detectably-labeled to identify excitatory chemical synapses in vitro or in vivo.
The SYNGAP and SYNGAP-related molecules of this invention have additional uses and advantages in vitro and in vivo. For example, the molecules can be employed in functional, cellular and molecular assays (e.g., screens) and in structural analysis, including X-ray crystallography, nuclear magnetic resonance imaging (NMRI), computational techniques. Of particular interest are those techniques involving computer-assisted simulation of synapses (i.e. formulation of virtual synapses). Also included are useful techniques for using the SYNGAP and SYNGAP-related molecules as markers that can provide diagnostic imaging of excitatory chemical synapses in vivo. By way of illustration, certain molecules of this invention can be employed to visualize specific neurons and particularly chemical synapses including SYNGAP in the brain of a living patient. In this embodiment, the desired molecule will usually be detectably-labeled with a suitable tag such as a radionuclide or other suitable imaging component known in the field.
Specific SYNGAP and SYNGAP-related molecules of this invention can be provided in a kit form or other convenient form to facilitate manufacture, packaging, dissemination, storage, and/or use of the present invention.
Accordingly, in one aspect, the invention provides isolated polynucleotides (RNA, mRNA, cDNA, genomic DNA, or chimeras thereof) that encode SYNGAP or a fragment or derivative of SYNGAP. Illustrative of such polynucleotides include those encoding a mammalian SYNGAP, e.g., a primate and particularly a rat or a human SYNGAP. In one embodiment, the polynucleotide encodes a mammalian SYNGAP having a molecular weight of between about 100 to about 15 kDa or greater. In another embodiment, the polynucleotide has at least about 70 percent sequence identity to any of the nucleotide sequences shown in SEQ ID NOS. 1, 3, or 5. Such sequence similarity (i.e. about 70%) or greater similarity will sometimes be referred to herein as xe2x80x9csubstantial homologyxe2x80x9d or like term. Specifically preferred polynucleotides of the invention encode the rat SYNGAP shown in any of the SEQ ID Nos. 2, 4, or 6.
In another aspect, the present invention provides an isolated polynucleotide that is capable of hybridizing to the nucleotide sequence shown n SEQ ID Nos. 1xe2x89xa72, 4-5, or 7-8 under moderate stringency hybridization conditions. In a preferred embodiment, the polynucleotide will also hybridize to those specific sequences shown in SEQ ID NOS. 1-2, 4-5, or 7-8 under high stringency conditions. The terms xe2x80x9cmoderatexe2x80x9d and xe2x80x9chighxe2x80x9d hybridization stringency have readily understandable meaning to those of skill in this field. Exemplary stringency conditions are disclosed in the discussion and examples which follow. In one embodiment, the polynucleotide capable of hybridizing to the SEQ ID NO: 1 or under high stringency conditions is between from about 12 to about 50 nucleotides in length. Illustrative of such polynucleotides are oligonucleotide primers made by conventional synthetic methods. In another embodiment, the polynucleotide is between about 100 to about 3500 nucleotides in length or greater. Illustrative of such polynucleotides are restriction enzyme fragments or chemically synthesized fragments complementary to the sequences shown in SEQ ID Nos. 1, 3, or 5.
In another embodiment of the invention, the polynucleotide capable of hybridizing to the nucleotide sequence shown in SEQ ID Nos. 1, 3, or 5 under high stringency conditions has a length of between about 100 up to about 4000 nucleotides or greater. In a preferred embodiment, the polynucleotide is a cDNA encoding an amino acid sequence capable of modulating the proto-oncogene Ras as determined, e.g., by a standard Ras GTPase activity assay. In a specific embodiment, the amino acid sequence encoded by the polynucleotide is capable inhibiting the Ras protein by at least about 10% up to about 100% in the standard Ras GTPase assay. Additionally preferred are polynucleotides that encode polypeptides capable of modulating Ras-mediated signaling transduction pathways and optionally signal transduction molecules xe2x80x9cdownstreamxe2x80x9d of Ras (directly or indirectly). Examples of such downstream molecules include recognized signaling and effector molecules. Illustrative methods for identifying such polynucleotides and amino acid sequences are described below.
One or a combination of standard approaches can be used to monitor Ras GTPase activity in cells or cell lysates. Preferably, the cell or cell lysate will include a naturally-occurring or recombinant Ras protein. Preferred assays for measuring and quantitating the Ras GTPase activity are discussed below.
Additionally preferred polynucleotides of this invention encode a mammalian SYNGAP or fragment or derivative thereof that is capable of significantly reducing inositol triphosphate signaling as determined, e.g., by modulation of phospholipase C activity in a standard phospholipase C enzyme assay. Preferably, the polynucleotide is a cDNA that is capable of increasing or decreasing the enzyme activity by at least about 10% or more up to about 100% relative to a suitable control assay. Specific phospholipase C assays are described below.
Additionally preferred polynucleotides of this invention encode a mammalian SYNGAP or a SYNGAP-related amino acid sequence that is capable of binding at least from about 1, 2 or 3 up to about 10 PDZ domains as determined by a standard PDZ domain binding assay. As noted above, the PDZ domain has been reported to be present in a variety of membrane proteins, e.g., the PSD95/SAP90 and SAP 120 proteins, and is believed to significantly impact chemical synapses.
By the term xe2x80x9cSYNGAP-relatedxe2x80x9d nucleotide or amino acid sequence or similar term is meant a fragment or derivative of SYNGAP sequence (polynucleotide or polypeptide as described below.
A variety of methods are known in this field for detecting and quantifying, if desired, PDZ binding. In general, the methods are capable of detecting formation of a binding complex and can be optimized to provide qualitative or quantitative characterization of those binding complexes. Illustrative methods for detecting the PDZ binding are more fully disclosed in the discussion and examples that follow.
Additionally preferred polynucleotides are those molecules that encode an amino acid sequence that includes at least a Ras GTPase Activating Protein (GAP) domain up to about 2 to about 3 GAP domains, and a C-terminal sequence that includes at least the following general amino acid sequence: (T or S), X V; wherein X is an amino acid, preferably one of the 20 natural amino acids.
Further preferred are those polynucleotides that encode an amino acid sequence that includes at least one pleckstrin homology (PH) up to about 3 PH domains and at least one C2 domain up to about 3 C2 domains.
Particularly preferred polynucleotides of this invention encode an amino acid sequence that includes in an N- to C-terminal orientation: at least one PH homology, preferably one PH homology; at least one C2 domain, preferably one C2 domain; at least one GAP domain, preferably one GAP domain; and the C-terminal sequence having the sequence (T or S), X V described above. Additionally preferred is a polynucleotide encoding an amino acid sequence that includes in an N- to C-terminal direction at least about the following amino acids of SEQ ID NO. 6, 4 to 72, 87 to 190, 266 to 502 and 1132 to 1135.
Specifically preferred are those polynucleotides that encode a rat SYNGAP as represented by any one of SEQ ID NOS. 2, 4 or 6 including fragments or derivatives thereof.
One or a combination of different strategies can be used to analyze the SYNGAP or SYNGAP-related molecules disclosed herein, e.g., to detect homologous molecules or to identify protein domains (e.g., GAP, PH, and C2). Specifically included are biochemical, immunological and biosensor-type assays as well as certain well-known computer-assisted manipulations.
Additionally provided by the present invention are fragments or derivatives of the polynucleotides encoding mammalian SYNGAP or SYNGAP-related molecules, as well as recombinant vectors including the polynucleotides or the fragments or derivatives thereof It is generally preferred that the recombinant vector be capable of propagating the isolated polynucleotide in a suitable prokaryotic or eukaryotic host cell. Additionally preferred recombinant vectors are capable of expressing that isolated polynucteotide as RNA and preferably mRNA, in a suitable cell expression system. The recombinant vector can include nearly any number of useful elements, however in most cases the vector will include control elements operably linked to the inserted nucleic acid (e.g., promoter, leader, and/or enhancer elements) which control elements can be selected to optimize replication and/or transcription of the vector in the cells.
As noted, polynucleotides of the invention generally encode mammalian SYNGAP or a fragment or derivative thereof In one embodiment, the polynucleotides are substantially homologous to the SYNGAP sequence shown in SEQ ID NO: 2, 4, or 6. In a specific embodiment, the isolated polynucleotides include or consists of cDNA and have a length of between about 50 to about 100 nucleotides up to about 4000 nucleotides or more, as determined by standard nucleic acid sizing methods. In another embodiment, the isolated nucleic acid includes or consists of RNA and particularly mRNA that is also substantially homologous to the specific rat SYNGAP sequence and which can have substantially the same length as the cDNA.
Also provided are host cells that include a polynucleotide as disclosed herein including those that express SYNGAP, a SYNGAP-related molecule or a fragment or derivative thereof under suitable cell culture conditions. Preferably, the host cells are capable of expressing the desired amino acid sequence in the host cell, cell medium, or both.
The invention also includes methods for isolating a polynucleotide encoding a mammalian SYNGAP or SYNGAP-related molecule such as the rat SYNGAP sequence specifically described below. In general, the methods include introducing the polynucleotide into host cells, typically as a recombinant vector including the polynucleotide, culturing the host cells under conditions suitable for propagating the polynucleotide and purifying the polynucleotide from the host cells to obtain larger portion of the isolated polynucleotide therefrom. Host cells useful for propagating the polynucleotides and/or polypeptides of this invention can be eukaryotic, prokaryotic, more particularly, fungal, yeast, animal or insect as desired. Host cells amenable for large scale production of mammalian SYNGAP are especially useful for commercial and industrial applications. Alternatively PCR amplification or related amplification methods can be used to isolate the polynucleotide.
Further provided are cultured host cells which have been transformed, transfected or infected either transiently or stably by at least one recombinant vector of the invention which vector includes an isolated polynucleotide that encodes a mammalian SYNGAP or a fragment or derivative thereof (DNA or RNA).
