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:3.6.1.3 (ATPase)
65,361 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

1. Photophosphorylation was studied in spinach chloroplasts on illumination, from the dark state, with saturating short ("single turnover") flashes of light. 2. At rapid flash rates (100 Hz), phosphorylation began within the first five flashes. The ATPase inhibitor protein appeared to be displaced from its inhibitory site on the ATPase also within five flashes, as deduced from the flash-induced ATPase activity. 3. At slower flash rates, or if the rate of electron transfer were reduced with 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU), phosphorylation began only after a larger number (50--60) of flashes. The displacement of the ATPase inhibitior protein was similarly delayed. 4. Partial displacement of the inhibitor protein from its inhibitory site on the ATPase (by pretreatment with dithioerythritol) allowed phosphorylation to proceed without a perceptible lag, even in the presence of DCMU. It was concluded that the ATPase inhibitor protein must be displaced on the ATPase before phosphorylation can begin, and that this process is energy dependent. 5. During the flash regime used, release of inhibitor from its inhibitory site seemed to be governed largely by the membrane potential. The light-induced pH gradient seemed to have little effect under these conditions. Our results are not compatible with a direct conformational interaction between the electron transfer chain and the ATPase causing displacement of the inhibitor. 6. The maximal rate of photophosphorylation induced by less than 200 flashes was 0.12--0.15 mol ATP made/mol ATPase per flash. This rate seemed to be limited not be the supply of energy to the ATPase molecules, nor by the maximal turnover capacity of the ATP synthesising system, but by the number of ATPase molecules which were active in synthesis, i.e., which lacked the inhibitor protein. 7. The bound nucleotides of the coupling ATPase exchanged with added nucleotides during single turnover flashes. At high flash rates, exchange began within 5 flashes. The average amount of nucleotide exchanged per flash over 100 flashes was about one tenth the amount of ATP synthesised in each flash. 8. It is concluded that, during phosphorylation, a steady state level of active coupling ATPases is set up. The energy-dependent displacement of the inhibitor protein, and its (energy-independent) relaxation back on to the inhibitory site are the two opposing factors involved in this steady state.
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PMID:The initial stages of photophosphorylation. Studies using excitation by saturating, short flashes of light. 14 4

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

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

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

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

Conditions were selected which enable a quantitative assay of the ATPase inhibitor protein in submitochondrial particles. It was found that the isolated soluble inhibitor exhibits a marked pH-dependent hysteretic behaviour, i. e., an instant jump of pH for the inhibitor solution from 4.8 to 8.2 induced a slow alteration of its activity as measured by the inhibition of ATP hydrolysis by submitochondrial particles. In acid media (pH less than 6.8), the inhibitor is in the active, whereas in alkaline media (pH greater than 6.8) in the inactive state; the apparent pKa value for the cooperative active/inactive transition is 6.8. Treatment of the inhibitor protein with diethylpyrocarbonate, a specific reagent for histidine, completely abolishes its inhibitory activity. Two types of the inhibitor protein--ATPase interaction were revealed, i.e., reversible (ATP-independent) and irreversible (ATP-dependent) ones. Both reactions, i.e., ATP hydrolysis and ATP inhibition by the inhibitor in the presence of Mg2+ are characterized by a hyperbolic dependence of the reaction rate on ATP concentration; however, for both reactions the apparent KmATP values (50 and 5 microM, respectively) differ significantly (pH 8.0). Thus, the inhibitor--ATPase interaction shows that there exists a specific site for ATP in the ATPase which is different from the catalytic one. A model for the inhibitor protein interaction with ATPase which takes account of a slow pH-dependent conformational transformation of the inhibitor protein is proposed.
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PMID:[Kinetics of the interaction of ATPase of submitochondrial fragments and a natural protein-inhibitor]. 252 66

The urinary volume (U.V.), Na excretion (UNaV) and K excretion (UKV) have been reported to show a circadian rhythm in man, but the mechanism of this rhythm has not been made clear. To investigate how atrial natriuretic peptide (ANP) and endogenous digitalis-like substance (DLS) participate in the circadian change in urinary electrolyte, the circadian changes in ANP and DLS (digoxin-like immunoactivity: DLI, Na-K-ATPase inhibitor: ATPI, ouabain binding inhibitor to Na-K-ATPase: OBI) were evaluated in 5 normal man. ANP, DLI and OBI showed no significant correlation with urinary electrolyte excretion, but there was a significant positive correlation between plasma ATPI and urinary Na excretion. From these results it is suggested that circulating Na-K-ATPase inhibitor (plasma ATPI) may be involved in the regulation of the circadian rhythm of urinary Na excretion.
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PMID:The possible role of endogenous digitalis-like substance in the regulation of circadian changes in urinary electrolyte excretion in man. 256 Dec 76

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

Mitochondrial H+ -ATPase complex, purified by the lysolecithin extraction procedure, has been resolved into a "membrane" (NaBr-F0) and a "soluble" fraction by treatment with 3.5 M sodium bromide. The NaBr-F0 fraction is completely devoid of beta, delta, and epsilon subunits of the F, ATPase and largely devoid of alpha and gamma subunits of F1, where F0 is used to denote the membrane fraction and F1, coupling factor 1. This is confirmed by complete loss of ATPase and Pi-ATP exchange activities. The addition of F1 (400 micrograms X mg-1 F0) results in complete restoration of oligomycin sensitivity without any reduction in the F1-ATPase activity. Presumably, this is due to release of ATPase inhibitor protein from the F1-F0 complex consequent to sodium bromide extraction. Restoration of Pi-ATP exchange and H+ -pumping activities require coupling factor B in addition to F1-ATPase. The oligomycin-sensitive ATPase and 32Pi-ATP exchange activities in reconstituted F1-F0 have the same sensitivity to uncouplers and energy transfer inhibitors as in starting submitochondrial particles from the heavy layer of mitochondria and F1-F0 complex. The data suggest that the altered properties of NaBr-F0 observed in other laboratories are probably inherent to their F1-F0 preparations rather than to sodium bromide treatment itself. The H+ -ATPase (F1-F0) complex of all known prokaryotic (3, 8, 9, 10, 21, 32, 34) and eukaryotic (11, 26, 30, 33, 35-37) phosphorylating membranes contain two functionally and structurally distinct entities. The hydrophilic component F1, composed of five unlike subunits, shows ATPase activity that is cold labile as well as uncoupler- and oligomycin-insensitive. The membrane-bound hydrophobic component F0, having no energy-linked catalytic activity of its own, is indirectly assayed by its ability to regain oligomycin sensitive ATPase and Pi-ATP exchange activities on binding to F1-ATPase (33). The purest preparations of bovine heart mitochondrial F0 show seven or eight major components in polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate or SDS-PAGE (1, 2, 12, 14), ranging from 6 to 54 ku in molecular weight (12). The precise structure and polypeptide composition of mitochondrial F0 is not known. The F0 preparations from bovine heart reported so far have been derived from H+ -ATPase preparations isolated in the presence of cholate and deoxycholate (11, 33, 36, 37).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Resolution and reconstitution of H+ -ATPase complex from beef heart mitochondria. 285 48


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