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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 surface activity and enzymic properties of the factor F1, the catalytic moiety of Streptococcus faecalis H+-ATPase, has been studied at the air-water and phospholipid-water interfaces. F1 does not interact with the monolayer phospholipids, hence its adsorption on a biological membrane must be due mainly to its recognition of proteins of the hydrophobic complex. The dimensions of the F1 molecule at the air-water interface have been estimated. In the presence of Mg2+, base area is S = 1.8 . 10(4) A2, height h = 27 A. Bearing in mind the size of a globular subunit, it follows from the measurements that the major F1 subunits should all lie in the same plane. The ATPase activity of F1 at the interface is inversely proportional to the monolayer density. With low density monolayer, the specific ATPase activity is higher at the interface than in the bulk of the solution. Adsorption of F1 at the interface shifts the isoelectric point of tiscussed relative to the proton-active transport mechanism.
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PMID:A study of the surface-active properties of the Mg2+-activated ATPase from cytoplasmic membranes of Streptococcus faecalis. 3 96

A purification procedure described previously resulting in electrophoretically pure Bacillus subtilis ATP-dependent DNAse has now been modified by adding a fractionation stage with Polymin P to permit large-scale isolation of the enzyme. It has been found that the enzyme molecule (Mr = 300000) consists of two large subunits with Mr 155000 and 140000. The purified enzyme has three activities: (1) DNAse on linear single-stranded and double-stranded DNAs (2) DNA-unwinding and (3) ATPase. Circular DNAs were not affected by the enzyme. Study of the dependence of these activities on temperature, pH, and ATP and Mg2+ concentrations has revealed two different states of the enzyme. At low ATP concentrations and alkaline pH, it showed chiefly nuclease action, degrading considerable amounts of DNA to small fragments five residues long on average. At higher ATP concentrations and neutral pH (more physiological conditions) it predominantly unwound DNA. Simultaneously it cut preferentially one of the duplex strands to fragments more than 1000 residues in length. The results obtained suggest that the energy of the enzyme-cleaved ATP is mainly expended on unwinding rather than on degrading DNA molecules.
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PMID:Properties of Bacillus subtilis ATP-dependent deoxyribonuclease. 3 53

1. The cell-membrane ATP phosphohydrolase of vegetatively grown Clostridium pasteurianum was specifically Mg2+-dependent, but demonstrated significant activity with GTP, CTP and UTP. It displayed approximate Michaelis-Menten kinetics only in the presence of certain effectors (e.g. phosphoenolpyruvate, fructose 1,6-bis-phosphate) which decreased the Km for ATP (to below 2 mM) but also V, whilst extending to pH 5.8 the effective pH range of activity of the enzyme. 2. ATP phosphohydrolase activity of the membrane ATPase (BF0F1) was inhibited by N,N'-dicyclohexylcarbodiimide, butyricin 7423, Dio-9, 4-chloro-7-nitrobenzofurazan, efrapeptin, leucinostatin and quercetin, and to a lesser degree by aurovertin and citreoviridin. The enzyme was not inhibited by oligomycin, spegazzinine, tributyl tin, triethyl tin or venturicidin. The soluble ATPase (BF1) component differed in not being inhibited by N,N'-dicyclohexylcarbodiimide, butyricin 7423 or leucinostatin. 3. The ATPase (BF0F1) complex and its soluble (BF1) component were separately purified. 4. Dodecylsulphate/polyacrylamide gel electrophoresis separated only four polypeptide components in the purified ATPase (BF0F1), with approximate molecular weights (+/- 10%) as follows: subunit a, 65 500; subunit c, 57 500; subunit da, 43 000; subunit fa, 15 000. The soluble (BF1 component contained only the three polypeptide subunits a, c and da. These were present in the BF0F1 preparation in the ratio 2 : 1 : 2; the contribution of subunit fa could not satisfactorily be quantified. 5. Subunit a was identified as the component binding 4-chloro-7-nitrobenzofurazan and subunit fa as the component binding N,N'-dicyclohexylcarbodiimide. The ATP phosphohydrolase activity of the membrane ATPase was not activated by trypsin treatment and the ATPase (BF0F1) contained no trypsin-sensitive inhibitor protein subunit. 6. Purified ATPase (BF0F1) was incorporated into artificial proteoliposomes which demonstrated ATP-dependent enhancement of 8-anilinonaphthalene-1-sulphonate fluorescence and ATP-dependent proton influx. These reactions were abolished by proton conductors (e.g. carbonylcyanide m-chlorophenylhydrazone) by valinomycin in the presence of a high external concentration of K+, or by N,N'-dicyclohexylcarbodiimide, butyricin 7423, Dio-9, 4-chloro-7-nitrobenzofurazan or leucinostatin. Oligomycin, tributyl tin, triethyl tin and venturicidin were not inhibitory. 7. When stripped of the soluble BF1 component, such ATPase-proteoliposomes demonstrated nil ATP phosphohydrolase activity and did not display ATP-dependent enhancement of 8-anilino-naphthalene-1-sulphonate fluorescence or ATP-dependent protein influx. All of these activities were restored by incubation of the BF1-depleted proteoliposomes with a purified preparation of the soluble BF1 component.
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PMID:The proton-translocating adenosine triphosphatase of the obligately anaerobic bacterium Clostridium pasteurianum. 1. ATP phosphohydrolase activity. 3 58