Recombinant vectors of the invention can be introduced into suitable cells or groups of such cells including tissue or organs if desired either in vitro or in vivo. Preferably, the cells are capable of expressing the recombinant vector at detectable levels. Host cells comprising the vectors can be cultured in medium capable of supporting propagation and/or expression of the vectors in the cells. The cells can be eukaryotic cells, preferably mammalian cells such as neurons and neuron-associated cells (e.g., glia) which cells are capable of expressing desired sequences in the recombinant vector. The cells can be primary cells or the cells can be immortalized. In some instances it will be desirable to introduce the vector into a suitable prokaryotic host e.g., bacteria, insect, yeast or fungal cells to propagate the vector.
The present invention also provides useful oligonucleotide primers, typically single-stranded primers, which oligonucleotide primers are complementary to a polynucleotide encoding a mammalian SYNGAP. For example, in one embodiment, the oligonucleotide primers are complementary to the rat SYNGAP sequence shown in any one of SEQ ID Nos. 1, 3, or 5. The oligonucleotide primers have a variety of useful applications, e.g., to detect or amplify a mammalian SYNGAP of interest. Exemplary oligonucleotide primers will generally have length of between about 12 to about 70 nucleotides although somewhat larger or smaller primers are useful for some applications.
Additional polynucleotides of the present invention have important uses. For example, as discussed, the invention provides for recombinant vectors that include an isolated polynucleotide relating to SYNGAP. Specific recombinant vectors can be used to produce significant amounts of nucleic acid sequence that can be sense or anti-sense, single-stranded or double-stranded as needed. Generally, RNA transcribed from DNA is referred to as the xe2x80x9csensexe2x80x9d RNA strand and oppositely oriented RNA is termed antisense RNA. Antisense polynucleotides, then, refer to sequences of DNA or RNA which can bind in a Watson-Crick fashion to a sequence on a target mRNA. See generally Bentley, D. L. and Groudine, M. (1986) Nature 321:702; and Kimelman, D. Gene regulation: Biology of Antisense RNA and DNA, R. P. Erickson, J. G. Izant, eds. (Raven Press, New York).
Mammalian SYNGAP RNA and particulary a SYNGAP mRNA existing in a biological sample such as in an excitatory chemical synapse (in vivo or in vitro) will sometimes be referred to herein as a xe2x80x9ctargetxe2x80x9d to denote potential for specific binding between a polynucleotide of interest, e.g., a suitable anti-sense RNA, and the SYNGAP mRNA in the sample.
In one preferred embodiment, the recombinant vectors include DNA sequences that encode an anti-sense RNA which RNA is substantially homologous to mammalian SYNGAP polynucleotides of this invention e.g., a rat SYNGAP. It is preferred that the recombinant vectors include cDNA sequences. In this instance it will be understood that the anti-sense RNA will usually include a uracil (U) in place of thymidine (T) where the cDNA sequence has a thymidine. In a preferred embodiment, the anti-sense RNA has a length of at least about 20 to about 50 nucleotides, at least about 100 to about 250 nucleotides, at least about 300 to about 700 nucleotides, at least about 1000 to about 2000 nucleotides and up to about 2500 to about 4000 nucleotides as determined by standard polynucleotide sizing methods. In most cases, the length of the anti-sense RNA will be guided by intended use including the length of the target and the level of anti-sense suppression desired.
The antisense RNA encoded by specific recombinant vectors of this invention is usually designed to undergo complementary base pairing (hybridization) with the target, rendering the target essentially unavailable for translation in most cases. In some instances, the antisense RNA will render the target susceptible to degradation, thereby substantially reducing the amount of the target in relevant cells or tissue. Accordingly, the recombinant vectors of the invention can be used to control the synthesis and/or expression of SYNGAP in desired neurons in vitro or in vivo.
Specific recombinant vectors of this invention that are capable of producing anti-sense RNA complementary to a mammalian SYNGAP mRNA (or more than one of such mRNA) can be used therapeutically to reduce levels of the target in vivo or in vitro. For example, in one embodiment, a desired recombinant vector is administered to a patient suffering from or suspected of suffering from a SYNGAP related disorder such as those specified herein. In this instance, the patient will benefit from substantially reduced or totally absent levels of the target. Preferred administration is sufficient to reduce levels of the target mRNA in the patient. Efficacy of the technique can be monitored and quantified if desired by a variety of techniques including Northern and Western blotting.
In another embodiment, the recombinant vector is formatted to produce anti-sense DNA of about the same size as the anti-sense RNA.
In addition, the polynucleotides of this invention and particularly the isolated nucleic acids and recombinant vectors described herein can be used as important controls for detecting and analyzing normal and aberrant mammalian SYNGAP expression in vitro and in vivo.
In another aspect, the present invention provides isolated mammalian SYNGAP preferably having an apparent molecular weight of between about 100 to about 150 kDa. The molecular weight of the SYNGAP can be determined by a variety of standard means including polyacrylamide gel electrophoresis. Preferred are polypeptide sequences having at least about 200 amino acids up to about 1000 amino acids or greater including about 1100 amino acids. Specifically provided is rat SYNGAP as shown at the amino acid sequence level in any one of SEQ ID NOS. 2, 4, or 6.
In another aspect, there is provided an isolated polypeptide having at least about 170 percent amino acid sequence homology to the sequence illustrated in SEQ ID NO: 2, 4, or 6. Additionally provided are isolated fragments or derivatives of that polypeptide.
The invention also provides methods for producing a mammalian SYNGAP in which the method includes culturing the host cell in medium under conditions suitable for expression of the SYNGAP in the host cell or medium.
In another aspect, the present invention provides an antibody or antigen-binding fragment thereof capable of binding the amino acid sequence shown in SEQ ID NO. 6. Preferably, the binding of the antibody or antigen-binding fragment is blocked by at least about 80%, 90% or more up to about 100% by contact with the amino acid sequence shown in SEQ ID NO:21 or a sequence substantially homologous thereto. The percent blocking by the amino acid sequence can be determined by a variety of means including a conventional immunoprecipitation assay or a Western immunoblot.
In one preferred embodiment, the antibody is a capable of binding excitatory synapses as determined by microscopy. In this embodiment, the antibody can be a monoclonal or a polyclonal antibody or an antigen-binding fragment thereof. In most cases, the antibody or antigen-binding fragment will be detectably-labeled or will be capable of generating a detectable label as specified below to help visualize and optionally quantitate synaptic binding.
Additionally, the invention pertains to methods for making specific immune system molecules and particularly antibodies (polyclonal, monoclonal or chimeric molecules) which bind certain SYNGAP or a SYNGAP-related molecules described herein. The methods generally include using a substantially purified sample of a desired SYNGAP polypeptide, as immunogen. Exemplarly antibodies are monoclonal antibodies obtained by conventional hybridoma manipulations. The antibodies can also be generated from an immunogenic peptide that comprises one or more epitopes of the polypeptide. It will be useful in some settings to covalently attach a suitable cytotoxic, anti-metabolic, or detectable label to the antibody by methods well-known in the field to help detect and/or modulate desired synapses that include mammalian SYNGAP and particularly human or rat SYNGAP in vitro or in vivo.
It will be understood that an xe2x80x9cimmune system moleculexe2x80x9d generally relates to antibodies and antigen-binding fragments of those antibodies that are derived from the immune system of a mammal such as a mouse, rat, rabbit, human, and the like.
The immune system molecules of this invention provide a number of uses and advantages. For example, the immune system molecules can be used to modulate mammalian SYNGAP expression in vitro or in vivo, e.g., by employing conventional microinjection techniques. In one approach, a suitable amount of a desired SYNGAP antibody is suspended in a physiologically acceptable buffer and injected into a desired cell or group of cells including tissue or an organ, which amount is sufficient to reduce or eliminate SYNGAP expression as determined, e.g., by the biochemical and functional assays described herein. In some instances, such approaches will be well-suited to modulate SYNGAP function, e.g., to impact neuronal plasticity in vitro or in vivo.
In another aspect, the present invention provides a kit that typically includes at least one container means comprising at least one of: 1) an antibody or antigen-binding fragment thereof capable of binding mammalian SYNGAP, 2) an isolated polynucleotide comprising sequence with at least about 70% sequence homology to any one of the sequences shown in SEQ ID NOs: 1, 3, or 5; 3) a pair of oligonucleotide primers capable of hybridizing to any one of the sequences shown in SEQ ID NOs: 1, 3, or 5, preferably under high stringency conditions; and 4) a polypeptide with at least about 70% to about 100% sequence homology to the sequence shown in any one of the sequences shown in SEQ ID NOs: 2, 4, or 6 a fragment or a derivative thereof.
In one embodiment, the kit includes a system for: 1) treating or preventing a disorder in a mammal associated with the SYNGAP, 2) detecting excitatory synapses in a cell or group of cells in vitro or in vivo, or both.
Isolated polynucleotides and polypeptides of the invention can be obtained as a substantially pure preparation if desired. That is, the nucleic acids and polypeptides can be isolated in substantially pure form by standard methods and can be provided as sterile preparations if desired. Methods for providing substantially pure preparations of nucleic acids and polypeptides are discussed below.
In another aspect of the present invention, there is provided methods for modulating excitatory synapse function in a cell or group of cells in vitro or in vivo. In this embodiment, the method includes administering to the cells a modulation effective amount of at least one polynucleotide of this invention or fragment or derivative thereof By the term xe2x80x9cmodulation effectivexe2x80x9d is meant a change in exciting synapse function (e.g., amout of SYNGAP).
The present invention also includes methods for modulating excitatory synapse function in a cell or group of cells in vitro or in vivo. In this embodiment, the method includes administering to the cells a modulation effective amount of a mammalian SYNGAP of this invention, including a fragment or derivative thereof
In embodiments of the methods for modulating excitatory synapse function, the modulation can include an increase in excitatory synapse number. Excitatory synapses can be visualized and quantified, if desired, by a variety of means including microscopy or centrifugation.
