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)

Reversible gamma-PO3 transfer in ATP reactions can be recognized by exchange of 18O from the beta,gamma-bridge position to the beta-P-nonbridge positions: (see article). Such intramolecular exchange is less demanding for the detection of the bond cleavage than the usual ATP:ADP isotope exchange because it does not require dissociation of bound ADP from the intermediate complex. Acyl phosphate intermediates are indicated for the glutamine synthetase and carbamyl-P synthetase reactions by their extreme requirements for glutamate and bicarbonate, respectively, for positional oxygen exchange. No support is given for E-P or concerted mechanisms. No support is found for an active CO2 in the latter reaction, although this is not ruled out by the data. Positional isomerization in ATP occurs with lamellae from spinach chloroplast only in the light. When the ATP molecule interacts, it also undergoes complete exchange of the gamma-PO3 oxygen with water before it rejoins the pool of free ATP. The difference in rates of the two exchanges suggests that the torsional motion of ADP-beta-PO3 is greatly hindered on the enzyme. This may explain, by the argument of substrate activation, the rapid reversibility of the ATPase reaction on the enzyme.
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PMID:Enzyme reactions of ATP studied by positional isotope exchange. 3 5

Freshly isolated sarcoplasmic reticulum vesicles contain 0.05 mol of tightly bound ADP and 0.03 mol of tightly bound ATP per mol of Ca2+, Mg2+-ATPase (ATP phosphohydrolase, EC 3.6.1.3). These values were increased to 0.1-0.2 mol ADP and 0.2-0.3 mol ATP per mol of ATPase after incubation of vesicles in the presence of MgATP and Ca2+ at 25 degrees C and pH 7.0. Half-maximal enrichment of tightly bound nucleotides was obtained with 2.5 mM ATP and 0.32 microM free Ca2+. Uncoupling of calcium transport from ATPase activity by mild acidic conditions or with ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid at pH 7.0 decreased the ability of the membranes to be enriched with tightly bound nucleotides and also decreased the content of tightly bound nucleotides of previously enriched membranes. Tightly bound [3H]nucleotides could only be partially displaced by reincubation under enrichment conditions. Tightly bound nucleotides are associated with energized calcium translocation but do not appear to be directly involved in the catalytic cycle.
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PMID:Occurrence and role of tightly bound adenine nucleotides in sarcoplasmic reticulum of rabbit skeletal muscle. 4 Feb 28

1. Phenylglyoxal reacts rapidly with isolated myosin heads (subfragment 1) and induces two successive and distinguishable effects on their enzymic properties: first, a twofold activation of the Ca2+ and Mg2+-dependent ATPases with no effect onthe K+-ATPase followed by inhibition of the K+, Ca2+ and actin-activated Mg2+-ATPases. A specific protein-reagent reagent complex is formed during the second phase of the modification reaction (Ki approximately 5 x 10(-3) M). 2. ADP and ATP with or without cations provide efficient protection only against the loss of ATPase activities, suggesting that the second inhibitory process is occurring at or close to the active site. 3. On the basis of [14C]phenylglyoxal-labelling experiments and the composition of modified subfragment-1 derivatives, it is demonstrated that the sequential modification of two reactive arginyl residues is responsible for the observed activation-inhibition phenomena. Blocking of the first reactive residue produces a shift in the pH/activity curves related to the Ca2+ and Mg2+-dependent ATPases with an apparent activation effect. Modification of the second guanidino group does not destroy the affinity of the protein for the nucleotide substrates but does alter the nucleotide binding site as reflected in the inability of Mg2+. ATP to dissociate the modified subfragment-1--actin complex. It is concluded that electrostatic interactions between this positively charged group and the negatively charged ATP and ADP molecules may be critical for the hydrolytic efficiency of myosin heads. 4. After dissociation and separation of the polypeptide constituents of the protein in acetic acid medium, both labelled sites are found to reside in the heavy chain.
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PMID:Involvement of an arginyl residue in the catalytic activity of myosin heads. 4 10

The reactions of adenosine 14C-and gamma 32P-labelled ATP with isolated membranes from catecholamine storage vesicles of the bovine adrenal medulla were studied. In presence of Mg2+ about twice as much of 32P-radioactivity combined with the membrane as 14C-adenosine compounds at 31 degrees C and also at 0 degrees C, while in the absence of Mg2+ the amounts of 14C and 32P incorporated were similar for both substances. Autoradiography of the SDS-polyacrylamide gel after electrophoresis of the 32P-ATP-treated membrane protein showed two distinct zones corresponding to protein bands. Sonication released twice as much 32P-ATP as 14C-ATP from the space within the membrane particles indicating that at least half of the ATP present in space did not contain its original terminal phosphate group. About 40--45% of the 32P-radioactivity was incorporated in the membrane lipids, whereas only small amounts of 14C-radioactivity were extracted with lipids. About 1/3 of the incorporated 14C-radioactivity was not extractable with acids. The same amount remained in the 32P-ATP treated preparation acid-stably bound after extraction of the lipids and hus must be firmly bound ATP. When the reaction of the membrane preparation with labelled ATP was performed at 0 degrees C the fractions of the acid-stably bound 32P- and 14C-radioactivity increased. About 1 nmole/mg of protein (10--15%) of the bound 32P-radioactivity was exchangeable against unlabelled ATP, while only a very small fraction (less than 0.5 nmol/mg protein) of the 14C-radioactivity was exchanged against unlabelled ATP. Preincubation of the membrane particles with ATP-Mg2+ at 0 degrees C induced 30% inhibition of the ATPase activity and abolition of the net uptake of catecholamines. Different Km values obtained from initial velocity studies of ATPase activity and the overall-incorporation of 32P-radioactivity indicated that a direct correlation between these processes did not exist. Different strong inhibitory effects exerted by ADP on the ATPase activity and net uptake of catecholamine at the one hand and the overall 32P-and 14C-incorporation at the other hand supported that view. It is concluded that small fractions of the observed 32P-and 14C-incorporation can be involved in the ATP hydrolyzing reaction.
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PMID:Distribution and metabolic fate of adenosine nucleotides in the membrane of storage vesicles from bovine adrenal medulla. 4 49

