Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.3.14 (ATP synthase)
7,042 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Beef heart mitochondrial ATPase, in both the membrane-bound and isolated form, contains tightly bound ATP and ADP. Each mol of ATPase contains about 2.2 mol ATP and 1.3 mol ADP. 2. In the absence of ATPase activity, these nucleotides exchange only slowly with nucleotides in solution. The exchange rate is increased during coupled ATPase activity, but not when the ATPase is uncoupled. 3. Oligomycin and dicyclohexylcarbodiimide inhibit exchange of the bound nucleotides, as does the ATPase inhibitor protein, although in each case some residual exchange occurs. Aurovertin, although inhibiting phosphorylation, does not inhibit the exchange. This is discussed in terms of the reversibility of these inhibitors. 4. The stimulation of exchange seen during coupled ATPase activity requires energisation of the ATPase molecule. Using the exchange reaction as a probe of energisation, it is deduced that energy can be transferred between different ATPase molecules. 5. It is proposed that coupled ATPase activity and phosphorylation in submitochondrial particles involve the tight nucleotide binding sites and the (weak) ATPase site, while uncoupled ATPase activity involves only the weak site.
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PMID:Tightly bound nucleotides of the energy-transducing ATPase, and their role in oxidative phosphorylation. II. The beef heart mitochondrial system. 13 63

Spinach leaf mitochondrial F0F1 ATPase has been purified and is shown to consist of twelve polypeptides. Five of the polypeptides constitute the F1 part of the enzyme. The remaining polypeptides, with molecular masses of 28 kDa, 23 kDa, 18.5 kDa, 15 kDa, 10.5 kDa, 9.5 kDa and 8.5 kDa, belong to the F0 part of the enzyme. This is the first report concerning identification of the subunits of the plant mitochondrial F0. The identification of the components is achieved on the basis of the N-terminal amino acid sequence analysis and Western blot technique using monospecific antibodies against proteins characterized in other sources. The 28-kDa protein crossreacts with antibodies against the subunit of bovine heart ATPase with N-terminal Pro-Val-Pro- which corresponds to subunit F0b of Escherichia coli F0F1. Sequence analysis of the N-terminal 32 amino acids of the 23-kDa protein reveals that this protein is similar to mammalian oligomycin-sensitivity-conferring protein and corresponds to the F1 delta subunit of the chloroplast and E. coli ATPases. The 18.5-kDa protein crossreacts with antibodies against subunit 6 of the beef heart F0 and its N-terminal sequence of 14 amino acids shows a high degree of sequence similarity to the conserved regions at N-terminus of the ATPase subunits 6 from different sources. ATPase subunit 6 corresponds to subunit F0a of the E. coli enzyme. The 15-kDa protein and the 10.5-kDa protein crossreact with antibodies against F6 and the endogenous ATPase inhibitor protein of beef heart F0F1-ATPase, respectively. The 9.5-kDa protein is an N,N'-dicyclohexylcarbodiimide-binding protein corresponding to subunit F0c of the E. coli enzyme. The 8.5-kDa protein is of unknown identity. The isolated spinach mitochondrial F0F1 ATPase catalyzes oligomycin-sensitive ATPase activity of 3.5 mumol.mg-1.min-1. The enzyme catalyzes also hydrolysis of GTP (7.5 mumol.mg-1.min-1) and ITP (4.4 mumol.mg-1.min-1). Hydrolysis of ATP was stimulated fivefold in the presence of amphiphilic detergents, however the hydrolysis of other nucleotides could not be stimulated by these agents. These results show that the plant mitochondrial F0F1 ATPase complex differs in composition from the other mitochondrial, chloroplast and bacterial ATPases. The enzyme is, however, more closely related to the yeast mitochondrial ATPase and to the animal mitochondrial ATPase than to the chloroplast enzyme. The plant mitochondrial enzyme, however, exhibits catalytic properties which are characteristic for the chloroplast enzyme.
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PMID:Plant mitochondrial F0F1 ATP synthase. Identification of the individual subunits and properties of the purified spinach leaf mitochondrial ATP synthase. 131 68