In another aspect, the present invention includes methods for treating a disorder associated with a mammalian SYNGAP including administering to a patient suffering from or susceptible to such disorder an effective amount of at least one isolated polynucleotide of this invention or fragment or derivative thereof
Additionally provided are methods for treating a disorder associated with a mammalian SYNGAP comprising administering to a patient suffering from or susceptible to such disorder an effective amount of an isolated SYNGAP of this invention or fragment or derivative thereof
In embodiments of the methods for treating SYNGAP-related disorders, the disorder can be a neurological disorder of the central (CNS) or peripheral (PNS) nervous system. In a specific embodiment, the CNS disorder is at least one of an affective disorder, a cognitive disorder, or a neurodegenerative disorder. For example, the affective disorder can be depression; the cognitive disorder can be at least one of memory loss, a learning disability, or schizophrenia; and the learning disability can be attention deficit disorder (ADD). In another specific embodiment, the degenerative disorder can be at least one of Parkinson""s disease (PD), Huntington""s disease (HD), senile dementia, or Alzhemier""s disease (AD). In a further specific embodiment, the PNS disorder is amyotrophic lateral sclerosis. In another specific embodiment, the neurological disorder is associated with at least one of trauma, an immune response or ischemia.
The methods for treating or preventing the SYNGAP-related disorders will also find applicability in the manipulation of habituation, sensitization, learning and memory in research settings.
As noted, the present invention provides a variety of screening assays for detecting compounds and especially pharmaceutical agents for use in the diagnosis or treatment of disorders impacted by mammalian SYNGAP.
In one embodiment, there is provided methods for identifying a compound useful in the diagnosis or treatment of a disorder relating to the SYNGAP. In a specific embodiment, the method includes at least one of the following steps:
a) culturing cells capable of forming synapses comprising SYNGAP under conditions conducive to forming or maintaining synapses,
b) contacting the cells with a candidate compound,
c) analyzing the cells for an increase or decrease in the number of synapses; and
d) detecting the increase or decrease as indicative of the compound useful in the diagnosis or treatment of the disorder relating to SYNGAP.
In another embodiment, the invention provides methods for detecting a compound capable of modulating a Ras-activated second messenger pathway. In this embodiment, the method includes at least one of the following steps:
a) providing a Ras response system comprising a recombinant mammalian GTPase Activating Protein at Synapses (SYNGAP),
b) contacting the Ras response system with a candidate compound,
c) analyzing the Ras response system for an increase or decrease in Ras activity; and
d) detecting the increase or decrease in the Ras activity as indicative of the compound capable of modulating the Ras-activated second messenger pathway.
In preferred embodiments, the Ras response system which includes at least one of Ras a nucleotide di- or tri-phosphate; adenylate cyclase; and an isolated polynucleotide encoding the SYNGAP or a fragment or a derivative thereof. The Ras response system can be provided by one or a combination different strategies including being provided in a host cell or a group of host cells (e.g., tissue or an organ), or a lysate of the cells, tissue or organ.
In another embodiment, the invention includes methods for detecting a compound capable of modulating a Ras-Raf (MAP kinase) cascade. The RAS-GTPase activating activity of SYNGAP can be measured using the MAP kinase as a reporter in a suitable primary or cultured cells. In this embodiment, the method includes at least one of the following steps:
a) transfecting a cells with a reporter gene construct capable of being modulated by Ras-Raf (MAP kinase),
b) transfecting the cells with a polynucleotide encoding any one of the SYNGAP sequences shown in SEQ ID Nos. 2, 4, or 6,
c) contacting the cells with a candidate compound; and
d) detecting the reporter gene construct as being indicative of the compound capable of modulating the RasGTPase activity of SYNGAP.
It will be appreciated that in some embodiments of the method, the SYNGAP cDNA can be provided before, during or after transfection of the reporter gene construct.
In another embodiment, the invention includes methods for detecting a compound capable of modulating a phospholipid-activated second messenger pathway. In this embodiment, the method includes at least one of the following steps:
a) providing an inositol triphosphate response system comprising a recombinant SYNGAP,
b) contacting the inositol triphosphate response system with a candidate compound,
c) analyzing the inositol triphosphate response system for an increase or decrease in phospholipase activity; and
d) detecting the increase or decrease in the phospholipase activity as indicative of the compound capable of modulating the phospholipid-activated second messenger pathway. In preferred embodiments, the phospholipase is a phosphoinositide-specific enzyme such as phospholipase C.
In one embodiment of the method, the inositol triphosphate response system further includes at least one of phosphatidylinositol (PI) and an isolated polynucleotide encoding the SYNGAP or a fragment or a derivative thereof In a specific embodiment, the inositol triphosphate response system includes at least one of diacylglycerol, protein kinase C, and inositol 1,4,5 triphosphate (INSP3). In a related embodiment, the phospholipid-activated second messenger pathway is capable of at least one of calcium (Ca+2) release or protein phosphorylation. The inositol triphosphate response system can be provided by one or a combination different strategies including being provided in a host cell or a group of host cells (e.g., tissue or an organ), or a lysate of the cells, tissue or organ.
In another embodiment of the method, at least the pleckstrin (PH) homology of SYNGAP is provided for use with the inositol triphosphate response system. In another embodiment of the method, at least the PH homology and the SYNGAP C2 domain is used with the inositol triphosphate response system.
Additionally provided by the present invention are methods for detecting a compound capable of modulating phospholipid-dependent calcium (Caxe2x88x922) binding to at least the SYNGAP C2 domain. In this embodiment, the method includes at least one of the following steps:
a) mixing a phospholipid, calcium (Ca+2) and at least the SYNGAP C2 domain, the mixing being under conditions conducive to forming a complex,
b) contacting the mixture with a candidate compound,
c) analyzing the mixture for formation of the complex; and
d) detecting the complex as indicative of the compound capable of modulating the phospholipid-dependent binding between the calcium (Ca+2) and at least the C2 domain of SYNGAP.
In another embodiment of the method, at least the PH homology and the SYNGAP C2 domain is used with the inositol triphosphate response system.
In embodiments of the present invention in which use of rat SYNGAP is desired, it will be possible to use a polypeptide sequence that includes and preferably consists of the sequence represented by any one of SEQ ID Nos: 2, 4 or 6. Also contemplated are embodiments of the present invention which include use of a fragment or a derivative of the rat SYNGAP sequence as show in SEQ ID NOs: 2, 4, or 6.
The methods for detecting the compound capable of modulating the phospholipid-activated second messenger pathway are flexible and can be used if one or more components of the inositol triphosphate response system, e.g., PI, act xe2x80x9cupstreamxe2x80x9d or xe2x80x9cdownstreamxe2x80x9d of SYNGAP.
In another embodiment of the present invention, there is provided methods for detecting a test amino acid sequence capable of binding a mammalian SYNGAP. In one embodiment, the method includes contacting the test amino acid sequence with the mammalian SYNGAP or a binding fragment or the derivative thereof under conditions conductive to forming a complex; and detecting the complex as indicative of the test amino acid sequence capable of binding the SYNGAP. For example, in a specific embodiment, the mammalian SYNGAP can be the rat SYNGAP sequence shown in SEQ ID NO: 6 or a suitable fragment or derivative thereof. Detection of the binding can be acomplished by one or a combination of different strategies such as at least one of immunoprecipitation, affinity chromatography, or a suitable biosensor assay.
In another embodiment of the method for detecting a test amino acid sequence capable of binding a mammalian SYNGAP, the contacting step can be performed in cells and the detecting step can include monitoring expression of a detectable gene product expressed by the cells In a preferred embodiment, the steps are performed in yeast cells.
Particulary preferred are those yeast cells (strains) that are suited for performing what is generally known in the field as a xe2x80x9ctwo-hybridxe2x80x9d assay or a related term. In other embodiments, the detecting step of the methods can include screening a polypeptide expression library or a combinatorial peptide library, e.g., by a hybridization type assay using a suitable polynucleotide of this invention, for the test amino acid sequence.
In preferred instances, the methods for detecting the binding between the test amino acid sequence and the SYNGAP will typically register binding between at least one PDZ domain in the test amino acid sequence and a C-terminal sequence in the SYNGAP or fragment or derivative thereof. Preferably, the C-terminal sequence has the following general sequence: (T or S) XV; wherein X is an amino acid as defined below. The SYNGAP sequence used can be the rat SYNGAP sequence disclosed in SEQ ID NO: 2, 4, or 6 as well as suitable fragments or derivatives of that sequence.
Additionally provided are amino acid sequences detected by any of the methods for detecting the test amino acid sequence capable of binding a mammalian SYNGAP.
The present invention includes, in another aspect, methods for detecting excitatory synapses in a cell or group of cells. In one embodiment, the methods include contacting the cells or group of cells with an antibody or antigen-binding fragment of this invention under conditions sufficient to detect the excitatory synapses in the cells or group of cells. In a specific embodiment, the antibody or antigen-binding fragment is detectably-labeled or is capable of producing a detectable label. Illustrative of such labels include a radionuclide; a protein tag; a chromophore; a fluorescent, chemiluminescent or phosphorescent molecule. In another specific embodiment the antibody or antigen-binding fragment thereof is labeled with at least one enzyme capable of producing a chromophore, a fluorescent, chemiluminescent or phosphorescent molecule.
In another aspect, the present invention provides a library that includes a plurality of the polynucleotides or the polypeptides of this invention including fragments or derivatives of those polynucleotides or polypeptides. Illustrative of such libraries include cDNA and genomic DNA libraries, combinatorial and peptide expression libraries.

