Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

There are alterations in the proteins synthesized during different stages of development of Schistosoma mansoni. The protein profiles at different stages were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. When stained by Coomassie blue, no significant differences were seen in protein profiles derived from cultured schistosomula from Days 0 to 6 and from adults. Newly synthesized proteins were detected by [35S]methionine incorporation. There were only a few differences in the protein profiles of schistosomula from Days 1 to 6 and from adults. Profiles derived from Day 0 schistosomula showed striking differences. Only a few proteins appear to be synthesized on Day 0 under these conditions. Schistosomula on Day 0 synthesized several minor proteins as well as a major protein of approximately 69,000 Da. This protein was immunoprecipitated by rabbit antiserum against bovine uncoating adenosine triphosphatase which recognizes the constitutive and induced 70,000 Da heat shock proteins in a wide variety of eukaryotic cells. More significant differences were observed when the newly synthesized proteins were analyzed by two dimensional gel electrophoresis. The profiles of newly synthesized proteins showed a specific repertoire of expression during the early stages of development in the parasite. A shift in temperature and medium during transformation from aquarium water to isotonic medium may initiate the synthesis of heat shock protein in these parasites.
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PMID:Schistosoma mansoni: protein composition and synthesis during early development; evidence for early synthesis of heat shock proteins. 358 70

A cDNA clone complementary to an interferon (IFN)-induced mRNA approximately 3 kb in length was identified and sequenced revealing homology with the endoplasmic reticular heat shock protein/ATPase gp96. Both IFN-alpha and -gamma transcriptionally upregulate expression of this gene. gp96 transcripts, protein, and ATPase activity are shown to be enhanced as a result of IFN treatment in two human cell lines and this effect requires de novo protein synthesis. gp96 molecules have recently been implicated in the presentation of endogenous antigens. A number of the key elements in this pathway, the transporter proteins, the major histocompatibility complex (MHC)-linked units of the proteasomes and the MHC class I molecules are known to be IFN inducible. Our results show that yet another molecule suggested to play an accessory role in the endogenous presentation pathway is IFN inducible. Further, our studies represent the first demonstration of modulation of expression of a heat shock protein by a cytokine and identify a new enzymatic activity upregulated in IFN-treated cells.
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PMID:The endoplasmic reticular heat shock protein gp96 is transcriptionally upregulated in interferon-treated cells. 752 74

BiP is a member of the Hsp70 heat shock protein family found in the lumen of the endoplasmic reticulum, that binds to a variety of proteins destined to be secreted. Substance P (SP) has been used as a model peptide to study the interaction of BiP with protein substrates. SP stimulates BiP ATPase activity and forms a stable complex with BiP that is dissociated in the presence of levels of ATP > 50 microM. At lower concentrations of ATP, the SP remains bound to BiP, and the results are consistent with the view that a BiP-ATP complex is initially formed that reacts with SP to form a ternary complex, SP-BiP-ATP. Hydrolysis of ATP in this complex yields a SP-BiP-ADP complex. An exchange of ATP with ADP bound to BiP has also been demonstrated, and the results suggest that the interactions of BiP with ATP resemble those seen with GTP-binding proteins and GTP.
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PMID:Similarity of nucleotide interactions of BiP and GTP-binding proteins. 752 51

Many proteins in the cell require assistance from molecular chaperones at stages in their life cycles in order to attain correctly folded states and functional conformations during protein synthesis or during recovery from denatured states. A recently discovered molecular chaperone, which is abundant in the eukaryotic cytosol and is called the chaperonin containing TCP-1 (CCT), has been shown to assist the folding of some proteins in cytosol. This chaperone is a member of the chaperonin family which includes GroEL, 60-kDa heat shock protein (Hsp60), Rubisco subunit binding protein (RBP) and thermophilic factor 55 (TF55), but is distinct from the other members in several respects. Presently the most intriguing feature is the hetero-oligomeric nature of the CCT; at least eight subunit species which are encoded by independent and highly diverged genes are known. These genes are calculated to have diverged around the starting point of the eukaryotic lineage and they are maintained in all eukaryotes investigated, suggesting a specific function for each subunit species. The amino acid sequences of these subunits share approximately 30% identity and have some highly conserved motifs probably responsible for ATPase function, suggesting this function is common to all subunits. Thus, each subunit is thought to have both specific and common functions. These observations, in conjunction with biochemical and genetic analysis, suggest that CCT functions as a very complex machinery for protein folding in the eukaryotic cell and that its chaperone activity may be essential for the folding and assembly of various newly synthesized polypeptides. This complex behaviour of CCT may have evolved to cope with the folding and assembly of certain highly evolved proteins in eukaryotic cells.
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PMID:The chaperonin containing t-complex polypeptide 1 (TCP-1). Multisubunit machinery assisting in protein folding and assembly in the eukaryotic cytosol. 760 Nov 14