The genes encoding body-wall-specific glyceraldehyde-3-phosphate dehydrogenase from Caenorhabditis briggsae were sequenced and compared to the homologous genes from Caenorhabditis elegans. The direct tandem organization of these genes, gpd-2 and gpd-3, and the size and location of the two introns in each gene are the same in C. elegans and C. briggsae. Primer-extension studies demonstrated that the two genes in C. briggsae are trans-splice differentially with the same splice leader (SL) RNAs as are observed in C. elegans. The gdp-2 gene is trans-spliced with SL1 while gdp-3 is trans-spliced with SL2. Significant sequence conservation was observed within the promoter regions of each species and may indicate those regions responsible for body-wall-muscle-specific gene expression and/or differential trans-splicing. Comparisons of the sequences suggest that the tandem repeat of the genes has been subjected to concerted evolution and that C. briggsae and C. elegans diverged much earlier than would be anticipated based on morphological similarities alone. Finally, an open reading frame found several hundred nucleotides upstream from gpd-2, in both species, appears to be homologous to the ATP synthase subunit, ATPase inhibitor protein, from bovine mitochondria.
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PMID:Conservation of gene organization and trans-splicing in the glyceraldehyde-3-phosphate dehydrogenase-encoding genes of Caenorhabditis briggsae. 144 20

The binding of ATP radiolabeled in the adenine ring or in the gamma- or alpha-phosphate to F1-ATPase in complex with the endogenous inhibitor protein was measured in bovine heart submitochondrial particles by filtration in Sephadex centrifuge columns or by Millipore filtration techniques. These particles had 0.44 +/- 0.05 nmol of F1 mg-1 as determined by the method of Ferguson et al. [(1976) Biochem. J. 153, 347]. By incubation of the particles with 50 microM ATP, and low magnesium concentrations (less than 0.1 microM MgATP), it was possible to observe that 3.5 mol of [gamma-32P]ATP was tightly bound per mole of F1 before the completion of one catalytic cycle. With [gamma-32P]ITP, only one tight binding site was detected. Half-maximal binding of adenine nucleotides took place with about 10 microM. All the bound radioactive nucleotides were released from the enzyme after a chase with cold ATP or ADP; 1.5 sites exchanged with a rate constant of 2.8 s-1 and 2 with a rate constant of 0.45 s-1. Only one of the tightly bound adenine nucleotides was released by 1 mM ITP; the rate constant was 3.2 s-1. It was also observed that two of the bound [gamma-32P]ATP were slowly hydrolyzed after removal of medium ATP; when the same experiment was repeated with [alpha-32P]ATP, all the label remained bound to F1, suggesting that ADP remained bound after completion of ATP hydrolysis. Particles in which the natural ATPase inhibitor protein had been released bound tightly only one adenine nucleotide per enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Binding of adenine nucleotides to the F1-inhibitor protein complex of bovine heart submitochondrial particles. 161 Aug 24

An oligomycin-sensitive F1F0-ATPase isolated from bovine heart mitochondria has been reconstituted into phospholipid vesicles and pumps protons. this preparation of F1F0-ATPase contains 14 different polypeptides that are resolved by polyacrylamide gel electrophoresis under denaturing conditions, and so it is more complex than bacterial and chloroplast enzymes, which have eight or nine different subunits. The 14 bovine subunits have been characterized by protein sequence analysis. They have been fractionated on polyacrylamide gels and transferred to poly(vinylidene difluoride) membranes, and N-terminal sequences have been determined in nine of them. By comparison with known sequences, eight of these have been identified as subunits beta, gamma, delta, and epsilon, which together with the alpha subunit form the F1 domain, as the b and c (or DCCD-reactive) subunits, both components of the membrane sector of the enzyme, and as the oligomycin sensitivity conferral protein (OSCP) and factor 6 (F6), both of which are required for attachment of F1 to the membrane sector. The sequence of the ninth, named subunit e, has been determined and is not related to any reported protein sequence. The N-terminal sequence of a tenth subunit, the membrane component A6L, could be determined after a mild acid treatment to remove an alpha-N-formyl group. Similar experiments with another membrane component, the a or ATPase-6 subunit, caused the protein to degrade, but the protein has been isolated from the enzyme complex and its position on gels has been unambiguously assigned. No N-terminal sequence could be derived from three other proteins. The largest of these is the alpha subunit, which previously has been shown to have pyrrolidonecarboxylic acid at the N terminus of the majority of its chains. The other two have been isolated from the enzyme complex; one of them is the membrane-associated protein, subunit d, which has an alpha-N-acetyl group, and the second, surprisingly, is the ATPase inhibitor protein. When it is isolated directly from mitochondrial membranes, the inhibitor protein has a frayed N terminus, with chains starting at residues 1, 2, and 3, but when it is isolated from the purified enzyme complex, its chains are not frayed and the N terminus is modified. Previously, the sequences at the N terminals of the alpha, beta, and delta subunits isolated from F1-ATPase had been shown to be frayed also, but in the F1F0 complex they each have unique N-terminal sequences.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Identification of the subunits of F1F0-ATPase from bovine heart mitochondria. 182 92