Signal transducing synaptic molecules and uses thereof


Disclosed system and methods for detecting malicious applications. The system provides a library of handler functions. The handlers functions control access of one or more applications to protected resources on a user device. The system also modifies the one or more applications to access the library of handler functions instead of corresponding application program interface (API) functions of the user device. The handler functions receive API function calls from a modified application. The system analyzes the received API function calls for malicious behavior characteristics. When the API function calls do not exhibit malicious behavior characteristics, the handler functions perform the API function calls to the protected resources. When the API function calls exhibit malicious behavior characteristics, the system prevents access of the modified application to the protected resources.

The number of mobile communication devices is constantly growing, and they have penetrated into all areas of human life. The most popular of these are telephones, smartphones, notebooks, tablets and navigation systems. Mobile devices are primarily used for communication and entertainment. In parallel with the development of these devices, new wireless communication systems, networks and protocols are also being developed. The transmission of data from one device to another and the exchange of information are tasks that even an inexperienced users can perform without difficulty. But the development of new technologies brings not only benefits, but also creates problems.
On one hand, the development of mobile devices and wireless communication systems simplifies human interaction facilitates exchange of information, and erases geographical boundaries. On the other hand, new technologies create new security problems. The ease of copying and exchanging of information causes leaks, whether deliberate or accidental, of confidential personal or business information from organization networks. Therefore, information security specialists responsible for administration of information security infrastructure need to extra vigilant in protecting corporate IT systems in order prevent information leaks and various security breaches.
In addition, the growth in popularity of mobile platforms, such as Android and iOS, is attracting the attention of hackers who have various goals—from a desire to play jokes or show off, to the theft of confidential information for the purpose of extortion and blackmail. As a result of this, computer threats, such as viruses, worms and other types of malware, which were previously characteristic of desktop computers are beginning to appear on mobile platforms as well. New threats unique to mobile devices are also appearing, such as Trojan horses, which after being installed on a mobile device, send SMS messages to telephone numbers or use geolocation services to track the movements of the device user.
Therefore, there is a need for a security system to protect mobile devices from malware, information leaks, unauthorized access and other security threats.

System and method for detecting malware using isolated environment


An anti-backlash assembly of the type for transferring motion via a pin movable along a slot. The assembly comprises a pivot lever and a second lever member. The pivot lever comprises a body portion and a pivot lever portion extending from the body portion to a pivot lever distal end. The pivot lever portion defines a pin engagement slot extending from adjacent the pivot lever distal end toward the body portion. A pair of fingers extend in parallel and spaced relationship from the pivot lever distal end of the pivot lever portion toward the body portion to engage opposite sides of a pin of the engagement arm. Each of the fingers includes a pin engaging section extending to an inwardly extending distal end and a spring section extending in a for urging the fingers into clamping engagement with the pin.

1. Field of the Invention
An anti-backlash assembly of the type for transferring motion via a pin movable along a slot.
2. Description of the Prior Art
The lever assembly to which the subject invention pertains transfers rotary motion to motion of the pin along the a slot, or vice versa. One such lever is disclosed in U.S. Pat. No. 6,658,957 granted to Nishishita, which discloses an lever assembly with a pivot lever including a body portion disposed in a plane for disposition on a shaft on an axis and a pivot lever portion extending radially from the body portion to a pivot lever distal end. A second lever member is coupled to the pivot lever with the coupling including a pin engagement slot and a pin slideable along the slot.
Such prior art levers are successful, however, too much of a gap may exist between the pin and the slot, allowing too much tolerance. This gap can cause the pin to rattle within the slot throughout its range of movements.


The invention includes an etchant composition containing isopropyl alcohol and one or more of HF, NH4F and tetramethyl ammonium fluoride (TMAF). The invention encompasses a method of processing a substrate. A substrate is provided which has a first material containing at least one of polysilicon, monocrystalline silicon and amorphous silicon, and a second material. The substrate is exposed to an etch composition which comprises isopropyl alcohol and at least one of HF, NH4F and TMAF. The invention includes a method of processing a semiconductor construction including providing a construction which has a capacitor electrode material and an oxide material along at least a portion of the capacitor electrode material. At least some of the oxide material is removed by isotropic etching utilizing an etchant composition comprising isopropyl alcohol.