The heat shock protein GroEL from Escherichia coli is a tetradecameric oligomer that facilitates the refolding of nonnative polypeptides in an ATP-hydrolysis dependent reaction. A mutant in GroEL was prepared in which lysine 3 was substituted with glutamate, which destabilizes the oligomeric structure of GroEL (Horovitz, A., Bochkareva, E.S., and Girshovich, A.S. (1993) J. Biol. Chem. 268, 9957-9959). The highly expressed and purified GroELK3E was judged to be monomeric by sedimentation equilibrium, yielding a molecular weight of 54,500, despite a weak tendency of the mutant to reversibly form higher order aggregates above 4 mg ml-1. The monomeric variant appears to be folded based on the far UV circular dichroism spectrum, which shows significant alpha-helical content, but with slight differences in conformation relative to wild-type GroEL. The increase in exposed hydrophobic surface of the monomer was probed with the dye 4,4'-bis-1-anilino-3-naphthalenesulfonate (bis-ANS). The fluorescence of bis-ANS increases approximately 150-fold in the presence of the mutant, and about 4 mol of bis-ANS bind per mol of monomer, with a binding constant of 1.6 microM. Adenosine nucleotide binding to monomeric GroELK3E resulted in considerable quenching of bis-ANS fluorescence, correlating with significant structural changes as seen in the far UV circular dichroism, and permitted the measurement of binding isotherms for ATP and ADP. Hyperbolic ATP binding isotherms yield a dissociation constant of 82 microM, about 4-fold weaker than the K0.5 for ATP seen in steady-state kinetics assays of the wild-type GroEL ATPase.A similar difference was seen for ADP binding. These results suggest that the mutation disrupts the native tetradecameric quaternary structure through conformational changes that may also weaken nucleotide binding. The monomeric mutant exhibited no chaperone activity as evidenced by a filure to inhibit or facilitate the refolding of chemically denatured enolase, an inability to refold denatured rhodanese above spontaneous levels, and a lack of binding to alpha-casein, a competitor in many chaperonin-promoted refolding reactions. Thus, the formation of assembly incompetent monomers by the lysine 3 to glutamate mutation results in a dramatic decrease in the affinity for nonnative polypeptide chains and suggests that the oligomeric nature of GroEL is crucial for its molecular chaperone function.
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PMID:A monomeric variant of GroEL binds nucleotides but is inactive as a molecular chaperone. 765 15

To elucidate thermoresistance, a gene of a hyperthermophilic heat shock protein (HHSP) was isolated from the hyperthermophile Desulfurococcus strain SY which grows at 95 degrees C. The molecular weight of HHSP deduced from the open reading frame was 59,137 (545 amino acid residues). Sequence alignments of peptides reveal similarities (evolutionary distances) to the alpha (0.279) and beta (0.296) subunits of thermosome, TF55 (0.343) and human t-complex polypeptide 1. The structure of a thermophilic heat shock protein TGroEL (Tamada et al. (1991) Biochem, Biophys. Res. Commun. 179, 565) was quite different from that of HHSP. TGroEL and HSP60 have sequences identical to HHSP at its equatorial domain, while those identical to the alpha subunit of F-type ATPase are at its apical domain.
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PMID:Gene of heat shock protein of sulfur-dependent archaeal hyperthermophile Desulfurococcus. 767 88

The import of preproteins into mitochondria involves translocation of the polypeptide chains through putative channels in the outer and inner membranes. Preprotein-binding proteins are needed to drive the unidirectional translocation of the precursor polypeptides. Two of these preprotein-binding proteins are the peripheral inner membrane protein MIM44 and the matrix heat shock protein hsp70. We report here that MIM44 is mainly exposed on the matrix side, and a fraction of mt-hsp70 is reversibly bound to the inner membrane. Mt-hsp70 binds to MIM44 in a 1:1 ratio, suggesting that mt-hsp70 is localizing to the membrane via its interaction with MIM44. Formation of the complex requires a functional ATPase domain of mt-hsp70. Addition of Mg-ATP leads to dissociation of the complex. Overexpression of mt-hsp70 rescues the protein import defect of mutants in MIM44; conversely, overexpression of MIM44 rescues protein import defects of mt-hsp70 mutants. In addition, yeast strains with conditional mutations in both MIM44 and mt-hsp70 are barely viable, showing a synthetic growth defect compared to strains carrying single mutations. We propose that MIM44 and mt-hsp70 cooperate in translocation of preproteins. By binding to MIM44, mt-hsp70 is recruited at the protein import sites of the inner membrane, and preproteins arriving at MIM44 may be directly handed over to mt-hsp70.
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PMID:Mitochondrial protein import: biochemical and genetic evidence for interaction of matrix hsp70 and the inner membrane protein MIM44. 779 11

