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)

The isolated Saccharomyces cerevisiae vacuolar proton-translocating ATPase (V-ATPase) is composed of at least 10 subunits. We have identified VMA8, the gene encoding the 32-kDa subunit of the V-ATPase, by 100% match between the sequences of tryptic peptides and the predicted protein sequence of ORF11. The VMA8 gene contains a 768-base pair open reading frame encoding a 256-amino acid protein with a predicted molecular mass of 29,176 Da. Disruption of VMA8 resulted in a mutant exhibiting pH-sensitive growth, slowed growth under all conditions, and an inability to grow on nonfermentable carbon sources. Vacuolar membranes isolated from vma8 delta yeast cells exhibited no V-ATPase activity. Immunoblot analysis of vma8 delta cells revealed normal levels of both V1 and Vo subunits. Whereas the V1 subunits failed to associated with the vacuolar membrane in vma8 delta cells, the Vo polypeptides were transported to and stable in the vacuolar membrane. Density gradient fractionation revealed that Vma8p associated only with the fully assembled V-ATPase and did not associate with a separate lower density Vo subcomplex fraction. Finally, Vma8p was unable to assemble onto the vascular membranes in the absence of other V1 subunits.
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PMID:VMA8 encodes a 32-kDa V1 subunit of the Saccharomyces cerevisiae vacuolar H(+)-ATPase required for function and assembly of the enzyme complex. 779 85

Subunit D of vacuolar H(+)-ATPase (V-ATPase) from bovine chromaffin granules was subjected to partial proteolysis and amino acid sequencing. A cDNA encoding this subunit was isolated and sequenced. The predicted open reading frame encodes a protein of 247 amino acids with a calculated molecular weight of 28,336. Northern blot analysis revealed an mRNA distribution with higher transcript amounts in tissues that are active in secretion. A homologous gene was identified as open reading frame 11 in chromosome V of Saccharomyces cerevisiae. The two proteins exhibit 55% identity with several conservative replacements. Interruption of the yeast gene, denoted as VMA8, resulted in the null mutant delta vma8::URA3 that, like all the other V-ATPase null mutants, did not grow on medium buffered at pH 7.5 and showed no accumulation of quinacrine into their vacuoles. Transformation of the null mutant with a plasmid containing the VMA8 gene restored the wild-type phenotype. This supports the conclusion that subunit D is an integral subunit of the catalytic sector of V-ATPase and its structural analysis suggests analogy to the gamma subunit of F-ATPases.
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PMID:A bovine cDNA and a yeast gene (VMA8) encoding the subunit D of the vacuolar H(+)-ATPase. 783 18

To investigate the function of subunit D in the vacuolar H(+)-ATPase (V-ATPase) complex, random and site-directed mutagenesis was performed on the VMA8 gene encoding subunit D in yeast. Mutants were selected for the inability to grow at pH 7.5 but the ability to grow at pH 5.5. Mutations leading to reduced levels of subunit D in whole cell lysates were excluded from the analysis. Seven mutants were isolated that resulted in pH-dependent growth but that contained nearly wild-type levels of subunit D and nearly normal assembly of the V-ATPase as assayed by subunit A levels associated with isolated vacuoles. Each of these mutants contained 2-3 amino acid substitutions and resulted in loss of 60-100% of proton transport and 58-93% of concanamycin-sensitive ATPase activity. To identify the mutations responsible for the observed effects on activity, 14 single amino acid substitutions and 3 double amino acid substitutions were constructed by site-directed mutagenesis and analyzed as described above. Six of the single mutations and all three of the double mutations led to significant (>30%) loss of activity, with the mutations having the greatest effects on activity clustering in the regions Val(71)-Gly(80) and Lys(209)-Met(221). In addition, both M221V and the double mutant V71D/E220V led to significant uncoupling of proton transport and ATPase activity, whereas the double mutant G80D/K209E actually showed increased coupling efficiency. Both a mutant showing reduced coupling and a mutant with only 6% of wild-type proton transport activity showed normal dissociation of the V-ATPase complex in vivo in response to glucose deprivation. These results suggest that subunit D plays an important role in coupling of proton transport and ATP hydrolysis and that only low rates of turnover of the enzyme are required to support in vivo dissociation.
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PMID:Subunit D (Vma8p) of the yeast vacuolar H+-ATPase plays a role in coupling of proton transport and ATP hydrolysis. 1080 66

In the accompany paper (Mukhopadhyay, A., Avramova, L. V. and Weiner, H., Arch. Biochem. Biophys.), it was shown that Tom34, a previously proposed putative translocase of the mitochondrial outer membrane, binds to the mature region of a precursor protein and appears to be a cytosol protein. Here Tom34 was used as bait in a yeast two-hybrid screening to search for its potential binding partners. Two of the identified proteins were the ATPase-related valosin-containing protein (VCP) and the lysosomal H(+)-transporting ATPase member M (ATP6M). Tom34 was found primarily in the cytosol while VCP and ATP6M were found in the cytosol as well as in nonmitochondrial organelles. Tom34 formed a approximately 400-kDa complex with them in the cytosol. Tom34 was found to possess a weak ATPase activity that did not change when associated with VCP. The tetratricopeptide repeat (TPR) motif region of Tom34 (residue 201-256) was responsible for binding to the other proteins. Tom34 appears not to be a member of the mitochondrial outer membrane translocase family but might function as a chaperone-like protein during protein translocation.
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PMID:Yeast two-hybrid screening identifies binding partners of human Tom34 that have ATPase activity and form a complex with Tom34 in the cytosol. 1191 76