Etching technology is frequently utilized for removing one or more materials during a variety of technical fabrication processing including, for example, semiconductor processing. In many applications an etch chemistry is chosen to selectively remove a first material relative to a second material. An etch is considered to be selective for a first material relative to a second material (or to be selective to the second material) if the etch removes the first material at a faster rate than the second material.
During semiconductor processing, wet etch techniques are commonly utilized to isotropically etch dielectric materials selectively relative to silicon materials. Wet etchants for isotropic oxide removal include etchants referred to as buffered oxide etchants (BOEs). Although such etchants are highly selective to silicon, some silicon etching can occur during the wet etch process. The slow etching of Si which occurs during wet etch processing can result in roughening of silicon surfaces, or unintentional thinning or removal of silicon materials. For example, removal of a gate oxide material during semiconductor fabrication can result in roughening of an underlying silicon surface which can ultimately compromise gate oxide integrity and/or tunneling oxide properties. Additional problems can arise during memory array formation where wet etch processing is utilized to remove dielectric materials within the array area near the periphery. Specifically, unintentional removal of a protective silicon material can expose dielectric material in the periphery leading to periphery blowout, where the term "blowout" refers to unintentional etching of material.
It is desirable to develop alternative etchant compositions and methods of processing substrates and semiconductor constructions.

Etch compositions and methods of processing a substrate


A sulfopolyester containing residues of 2,2,4,4-tetralkylcyclobutane-1,3-diol such as 2,2,4,4-tetramethylcyclobutane-1,3-diol is highly dispersible in water. This allows one to reduce the content of sulfonate groups or reduce the amount of ethylene glycol or other hydrophilic glycols to retain good water resistance in cured coatings. The sulfopolyester may also be a reaction product of a 2,2,4,4-tetralkylcyclobutane-1,3-diol along with 1,4-cyclohexanedimethanol, neopentyl glycol, or a mixture thereof with an acid component. Coating compositions may also contain these sulfopolyesters along with water and a polymer resin.

Sulfopolyester polymers are generally referred to polyesters containing ionic sulfonate (SO3−) groups, particularly to those synthesized using a sulfomonomer such as, for example, 5-sodiosulfoisophthalic acid (5-SSIPA or SIP) or dimethyl 5-sodiosulfoisophthalate, as one of the diacids in the polyester compositions. Such sulfopolyester polymers are commercially available from Eastman Chemical Company under the Eastman AQ™ family of polymers. The polymers are linear, amorphous polyesters that can be dispersed in water without the assistance of surfactants or amines. This water dispersibility is attributed to the ionic nature of the sulfonate substituents attached to the polymer chains.
The common Eastman AQ™ polymers are Eastman AQ 38S, AQ 48 ultra, and AQ 55S, which have Tg ranging from 35° C. to 55° C. as denoted by the names of the products. Another product, Eastek™ 1200 polymer, has a higher Tg of about 63° C.; it has applications in printing inks, overprints, and primers for industrial wood coatings. The product is capable of providing excellent water resistance, alcohol resistance, and is fast drying. These properties are also desirable for top coat application in wood coatings; as a result, there have been increased interests in Eastek™ 1200 polymer for wood coating application as a top coat. In addition to sulfonate groups, sulfopolyester polymers typically contain the residue of a hydrophilic polyhydroxyl compound such as ethylene glycol (EG), diethylene glycol (DEG), or polyethylene glycol (PEG) in the polymer chains to further impart water dispersibility. Such EG-based glycols, however, are not preferred for applications in coatings due to their deficiency in hydrolytic stability. Although the water dispersibility of a sulfopolyester polymers without EG-based glycols can be retained by increasing the ratio of the sulfonate groups, it is desirable to keep the sulfonate content low in order to obtain adequate water resistance of the coatings. It is, therefore, an object of this invention to prepare sulfopolyester polymers with low sulfonate content, low EG-based glycol content, yet retain good water dispersibility.
The EG-based glycol components may be replaced by diols that are known to be more hydrolytically stable, such as neopentyl glycol (NPG) and cyclohexane dimethanol (CHDM). Sulfopolyester polymers based on such diols, however, are not as water dispersible as those based on EG, DEG, or PEG. A need therefore exists for a diol component that is not EG-based but is capable of providing good water dispersibility. The advantage of such a solution is to either improve the water dispersibility of such a polymer compared to other non-EG-based glycol modified sulfopolyesters having the same quantity of sulfonate groups, or retain an equivalent level of water dispersibility with a lower amount of hydrophilic groups, thereby improving the water resistance of the coating.

Sulfopolyester polymer compositions with improved water dispersibility


A method and apparatus for enabling communication between iSCSI/SAS host devices and iSCSI/SAS target devices via an iSCSI/SAS router is disclosed. In one embodiment, an iSCSI/SAS router includes iSCSI interfaces, SAS interfaces, and a mapping and routing logic. The mapping and routing logic is coupled to the iSCSI interfaces and the SAS interfaces. The mapping and routing logic virtualizes one or more SAS host and target devices to communicate with discovered one or more iSCSI host and target devices. Further, the mapping and routing logic virtualizes the one or more iSCSI host and target devices to communicate with discovered one or more SAS host and target devices.

As electronic business (e-business) grows, so does the need of better ways to share and manage large amounts of data. The amount of data storage required by today's e-businesses is staggering. Today almost all client access to large scale storage is accomplished by sending request through internet small computer system interface (iSCSI) based devices that connect an internet protocol (IP) network, example a local area network (LAN) or a wide area network (WAN), to storage network, example a storage area network (SAN).
However, serial attached SCSI (SAS) is an emerging industry standard as the enterprise-class standard storage interface. A typical SAS domain comprises SAS devices (e.g., host devices or initiators and target devices) interconnected with a set of point-to-point-links. For example, SAS can support up to 16,384 addressable devices in the SAS domain and point-to-point data transfer speeds up to 6 Gbit/s. In the SAS domain, the SAS devices use SAS commands for interacting with other SAS devices.
Another standard used for exchanging data between a host device and a target device using SCSI commands is iSCSI standard. iSCSI is a TCP/IP based protocol that carries the SCSI commands over an IP network to enable communication between iSCSI host and target devices. An advantage of iSCSI standard is that the iSCSI enables the iSCSI host and target devices to communicate with each other with no limitation to distance between them. Unlike the iSCSI, the SAS enables the SAS host and target devices to communicate with each other with limited operating distances. Currently, the SAS devices can span up to a range of about 6-7 meters. As a result, the SAS devices are mostly restricted to point-to-point connectivity and hence operate within limited operating distances.
Today, almost all backend storage devices are SAS devices or drives. The SAS devices are made part of the SAN through controllers which act as the SAN interface to the SAS devices. The SAS devices do not have any iSAN interface. Also currently, there are no SAS to iSCSI converters available for enabling the traffic between SAS devices and iSCSI devices.

Method and apparatus for enabling communication between iSCSI devices and SAS devices


Radically polymerizable macrocyclic polyethers or macrocyclic heteroanalogous polyethers, preferably crown ethers, heteroanalogous crown ethers and cryptands for use in dental materials is disclosed.

Curable dental materials based on an organic monomer matrix which can be cured during application by radical polymerization are known. Examples of such dental materials are adhesives, coatings, cements or composites. With all of these materials, a good substrate adhesion, whether to dentine or tooth enamel or to another dental material, is an important property which can, however, be achieved only with difficulty due to the poor compatibility of organic monomers or polymers on the one hand and the substrate, e.g. dentine or tooth enamel, on the other.
Macrocyclic polyethers (crown ethers) and their heteroanalogous compounds (coronands or corands) in which the O atoms are partially or completely substituted by heteroatoms, above all nitrogen and sulphur, and also bicyclic crown ethers (cryptands) are known in the state of the art (e.g. C. J. Pedersen, H. K. Frensdorf, Angew. Chem. 84 (1972) 16). Examples of crown ethers or their heteroanalogous compounds are [15]crown-5, [18]crown-6 or 1,10-diaza-[18]crown-6:

In a simplified nomenclature, a crown ether with x ring members and y oxygen atoms is called [x]crown-y (cf. Ullmann's Encyclopedia of Industrial Chemistry, 5th Ed., Vol. A8, 91 et seq.). Additional substituents are placed in front of the name as a prefix, such as e.g. dibenzo[18]crown-6. In the case of bicyclic crown ethers (cryptands), the bridging takes place via two nitrogen atoms, wherein in the designation the number of oxygen atoms of the first, second and third bridges are placed in front of the word cryptand separated by full stops, e.g. [2.2.2]cryptand:

Polymerizable macrocyclic polyethers, such as e.g. polymerizable crown ethers, are also known. Thus for example ion-binding polymers are obtained by radical polymerization of e.g. 4-vinylbenzo-[15]crown-5 (VB15C5) or 4-vinylbenzo-[18]crown-6 (VB18C6) (S. Kopolow, T. E. Hogen Esch, J. Smid, Macromolecules 6 (1973) 133).