Functionally inverted plasma membrane vesicles isolated from the eukaryotic microorganism Neurospora crassa catalyze Mg2+/ATP-dependent Ca2+ uptake. Inhibitors induced efflux studies and isotope-exchange experiments indicate that the Ca2+ is accumulated inside the vesicles against a concentration gradient of about 40-fold, and that the majority of the transported Ca2+ is present essentially in free solution. Comparisons of Mg2+/ATP-driven 45Ca2+ uptake and [14C]SCN-uptake with respect to the Mg2+/ATP concentration dependence, the effects of inhibitors, and the nucleotide and divalent cation specificities indicate that the energy for Ca2+ accumulation is derived from ATP hydrolysis catalyzed by the electrogenic plasma membrane ATPase. Energized Ca2+ uptake is stimulated by the permeant anion SCN- to a degree that varies reciprocally with the ability of this anion to dissipate the membrane potential, and is inhibited by K+ in the presence of nigericin. All of these data point to the conclusion that the active transport of Ca2+ across the Neurospora plasma membrane takes place via a Ca2+/H+ antiporter, which functions to pump Ca2+ out of the intact cell.
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PMID:Active transport of calcium in Neurospora plasma membrane vesicles. 4 Feb 23

Purified chondrocytic alkaline phosphatase (orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1) from bovine fetal epiphyseal cartilage hydrolyzes a variety of phosphate esters as well as ATP and inorganic pyrophosphate. Optimal activities for p-nitrophenyl phosphate, ATP and inorganic pyrophosphate are found at pH 10.5, 10.0 and 8.5, respectively. The latter two substrates exhibit substrate inhibition at high concentrations. p-Nitrophenyl phosphate demonstrates decreasing pH optima with decreasng substrate concentration. Heat inactivation studies indicate that both phosphorolytic and pyrophosphorolytic cleavage occur at the same site on the enzyme. Mg2+ (0.1-10.0 mM) and Mn2+ (0.01-0.1 mM) show a small stimulation of p-nitrophenyl phosphate-splitting activity at pH 10.5. Levamisole, Pi, CN-, Zn2+ and L-phenylalanine are all reversible inhibitors of the phosphomonoesterase activity. Pi is a competitive inhibitor with a Ki of 10.0 mM. Levamisole and Zn2+ are potent non-competitive inhibitors with inhibition constants of 0.05 and 0.04 mM, respectively. The chondrocytic alkaline phosphatase is inhibited irreversibly by Be2+, EDTA, EGTA, ethane-1-hydroxydiphosphonate, dichloromethane diphosphonate, L-cysteine, phenyl-methylsulfonyl fluoride, N-ethylmaleimide and iodoacetamide. NaCL, KCL and Na2SO4 at 0.5-1.0 M inhibit the enzyme. At pH 8.5, the cleavage of inorganic pyrophosphate (pyrophosphate phosphohydrolase, EC 3.6.1.1) by the chondrocytic enzyme is slightly enhanced by low levels of Mg2+ and depressed by concentrations higher than 1mM. Ca2+ show only inhibition. Similar effects of Mg2+ and Ca2+ on the associated ATPase (ATP phosphohydrolase, EC 3.1.6.3) activity were observed. Arrhenius studies using p-nitrophenyl phosphate and AMP as substrates have accounted for the ten-fold difference in V in terms of small differences in both the enthalpies and entropies of activation which are 700 cal/mol and 2.3 cal/degree per mol, respectively.
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PMID:Enzymatic characterization of the chondrocytic alkaline phosphatase isolated from bovine fetal epiphyseal cartilage. 4 Jun 3