The topography of the subunits of the membrane sector F0 of the ATP synthase complex in the bovine mitochondrial inner membrane was studied with the help of subunit-specific antibodies raised to the F0 subunits b, d, 6, F6, A6L, OSCP (oligomycin-sensitivity-conferring protein), and N,N' -dicyclohexylcarbodiimide (DCCD)-binding proteolipid and to the ATPase inhibitor protein (IF1) as an internal control. Exposure of F0 subunits in inverted and right-side-out inner membranes was investigated by direct antibody binding as well as by susceptibility of these subunits to degradation by various proteases as monitored by gel electrophoresis of the membrane digests and immunoblotting with the subunit-specific antibodies. Results show that subunits b, d, F6, A6L (including its C-terminal end) and OSCP were exposed on the matrix side. Sufficient masses of these subunits to recognize antibodies or undergo proteolysis were not exposed on the cytosolic side. This was also the case for subunit 6 and the DCCD-binding proteolipid on either side of the inner membrane. Quantitative immunoblotting in which bound radio-activity from [125I]protein A was employed to estimate the concentration of an antigen in a sample allowed the determination of the stoichiometry of several F0 subunits and IF1 relative to F1-ATPase. Results showed that per mol of F1 there are in bovine heart mitochondria 1 mol each of d, OSCP, and IF1, and 2 mol each of b and F6. Subunit 6 and the DCCD-binding proteolipid could not be quantitated, because the former transferred poorly to nitrocellulose and the latter's antibody did not bind [125I]protein A.
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PMID:Mitochondrial ATP synthase complex. Membrane topography and stoichiometry of the F0 subunits. 183 Mar 6

Deterioration of rat liver mitochondrial function during cold preservation with UW solution was studied. Mitochondria were isolated from control liver (0-hr preservation), 24-hr preserved liver, and 48-hr preserved liver with UW solution at 4 degrees C. Respiration assay revealed that phosphorylation was damaged more rapidly than oxidation. Inside-out submitochondrial particles prepared from each sample by sonication in the presence of EDTA were subjected to ATPase assay. ATP hydrolyzing activity of H(+)-ATPase (ATP synthase), which plays a key role in phosphorylation in mitochondria, decreased markedly to 58% after 24-hr preservation. After 48-hr preservation, reduction to 40% of control was noted. When an intrinsic H(+)-ATPase inhibitor protein was removed from ESMP by Sephadex gel filtration, decrease of the ATPase activity was enhanced down to 49% and 29% of the control for 24-hr and 48-hr preserved liver, respectively. Molecular damage of the enzyme was confirmed by SDS-PAGE. Alpha subunit of the enzyme decreased time-dependently, and H(+)-ATPase molecules that lost alpha subunit seemed to lose their catalytic activity. Although the cause of the molecular damage of H(+)-ATPase is not clear yet, some mitochondrial protease(s) may be involved.
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PMID:Molecular damage to rat liver mitochondrial H(+)-ATPase during cold preservation with UW solution. 183 87