CH-A-631 344 describes a fluoridating composition for dental enamel which, in addition to potassium or sodium fluoride and an organic solvent, such as e.g. acetone, acetonitrile or ethyl acetate, contains a complexing agent, preferably a crown ether (e.g. [18]crown-6) or a cryptand (e.g. [2.2.2]cryptand). The named macrocyclic polyethers are not polymerizable compounds. Rather, the crown ethers used bring about merely a dissolution of the fluoride, accompanied by formation of fluoride ions without a solvation shell. Poorly solvated fluoride ions are said to be absorbed better by the tooth enamel.

Use of polymerizable macrocyclic polyethers and macrocyclic heteroanalogous polyethers in dental materials


A method of assembling elements into a valve block is described, especially for use with slip-controlled brake systems, a sealing ball, valve covers or valve seats are inserted into the valve block to be sealed without using force fitting swaging or other mechanical working of the parts, to avoid contaminating abrasive particles generated by those processors. The elements are inserted into openings in the valve block housing with a clearance fit, and heated to a temperature higher than the austenite starting temperature causing a memory shape to be restored, which provides a sealing fixation of the elements in respective openings in the valve block.

The present invention is concerned with a valve block, in particular, for use with slip-controlled brake systems.
For example, DE-OS NO. 37 29 216 discloses a valve block in the valve block housing in which a plurality of hydraulic passageways establish communication between the pressure fluid supply and pressure fluid discharge ports. Valve magnets are provided in the valve block housing and are held on the valve domes, acting as adjusting elements. Passageways completely passing through the valve block housing have a ball element swaged in and acting as a blocking element to seal the passageways.
In the afore-described ball-type seal, there is a great danger of a passageway contamination as a result of abrasion during assembly of the ball. Such contamination has proved to be disadvantageous as having an adverse interfering effect on the magnetic valves during their adjusting movement, and this possible contamination requires that all residues in the passageways be removed during the rinsing operation of the valve block. The use of "king" expanders or plugs of equivalent shape has not proven effective to solve this problem.
Moreover, insertion of the valve domes and associated valve seats into the valve block housing has presented problems as these parts to be inserted also exhibit a surface-damaging force fit and are swaged, so that similarly to the afore-described ball installation, assembly of the valve domes and spots seats can add to contamination of the passageways.
It is, therefore, the object of the present invention to provide a valve block of the afore-mentioned type which, avoiding the afore-described disadvantages, permits an improved fixation of the parts inserted into the valve block without resulting in surface abrasion and thus contamination of the passages.

Valve block and method of assembling an element thereto


A process for making a continuous fiber reinforced thermoplastic article that is sufficiently strong and rigid for use as a structural member includes the steps of providing a tubular composite parison that has continuous, axially oriented glass fiber filaments dispersed in a thermoplastic resin and admitting pressurized air into the interior of the tubular parison to expand the tubular parison to a desired shape.

This invention relates to a process for making a thermoplastic article and more particularly to a process for making a fiber reinforced thermoplastic article that can be used as a structural member, such as a cross car beam or a structural instrument panel carrier.
U.S. Pat. No. 5,088,571 issued to Donald L. Burry and Leonard J. Pilato Feb. 18, 1992 discloses a modular instrument panel carrier that comprises a box shaped beam that extends transversely across the vehicle. The box shaped beam is made from two panels of fiber reinforced plastic that have flanged edges that are bonded together. The patent specification does not describe the manufacturing process for the panels or give any particulars for the reinforcement fibers or the plastic.
It is known that continuous fiber reinforced thermosetting plastic articles can be made by pultrusion. Pultrusion is a primary fabrication process for making continuous-length filament reinforced plastic composite profiles. In the common form, reinforcing filaments, such as glass fiber roving, saturated with catalyzed thermoset resin, are continuously pulled through a shaped orifice in a heated steel die. As the two materials, filaments and resin, pass through the die, polymerization of the resin occurs to continuously form a rigid cured profile corresponding to the die orifice shape. Resin materials include rigid and flexible polyesters and combinations thereof, vinyl esters, one-part epoxies that cure like polyesters, and high performance resins based on urethane reaction with vinyl esters and unsaturated polyesters. Reinforcing materials include glass fiber rovings, graphite fibers and nonwoven veils. While this process is capable of making structural members, the articles produced by this process are necessarily limited to straight, constant shaped products or profiles that can be configured by an orifice in a heated steel die or the like because the thermosetting plastic material is hardened permanently after it leaves the heated die.
It is also known from U.S. Pat. No. 3,023,461 granted to Orville B Sherman Mar. 6, 1962 and from U.S. Pat. No. 5,085,821 granted to Shigero Nohara Feb. 4, 1992 that small, hollow, non-structural thermoplastic articles of changing shape, such as double layered bottles, can be made by combining extrusion and blow molding processes. The thermoplastic articles produced by this combined process are not limited to straight, constant shape products or profiles that can be configured by an orifice of an extruder. Indeed, the combined process is capable of producing small, hollow products that are straight or curved and having either constant area and changing shape or changing area and changing shape. However, the physical properties, particularly the strength and structural rigidity, of these hollow thermoplastic articles are limited by the wall thickness and/or hollow shapes that can be generated by the combined process because no reinforcement is used.
On the other hand, it is further known from U.S. Pat. No. 4,731,216 granted to Alvin S. Topolski Mar. 15, 1988 that small, hollow, non-structural thermoplastic articles, such as bottles, can be made by a combined extrusion and blow molding process from a reinforced mixture comprising a thermoplastic matrix having particles of an incompatible thermoplastic material for reinforcement. However, the physical properties, particularly the strength and structural rigidity, of the hollow, reinforced thermoplastic article are limited to those that can be obtained from a mixture that is discontinuous by nature.

Process of making a continuous fiber reinforced thermoplastic article


A geothermal power plant operating on geothermal fluid includes a source of low pressure geothermal steam, and a turbo-generator including a low pressure steam turbine to which the geothermal steam is applied, and a generator coupled to the steam turbine. The low pressure geothermal steam is applied to the turbine, wherein expansion takes place driving the generator, and producing expanded steam that is exhausted from the turbine. A condenser for condensing the expanded steam includes a steam heat exchanger for receiving the expanded steam and a fan for cooling the expanded steam such that steam condensate is produced.

Producing power from geothermal heat sources currently is being carried out on a wide scale throughout the world. In many installations, hot geothermal fluid extracted from a production well is flashed into steam in a separator that produces geothermal steam rich in non-condensable gases, such as hydrogen sulfide, carbon dioxide, etc. The brine that remains is sometimes so corrosive, and sometimes contains such a high concentration of silica, that from an engineering standpoint, it must be disposed of in a re- injection well without using a significant amount of heat contained therein.
The geothermal steam produced from the separator is applied to a steam turbine coupled to a generator, the steam expanding in the turbine and driving the generator which produces electricity. Condensing expanded steam in the turbine exhaust is often a problem when an inadequate supply of ambient cooling water is available. Usually, the expanded steam contains noxious gases which, for environmental reasons, precludes the use of an open system condenser, such as cooling towers, for condensing the steam even when ambient water is available. Under such conditions, chemical pre-treatment of the exhaust steam is usually required to suppress escape of the non-condensable gases. This can be an expensive, and complex, solution to the problem.
It is therefore an object of the present invention to provide a geothermal power plant and a condenser therefor which is capable of operating without significant amounts of ambient cooling water, and which avoids the problems described above.

Geothermal power plant and condenser therefor


A method for hole-filling in 3D models includes identifying vertices adjacent to hole boundaries in a mesh of points on a digital image and constructing a signed distance function based on vertices adjacent to hole boundaries. A Radial Basis Function is fit based on the constructed signed distance function and evaluated on a grid, which include the hole. The points on the hole surface are extracted and meshed to fill the hole.