Growth of Clostridium perfringens was inhibited by compounds which dissipate or prevent the formation of electrochemical proton gradients. Membrane vesicles prepared from this organism exhibited Mg2+-dependent adenosine triphosphatase (ATPase) activity sensitive to N,N'-dicyclohexylcarbodiimide. Mg2+-ATPase activity was optimal of 50 degrees C, but no discrete pH optimum was observed. Adenosine triphosphate-dependent quenching of the fluorescence of the weak base quinacrine by everted membrane vesicles suggested that the Mg2+-ATPase is a proton pump capable of generating an electrochemical proton gradient. Adenosine triphosphate-dependent transport of Ca2+ by everted vesicles was sensitive to uncouplers and inhibitors of the Mg2+-ATPase.
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PMID:Properties and function of the proton-translocating adenosine triphosphatase of Clostridium perfringens. 4 Sep 63

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

An adenosine triphosphate-stimulated deoxyribonuclease was purified to about 4200 fold from Bacillus cereus. The enzyme activity of the crude extract increased by a factor of about 5 after dialysis. One of the low molecular weight inhibitors of the crude extract was found to be inorganic phosphate. During enzyme purification two nucleases were identified. One of them was specific to denatured DNA and the other one degraded both denatured DNA and native DNA. The activity towards native DNA could be increased several times by ATP. Through all steps of purification the ATP-independent DNase always accompanied the ATP-dependent one and the ratio of their activity was found to be constant. The ATP-dependent DNase also possessed ATPase activity stimulated both by native and denatured DNA. The fact that ATPase was stimulated by DNA and went together with ATP-dependent DNase during purification suggests that these functions belong to the same enzyme complex. Maximal activity of ATPase had broader pH, Mg2+ and ATP concentration ranges than that of DNase. Cooperation of the two functions may be limited only to a narrow range of ATP concentration. Km for ATPase was 1.6x10-4 M ATP.
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PMID:An adenosine triphosphate dependent deoxyribonuclease with adenosine triphosphatase, activity from Bacillus cereus. 4 71

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

Homogenates of Tritrichomonas foetus exhibited a Mg2+-dependent adenosine triphosphatase (ATPase) activity, with a pH optimum in Tris buffers of 8.2 to 8.3. The activity was not sensitive to oxygen. At high concentrations, quercetin and 4-chloro-7-nitrobenzofurazan inhibited ATPase activity in the cytoplasmic extract by 20 and 70%, respectively, whereas oligomycin, venturicidin, triethyltin, leucinostatin, dibutylchloromethyltin chloride, spegazzinine, efrapeptin, citreoviridin and sodium azide had no effect and N,N'-dicyclohexylcarbodi-imide stimulated the activity somewhat. The activity was localized in a population of small cytoplasmic particles which also contained an acid phosphatase. There was no indication of an association of ATPase with hydrogenosomes. The ATPase activity (or activities) in this aerotolerant anaerobe is different from the ATPases characteristic of mitochondria or of anaerobic bacteria.
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PMID:Adenosine triphosphatase activity of Tritrichomonas foetus. 4 53


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