As different structural states of the Na+-K+-ATPase (EC 3.6.1.3) may lead to a changed reactivity to antibodies, the influence of different ligands on the reaction between highly purified membrane-bound Na+-K+-ATPase and specific antibodies was investigated. The antigen antibody reaction was registered by measuring the antibody inhibition of Na+-K+-ATPase activity. It was found that Na+ decreased and K+ increased the antibody inhibition of the Na+-K+-ATPase activity of the membrane-bound enzyme if both Mg++ and ATP were present during the antigen antibody reaction. These effects were not observed if ATP was replaced by ADP or by the ATP analog adenylyl (beta-gamma-methylene) diphosphonate. If a solubilized enzyme preparation with the same specific activity was used the effects of Na+ or K+ which were demonstrated in the membrane-bound enzyme could not be detected. The study suggests that the Na+-K+-ATPase structure is altered by Na+ and K+, provided Mg++ and specifically the nucleotide ATP are also present. These structural changes are likely to occur during Na+-K+-transport and do not seem to be necessarily linked to the Na+, K+ and Mg++ stimulated ATP splitting of the enzyme.
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PMID:Immunological characterization of Na+ and K+ mediated structural states of rat kidney Na+-k+-ATPase. 8 Mar 2

We have recognized an experimental confluence between oxidative phosphorylation and chemical carcinogenesis and, therefore, became interested in the mitochondrial target of hydrazine, which is not only a potential environmental hazard as a carcinogen but is also a likely metabolite of many drugs. Hydrazine induced a Pi dependent transitory uncoupling of rat liver mitochondria when beta-hydroxybutyrate was the substrate. Uncoupling was inhibited by rutamycin; accordingly, the mitochondrial target for nucleophilic hydrazine is an electrophilic site, presumably involving activated Pi. The protective action of ATP2, ADP, PPi and Mg++ was attributed to a conformational change of the phosphorylating enzyme which participated in oxidative phosphorylation. In a mitochondrial system which included ATP gramicidin potassium ion and sulfate, hydrazine, acting as a large cation but not as a nucleophile, blocked mitochondrial swelling and the increment in ATPase activity associated with potassium ion. These data in conjunction with our previous reports dealing with other carcinogens and certain of their derivatives also contribute to an experimental confluence between oxidative phosphorylation and chemical carcinogenesis and are compatible with toxic effects of hydrazine on mitochondria observed previously by others.
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PMID:A requirement of Pi for the transitory uncoupling of rat liver mitochondria by hydrazine, when beta-hydroxybutyrate is the substrate. 8 99

As different structural states of the (Na+-K+)-ATPase (EC 3.6.1.3) may lead to a changed reactivity to antibodies, the influence of Na+, K+, Mg++, Pi and ATP on the reaction between highly purified (Na+-K+)-ATPase and antibodies directed against the membrane-bound enzyme was measured. The antigen antibody reaction was registered by measuring the antibody inhibition of (Na+-K+)-ATPase activity. In the membrane-bound but not in the solubilized enzyme four different degrees of antibody inhibition were obtained at equilibrium of the antigen antibody reaction if different combinations of Na+, K+, Mg++ and ATP were present during the incubation with the antibodies. Corresponding to the different degrees of inhibition, different rates of enzyme inhibition were measured. (a) The smallest degree of enzyme inhibition was obtained when (i) only Mg++, (ii) Mg++ and Na+ or (iii) Mg++ and K+ were present during the antigen antibody reaction. (b) The enzyme activity was inhibited more strongly if Na+, Mg++ and ATP were present together. (c) It was inhibited even more if only (i) Na+, (ii) K+, (iii) ATP or both (iv) ATP and Na+, (v) ATP and K+, (vi) ATP and Mg++, or if (vii) no ATP and activating ions were present. (d) The highest degree of antibody inhibition was obtained if Mg++, ATP and K+ were present together. In the presence of Mg++ plus ADP and in the presence of Mg++ plus the ATP analog adenylyl (beta-gamma-methylene) diphosphonate, Na+ and K+ did not influence the degree of antibody inhibition as they did in the presence of Mg++ plus ATP. It was further found that the degree of antibody inhibition in the presence of Mg++, ATP and K+ was affected by the sequence of which K+ and ATP were added to the enzyme prior to the addition of the antibodies. It is suggested that by antibody inhibition different conformations of the (Na+-K+)-ATPase could be detected. These conformations may possibly not occur in the solubilized enzyme and therefore do not seem to be necessarily linked to the intermediary steps of the ATP hydrolysis of the enzyme. The structural changes which are induced by Na+ and K+ in the presence of Mg++ plus ATP are proposed to occur during the Na+-K+ transport.
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PMID:Conformational changes of membrane-bound (Na+-K+)-ATPase as revealed by antibody inhibition. 8 16