1. Field of the Invention
The present invention relates generally to methods and systems for processing image data, and more particularly, to methods and systems for computing complete surfaces from collections of coordinates representing points on an object's surface to generate images or for producing three-dimensional physical replicas or models of objects.
2. Description of the Related Art
The creation of three-dimensional digital objects by scanning real objects or scenes has become common practice in computer graphics applications, computer-aided design and manufacturing, urban planning, training simulations and building construction. Computer graphics applications include computer games, e-commerce and virtual museums, for example. Computer-aided design applications include scanning objects to be used as starting points for the design of new products. For example, features on existing objects can be numerically deleted and added to create new designs.
Computer-aided manufacturing applications include measuring an object to generate a physical replica using a three-dimensional hard copy device that produces objects by building layers of polymeric material, or sintering successive layers of metallic powders. Scans on the scale of the size of cities may be obtained for planning purposes such as assessing the impact of a new building or highway. In military applications, scanned cities or buildings may be used in virtual reality training simulations. In building construction, an existing building may be scanned to generate "as built" drawings to be used as a starting point for building remodeling. The following description has been restricted for convenience to techniques involving instruments that capture range images (in which each pixel value represents depth).
In general, a variety of techniques can be used to capture digital models of physical objects, including CAT scans and structure from motion applied to video sequences. Popular devices for capturing range images include laser triangulation systems (for objects less than a meter in largest dimension), laser time of flight systems (for objects up to a few hundred meters in size) and LIDAR (LIght Detection And Ranging), for scenes on the scale of the size of cities.
The basic operations necessary to create a digital model from a series of captured images may be as follows. After outliers (stray or unnecessary digital points) are removed from the range images, they are in the form of individual height-field meshes. A first step aligns these meshes into a single global coordinate system. In some systems this alignment is performed by tracking, for example, by a coordinate measurement machine for small-scale scanners, or by GPS (global positioning system) for large-scale scanners. In many systems, registration is performed by manual alignment. The alignment is then refined automatically using techniques such as the Iterative Closest Point (ICP) algorithm of Besl and McKay (See, "A method for registration of 3-D shapes"; by Besl, P. J. and McKay, H. D. in Patten Analysis and Machine Intelligence, IEEE Transactions, Vol. 14, Issue 2, February 1992, pp. 239-256), which is known in the art.
After registration, scans do not form a single surface, but often interpenetrate one another due to acquisition errors, primarily along the line-of-sight in each scan. These need to be corrected in a scan integration.
At the end of scan integration, there may be holes in the final surface, which are not physically present in the object. These holes are the result of the scanning device not being able to sample parts of objects. Areas may not be sampled if they are dark and so reflect little light, if they are partially hidden by another part of the object, or in the case of urban modeling, if objects such as buildings are hidden by trees, etc. These are artificial holes that result from the scanning process and are unacceptable in almost all applications that use 3D models.
In any computer graphics applications or visual simulation, large black regions on an object are unrealistic and distracting. In computer-aided design and manufacturing it is physically impossible to reproduce an object with an incomplete surface definition.
It should be noted that in addition to holes occurring because of problems with data collection, holes may also be created by users in a computer aided-design system. In editing an object a user may want to delete a feature such as a handle or embossed pattern that was part of the original object. In this case also, a method for filling the hole that is left is needed.
Prior art methods for hole-filling are either implemented as a post-processing operation, applied after surface reconstruction, or they are integrated into a surface reconstruction technique. For hole-filling as post-processing, a widely used approach is to triangulate each connected component of the surface's boundary, which results in filling each hole with a patch that has the topology of a disk. This works well for simple holes in almost flat surfaces, but for more complex cases it results in self-intersecting surfaces.
Two methods attempt to avoid this problem, Davis et al., 3DPVT, Padua, June 2002, in "Filling Holes in Complex Surfaces Using Volumetric Diffusion" developed a technique for filling holes by using a diffusion operation to expand the "signed distance function" defined by the existing points to complete a volumetric representation of the surface. But, the method lacks control over properties of the surface, like curvature, which leads to surfaces not matching the topology of the object being modeled (scanned) in some cases.
Correct topology may require additional information from the user, such as the original lines-of-sight used in scanning which may not be available, or in the case of holes created by a user modifying an object in a computer-aided design may never have existed.
Verdera et al., IEEE ICIP, September 2003, in "Inpainting Surface Holes" represented the surface of interest as the zero level-set of a function and then used a system of coupled geometric partial differential equations to smoothly continue the surface into the hole. This method suffers from the same lack of user control as the volumetric diffusion method. It also requires that the user provide the line-of-sight information that may not be available.
For hole-filling techniques integrated into surface reconstruction methods, Curless et al in "A Volumetric Method for Building Complex Models from Range Images", ACM SIGGRAPH 1996, converts each model mesh into a signed distance function whose zero set is the observed surface, blends these distance functions together and extracts the zero set as the final surface. However, the method does not operate properly if the scanner lines of sight do not adequately cover the volume outside the object.
Carr et al., ACM SIGGRAPH 2001, in "Reconstruction and Representation of 3D Objects with Radial Basis Functions" fits a set of radial basis functions to the data, and a weighted sum of these functions forms a new function, a level set of which is the intended surface. The method rebuilds the entire surfaces based on radial basis functions, rather than just operating on the region near the holes. Consequently, this method is slow especially for large models. For large models, such as urban scenes or complex large works of art, the computational time is impractical for building a model in time to be used for the application.
Similarly, Dey and Goswami, Solid Modeling, June 2003, in "Tight Cocone: A Water-tight Surface Reconstructor" produce a model without holes by using an alternative technique for rebuilding the model from the original points. In their method, a 3 dimensional Delauney triangulation of the points is computed. The time to perform this calculation increases as N**2 for N points, making this approach not feasible for anything but relatively small point sets.
Therefore, a system and method are needed to fill in a surface in a hole left by inadequate data in a realistic manner and remain compatible with the original physical object. In addition, a system and method are needed for building a mesh for the points that are available in a scan, in a quick and efficient manner. The system and method also need to give the user easily specified parameters for operating on any remaining holes to produce a surface that is a faithful representation of the original object.

System and method for hole filling in 3D models

International Business Machines Corporation


A color selection electrode is suspended in a color display tube from pins which are provided in the corners of the display window. The suspension means comprise a resilient element which is secured to the color selection electrode, and a slide plate slidably secured relative to the flat resilient element rigidly secured to the pin. During the suspension the position of the color selection electrode is adjusted, and the slide plate is rigidly secured to the resilient element.

The invention relates to a colour display tube comprising an envelope having a substantially rectangular display window which is provided with an upright edge, and a substantially rectangular colour selection electrode having a great number of apertures, which electrode is suspended from pins secured in the upright edge of the display window by means suspension means, each of which means comprises a resilient element which is fixedly connected to the colour selection electrode, and a slide plate having an aperture which is rigidly secured to the resilient element, each pin having a free end portion which partly projects from the aperture in the slide plate.
The invention also relates to a method of manufacturing such a colour display tube.
Such a colour display tube is known from British Patent Specification 2,097,996. Therein, each suspension means comprises a wire spring which is connected at one end to the colour selection electrode and at the other end urges the flat resilient element onto the free end portion of the pin. During the suspension of the colour selection electrode, the slide plate is slidably secured relative to the flat resilient element, the wire spring being necessary to retain the slide plate before it is rigidly secured to the flat resilient element.
In order to obtain a proper suspension of the colour selection electrode, the wire spring must firmly urge the flat resilient element onto the free end portion. This pressure should be sufficiently large that in the case of vibrations or shocks the colour selection electrode is not subject to an unfavourable change in position. Since the wire spring is secured to the colour selection electrode at one end, the colour selection electrode may be subject to deformation when the pressure exerted by the wire spring is too large. For this reason, a wire spring having a limited pressure is used in practice, the disadvantage thereof being, however, that in the case of vibrations and shocks the colour selection electrode may assume a position which differs from that when the colour selection electrode is properly suspended.

Color display tube and method of manufacturing such a color display tube


The invention relates to methods for the temporal administration of EDA agonists, in particular EDI200, which correlate to optimal therapeutic response windows required for the formation of any EDA-dependent structures such as ectodermal appendages. Use of the methods described allow for the design of targeted therapeutic dosing and administration regimens in order to correct or alter abnormal phenotypes associated with genetic disorders, in particular, XLHED.