Mutants of CHO-K1 cells with varied levels of A system activity, probably the result of increases in absolute amount of the A system transporter, have corresponding increases in levels of peptides banding at 62-66 and 29 kDa. Mutant alar4-H3.9, showing the highest increase of A system activity and of 62- to 66- and 29-kDa peptides, was selected for this study. The N terminus 16-amino acid sequence of the 62- to 66-kDa peptide(s) of this mutant showed between 80% and 100% identity with the mammalian mitochondrial 60-kDa heat shock protein P1 (hsp60). Two-dimensional gel electrophoresis of the 62- to 66-kDa band showed two major, a minor, and several smaller spots (of same mass but different pI values) for both wild type (WT) and mutant, with the two major spots being of greater density in the mutant. Immunoblots with antibody to P1 identified the two major and minor peptides as P1 related. Two-dimensional gels of whole cell extracts of the WT and alar4-H3.9 confirmed these findings and indicated that the two major bands of the mutant were 2.4 times as abundant as that found for the WT. A plasma membrane fraction of the mutant, exhibiting 4.8 times more A system activity than the WT, contained 3.6 times as much P1 as the WT. Immunoblots with antibodies to P1, mitochondrial malate dehydrogenase, and to the mitochondrial F1/F0-ATPase demonstrated that the increased amount of P1 observed in the mutant was not the result of increases in amount of mitochondrial protein. Northern blot analysis demonstrated that the mutant had 2.5 times as much mRNA for P1 as the WT. The close analogy with the relationship between A system and Na+,K(+)-ATPase suggests that there is a coordinate regulation of the A system of amino acid transport, Na+,K(+)-ATPase, and P1 protein, probably as a result of mutation in a shared regulatory element. The possible role of P1 in A system function is discussed.
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PMID:Enhancement in amount of P1 (hsp60) in mutants of Chinese hamster ovary (CHO-K1) cells exhibiting increases in the A system of amino acid transport. 790 32

We have investigated the functional role of the coated vesicle-uncoating ATPase (UA), a cognate heat shock protein (hsc70), in receptor-mediated endocytosis. A monoclonal antibody against bovine brain UA/hsc70 was generated that recognizes a 26 kDa proteolytic fragment harbouring the putative clathrin-binding site. In vitro, this antibody blocked the UA/hsc70-mediated release of clathrin from isolated coated vesicles (CVs). Upon microinjection into tissue culture cells, it specifically inhibited the heat shock-induced nuclear migration of UA/hsc70. This antibody also interfered with endocytosis of ligand-receptor complexes in injected cells. Two different systems were studied: the uptake of aggregated human IgG by BHK cells transfected with a human Fc receptor (FcRII), and the internalization of LDL by human fibroblasts. Injection of the monoclonal antibody in concentrations yielding approximately equal molar ratios of antibody to enzyme resulted in a reduction of endocytosis to 20-30% of control values, as seen by conventional light and confocal laser scanning microscopy, and by electron microscopy. In the transfected BHK cells, the endocytosed ligand remained associated with the labeling for clathrin and was not delivered to the endosomal compartment within the period expected from control serum- or non-injected cells. Thin sections revealed an accumulation of coated structures in the antibody-injected cells as compared to controls. Thus, our data show that UA is essential for normal receptor-mediated endocytosis, and is presumably involved in the uncoating of CVs preceding their fusion with endosomes.
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PMID:Receptor-mediated endocytosis is sensitive to antibodies against the uncoating ATPase (hsc70). 792 28

A temperature-sensitive mutant of DnaK, the principal Escherichia coli member of the 70 kDa heat shock protein family, has been isolated. The mutation, dnaK25, lies in the putative ATP binding pocket of DnaK. It consists of a C to T transition that changes the highly conserved proline 143 to serine. Mutant strains do not support the propagation of bacteriophage lambda or of plasmids that require DnaA for replication. They are also defective in the utilization of mannose and sorbitol. ATPase activity of the mutant protein is reduced 20-fold relative to wild-type, while autophosphorylation is unaffected. DnaK25 has a fourfold faster rate of nucleotide exchange than wild-type DnaK; nucleotide exchange by both proteins is markedly increased by GrpE. The DnaK25 ATPase is still stimulated by DnaJ and GrpE and by peptide substrates. However, the affinity of most peptides tested for stimulating the DnaK25 ATPase is reduced significantly. These results indicate that a mutation in the N-terminal nucleotide binding domain can alter substrate interactions with the C-terminal substrate binding site. Nucleotide exchange by both wild-type DnaK and DnaK25 proceeds at a much faster rate than ATP hydrolysis, and therefore cannot be the rate limiting step of ATP hydrolysis under the conditions used in these experiments. Consistent with this, peptides, which stimulate ATP hydrolysis, have no effect on nucleotide exchange. Peptides thus appear to stimulate the ATPase by acting at another step, such as increasing the rate of phosphate bond cleavage.
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PMID:Isolation and characterization of an Escherichia coli DnaK mutant with impaired ATPase activity. 793 96


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