The requirement of actual splitting of ATP for endocytosis in erythrocyte ghosts has been confirmed by use of the ATP analog, 5'-adenylylimidodiphosphate, (AMP-P(NH)P). This compound, in which the oxygen connecting the beta and gamma phosphorus atoms was replaced by an NH group, did not cause endocytosis nor was it a substrate for ATPase activity. AMP-P(NH)P was a competitive inhibitor both for the endocytosis and the Mg2+-ATPase activities. The K1 of AMP-P(NH)P for Mg2+ ATPase activity was 2.0 - 10-4 M and, while the Km of ATP for this activity was also 2.0 - 10-4 M indicating nearly identical affinities of ATP and AMP-P(NH)P for the active site. ADP, or ADP plus orthophosphate, did not cause endocytosis, showing that endocytosis was not due to binding of the products of ATP hydrolysis. Sodium or potassium ion or ouabain had no effect on endocytosis, which eliminated the possibility of involvement of the Na+, K+ ATPase in the endocytosis process. Calcium could not be substituted for magnesium; rather it inhibited endocytosis at the concentration of 1 - 10-3 M. EGTA relieved the inhibitory effect of Ca, which indicated that the binding of calcium to the membrane was reversible. These experimental results reaffirm the conclusion that ATP must be split to engender endocytosis under these conditions. Some characteristic parameters of the hemoglobin-free porcine erythrocyte ghosts were studied in order to characterize the system more adequately.
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PMID:Energy-dependent endocytosis in erythrocyte ghosts. IV. Effects of Ca2+, Na+ +K+, and 5'-adenylylimidodiphosphate. 12 70

1. The bound nucleotides of the beef-heart mitochondrial ATPase (F1) are lost during cold inactivation followed by (NH4)2SO4 precipitation. The release of tightly bound ATP parallels the loss of ATPase activity during this process. 2. During cold inactivation, the sedimentation coefficient (s20, w) of the ATPase first declines from 12.1 S to 9 S, then to 3.5 S. (NH4)2SO4 precipitation of the 9-S component also leads to dissociation into subunits with s20, w of 3.5 S. 3. The 9-S component still contains the bound nucleotides, which are removed when it dissociated into smaller subunits. 4. Reactivation of cold-inactivated ATPase by incubation at 30 degrees C is increased by the presence of 25% glycerol. ATP, however, does not have any clearcut effect on the degree of reactivation in the presence of glycerol. 5. ADP is an inhibitor of the reactivation, probably because it exchanges during reactivation for bound ATP giving rise to an inactive 12-S component. 6. The exchange of tightly bound nucleotides with added adenine nucleotides is more extensive and faster with cold-inactivated ATPase than with the native enzyme. During reactivation up to 1.6 moles of ATP and 1.0 mole ADP can exchange per mole enzyme. 7. Incubation with GTP, CTP or inorganic pyrophosphate induces an increased activity of the ATPase, which, however, soon declines in the presence of ATP. It also disappears on precipitation of GTP-treated enzyme with (NH4)2SO4.
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PMID:Nucleotide-binding properties of native and cold-treated mitochondrial ATPase. 12 64

N-Ethylmaleimide, at millimolar concentrations, irreversibily inhibits photophosphorylation and ATPase activity of photosynthetic membranes from Rhodopseudomonas capsulata. The inhibitory effect of N-ethylmaleimide is evident only the membranes are preincubated with the inhibitor in the light and in the absence of phosphorylation substrates. ADP and orthophosphate (or arsenate) exert a protective effect against the inhibition if they are present during the preillumination stage. The energization of the membrane by ATP hydrolysis, measured as ATP-induced quenching of 9-aminoacridine fluorescence, also is inhibited irreversibly by N-ethylmaleimide. Uncouplers protect the ATPase from inhibition by N-ethylmaleimide at concentrations at which they inhibit photophosphorylation. The ATPase, as measured either in the dark or in the light, is also inhibited by carbonylcyanide p-trifluoromethoxypenylhydrazone in parallel with photophosphorylation. These results are interpreted as evidence that the high-energy state of the membrane induces a conformational change of the ATPase, making it sensitive to attack by N-ethylmaleimide; this conformational change might be related to the active state of the ATPase.
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PMID:Energy transduction in photosynthetic bacteria. VII. Inhibition of the coupling ATPase by N-ethylmaleimide related to the energized state of the membrane. 12 65


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