X-linked hypohidrotic ectodermal dysplasia (XLHED) is a disease identified in humans, dogs, mice and cattle. A subset of the human disease has been attributed to a defect in the EDA gene (formerly ED1) that encodes for the protein ectodysplasin (EDA, subtype EDA-A1) which has been shown to be involved in the morphogenesis of hair follicles and tooth buds during early development. The disease phenotype is sparse or absent hair, missing and/or malformed teeth, hypoplastic eccrine glands, recurrent benign infections, and increased susceptibility to bronchitis and pneumonia (Reed et al., 1970; Nordgarden et al., 2001). There is significant morbidity and mortality in affected children due to hyperthermia, caused by the inability to sweat. Significant morbidities include increased risk of respiratory tract infections, ocular disease due to dry eyes, as well as difficulties with mastication, growth retardation, poor appearance, and speech impairment resulting from tooth abnormalities (delayed dentition, conical tooth crowns (peg-shaped teeth) and oligodontia).
The first model of XLHED was identified in mice selected from the Black 6 strain for large size which resulted in the spontaneous appearance of a sub-strain with abnormal hair and tooth development. The affected animals (designated "Tabby mice" due to the resemblance of the fur patterning of the heterozygote females to that of the tabby cat) lack functional EDA protein due to a frame-shift mutation resulting in the absence of the domain necessary for receptor binding and signaling that is critical for normal tooth, hair and sweat gland morphogenesis (Ferguson et al., 1997; Srivastava et al., 1997). Consequently, these mice have no sweat glands and no hair on the tail. The Tabby mouse currently is a widely used model for XLHED.

This patent is subject to a terminal disclaimer.

Treatment of X-linked hypohidrotic ectodermal dysplasia with a fusion EDA1 protein


A hardener composition for alkaline phenolic resole resin binders which comprises a solution of a benzylic ether resole resin in an ester. Binder compositons comprising this hardener with alkaline phenolic resole resins are further provided as are foundry molding compositions and methods for making foundry cores and molds using the molding compositions.

Binders or binder systems for foundry cores and molds are well-known. In the foundry art, cores or molds for making metal castings are normally prepared from a mixture of an aggregate material, such as sand, and a binding amount of a binder or binder system. Typically, after the aggregate material and binder have been mixed, the resulting mixture is rammed, blown or otherwise formed to the desired shape or pattern, and then cured with the use of catalysts and/or heat to a solid, cured state.
Resin binders used in the production of foundry molds and cores are often cured at high temperatures to achieve the fast-curing cycles required in foundries. However, in recent years, resin binders have been developed which cure at low temperatures. These processes are preferred over high-temperature curing operations which have higher energy requirements and which often result in the production of undesirable fumes.
One group of processes which do not require heating in order to achieve curing of the resin binder are referred to as no-bake processes. In such processes, the binder components are coated on the aggregate material, such as sand, and the resulting mixture is rammed, blown or otherwise formed to the desired shape or pattern. Curing of the binder is achieved without heating.
One such no bake process employs an aqueous alkaline solution of a phenolic resole resin as the binder. In this process, the foundry sand is usually mixed with an ester hardener before the solution of resole resin is added to the mixture. The process is described in detail in U.S. Pat. No. 4,474,904 (Re No. 32 812) which is incorporated herein by reference in its entirety.
The ester cured process is superior to some of the earlier processes from an environmental standpoint. However, the initial tensile strengths of the cores made by this process tend to be somewhat lower than those prepared by other no bake processes.
In Japanese Patent Application No. 62-282743 the absolute amount of binder addition was increased, resulting in higher tensile strengths of ester cured resins. This was achieved by using a solution of a phenolformaldehyde resin in the organic ester as the hardening agent. Examples of both novolak and resole resins in the ester solution were given. However, this approach has its limitations. The novolak resins used dissolve only in gamabutyrolactone and are not soluble in other ester hardeners which are useful in many applications in alkaline phenolic resin binders. The resoles disclosed do not form stable solutions in a number of the ester hardeners.
We have now discovered that solutions of benzylic ether phenolic resole resins in ester solvents are excellent hardeners for use in the ester cured process. Higher initial tensile strengths of cores made using this binder system are achieved without increasing the absolute amount of binder addition. Furthermore, the benzylic ether phenolic resole resins form stable solutions in triacetin (glycerol triacetate), ethylene glycol diacetate and other ester hardeners in which ordinary resole resins are not stable.

Ester hardeners for phenolic resin binder systems


A visual method for enhancing search result navigation, comprising: obtaining a first search result from a search engine; clustering the first search result to get clustering information; calculating the correlations between the clustering information and the ranked list of the first search result, and performing visualization processing on the clustering information; and displaying the visual cluster hierarchy and the ranked list of the first search result in a joint manner based on the correlations. When a cluster is selected, further searching is performed and the search result is clustered again. Using the present invention, through combining a traditional ranked list of search results and the visual cluster hierarchy of these search results to display them in a joint manner, a convenient way is provided for the web user to find the potential correlations between the visual cluster hierarchy and the ranked list of the search results. Besides, through dynamically searching and clustering more search results again, the web user may get more detailed and more accurate search results easily.

With the wide application of the Internet, people can get lots of information from the web. However, due to the rapid growth of the web contents, it becomes more and more difficult for web users to find required information rapidly and accurately. Currently, web users mainly rely on search engines to find required information. Generally, the process in which a web user uses a search engine to find required information is as follows: the web user submits a query, which may be, for instance, a single keyword or a combination of keywords, to the search engine. Then, the search engine produces a ranked list of the search results based on the submitted query. The ranked list is returned and displayed on the browser used by the web user. The web user obtains the part of the search results of interest to him through viewing segments of the returned ranked list of the search results.
However, such a method of using a search engine to find information commonly has a problem that the search engine always returns too many search results. In this situation, since the screen size of a computer display is limited, it is impossible to display all the search results simultaneously to the web user. Thus, the web user usually needs to browse many Web pages to find the required information, resulting in low efficiency of the web user getting information. On the other hand, according to an investigation of web users, in most cases, a web user only looks at the first few web pages of the ranked list of search results. Thus, in fact, the search quality of a web user searching information is also very low.
In order to improve search quality, some methods for improving the browsability of search results have been proposed in recent years. Vivisimo Company proposed a solution in which the search results returned by a search engine are clustered and the clustering results are visually displayed together with the ranked list of the search results. Although this solution may provide a convenient way for web users to know the clustering of the search results, it only displays the clustering results and the ranked list of search results simultaneously but independently, without presenting their correlations clearly and visually to the web users. Besides, the solution only clusters and displays part of the search results (for instance, the first 210 search results); if the web user selects a clustering item in the clustering results, the search results contained in the clustering item will be displayed, but no more relevant search results can be produced, so the web user cannot get more information of interest.

Visual method and apparatus for enhancing search result navigation


A jaw crusher includes s a toggle beam having a piston rod arrangement seat, a toggle plate, a first toggle plate seat arranged between the toggle plate and a movable jaw, a second toggle plate seat arranged between the toggle plate and the toggle beam, and a positioning device having a movable piston rod arrangement received by the piston rod arrangement seat of the toggle beam. The jaw crusher further has a toggle plate squeezing device including a contraction member. The squeezing device has a first mounting member connected to the movable jaw, and a second mounting member connected to the piston rod arrangement, to exert a pressing force on the first toggle plate seat, the second toggle plate seat and the piston rod arrangement seat.

Jaw crushers are utilized in many applications for crushing hard material, such as pieces of rock, ore, etc. A jaw crusher has a movable jaw that cooperates with a stationary jaw. Between the jaws a crushing gap is formed. The size of the crushing gap is often adjustable by means of a hydraulic ram which is connected to the movable jaw via a toggle plate and a toggle beam. Adjustment of the movable jaw position may be carried out to compensate for wear of wear parts and/or to adjust the size of the crushed material. In order to secure that the toggle plate is seated in a proper manner a jaw crusher may be provided with an arrangement that exerts a tensioning force acting to tension the toggle plate against toggle plate seats.
U.S. Pat. No. 4,637,562 discloses a jaw crusher with a toggle plate tensioning device where a compression spring exerts a force acting to tension the toggle plate against its respective toggle seats. U.S. Pat. No. 4,637,562 also discloses a hydraulic mechanism for adjusting the position of the movable jaw.
However, with the design described in U.S. Pat. No. 4,637,562 it may be difficult to secure a proper positioning of the toggle plate due to backlash arising during a crusher operation. Especially, upon presence of un-crushable material in the crusher such backlash may arise.