Phosphorylation of ribosomal protein S6 of mammals precedes activation of cell growth in numerous biological systems. We have cloned a cDNA for ribosomal protein S6 from T-47D human breast cancer cells by immunoscreening a lambda gt11 expression library with antibody raised against the mitochondrial Ca(2+)-binding ATPase inhibitor protein (CaBI) of bovine heart mitochondria (Yamada & Huzel: J Biol Chem 263: 11498-11503, 1988). Similar clones were obtained by the immunoscreening of a rat heart expression library. In agreement with others, the open reading frames of the cDNAs from the two species coded for the same amino acid sequence. No difference in S6 of the human neoplastic cells compared to that of non-neoplastic cells was found. However, common antigenic determinants in S6 and CaBI were indicated. Accordingly, S6 was purified from rat liver ribosomes and antiserum prepared. Immuno-dot blot and Western blot analyses showed high specific reactivity between S6, the cloned chimeric beta-galactosidase fusion protein from a cDNA clone, and CaBI with anti-S6 and anti-CaBI antibodies. The antibodies also showed a high degree of discrimination for S6 and CaBI. Neither interacted with the other ribosomal proteins nor with another ATPase inhibitor protein from bovine heart mitochondria. Neither interacted with the Ca(2+)-binding proteins, calmodulin, oncomodulin, Protein C, or Factor X. Prothrombin was weakly reactive with anti-CaBI but not with anti-S6. Thus, the results fulfill the specific criteria for the concept and operational definition of common protein epitopes in S6 and CaBI. However, neither prothrombin nor S6 fusion protein inhibited mitochondrial ATPase activity even at 20 times the concentrations at which CaBI gave 97% inhibition.
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PMID:Antigenic reactivity of ribosomal protein S6 and the calcium-binding ATPase inhibitor protein of mammalian mitochondria. 183 89

An endogenous ATPase inhibitor protein has been identified and isolated for the first time from plant mitochondria. The inhibitor protein was isolated from potato (Solanum tuberosum) tuber mitochondria and purified to homogeneity. The isolated inhibitor is a heat-stable, trypsin-sensitive, basic protein, with a molecular mass approximately 8.3 kDa. Amino acid analysis reveals a high content of glutamic acid, lysine and arginine and the absence of proline; threonine and leucine. The interaction of the inhibitor with F1-ATPase requires the presence of Mg2(+)-ATP in the incubation medium. The ATPase activity of isolated F1 is inhibited to 50% in the presence of 14 micrograms inhibitor/mg F1. A stoichiometry of 1.3 mol inhibitor/mol F1 for complete inhibition can be calculated from this value. The potato ATPase inhibitor is also a potent inhibitor of the ATPase activity of the isolated yeast F1. The inhibitor resembles the ATPase inhibitors of yeast and mammalian mitochondria, and does not seem to be related to the inhibitory peptide, epsilon subunit, of chloroplast ATPase.
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PMID:Evidence for an endogenous ATPase inhibitor protein in plant mitochondria. Purification and characterization. 213 39

Submitochondrial particles (A particles) and phosphorylating electron-transport particles (ETPH) were prepared from bovine heart mitochondria. The A particles either were supplemented with or were depleted of the mitochondrial calcium-binding ATPase inhibitor protein (CaBI). The CaBI-depleted A particles still retained the Pullman-Monroy ATPase inhibitor protein (PMI), and the other particles all contained both CaBI and PMI. ATP synthase and ATPase activities of the particles were measured in similar reaction mixtures by luminescence of firefly luciferin-luciferase. Succinate was the respiratory substrate, and the adenylate kinase inhibitor P1, P5-di(adenosine-5') pentaphosphate was obligatory. The ATP synthase activity of CaBI-depleted A particles was 30-40% of that of the A and ETPH particles, and its ATPase activity was 7-8 times greater. Reconstitution of the CaBI-depleted A particles with CaBI restored the original ATP synthase and ATPase activities. ATP synthase activity rose about 1.7-fold when A particles were supplemented with additional CaBI and ATPase activity dropped to 9% of the original. Varying Ca2+ levels had little or no effect on the ATP synthase and ATPase activities of the CaBI-depleted A particles. In contrast, ATP synthase activity of the other particles was decreased by as much as 70% at the optimal Ca2+ concentration of 1 microM, and the ATPase activity of the A and EPTH particles rose concomitantly by 7-8-fold. The ATP synthase and ATPase activities of all the particles in microM Ca2+ became like those of the CaBI-depleted A particles. These changes were reversible; normal activities were restored as Ca2+ concentrations were raised above 1 microM.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Calcium-binding ATPase inhibitor protein of bovine heart mitochondria. Role in ATP synthesis and effect of Ca2+. 269 14


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