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)

The effects of the allosteric ligands UMP, IMP, and ornithine on the partial reactions catalyzed by Escherichia coli carbamyl phosphate synthetase have been examined. Both of these reactions, a HCO3(-)-dependent ATP synthesis reaction and a carbamyl phosphate-dependent ATP synthesis reaction, follow bimolecular ordered sequential kinetic mechanisms. In the ATPase reaction, MgATP binds before HCO3- as established previously for the overall reaction catalyzed by carbamyl phosphate synthetase [Raushel, F. M., Anderson, P. M., & Villafranca, J. J. (1978) Biochemistry 17, 5587-5591]. The initial velocity kinetics for the ATP synthesis reaction indicate that MgADP binds before carbamyl phosphate in an equilibrium ordered mechanism except in the presence of ornithine. Determination of true thermodynamic linked-function parameters describing the impact of allosteric ligands on the binding interactions of the first substrate to bind in an ordered mechanism requires experiments to be performed in which both substrates are varied even if only one is apparently affected by the allosteric ligands. In so doing, we have found that IMP has little effect on the overall reaction of either of these two partial reactions. UMP and ornithine, which have a pronounced effect on the apparent Km for MgATP in the overall reaction, both substantially change the thermodynamic dissociation constant for MgADP from the binary E-MgADP complex, Kia, in the ATP synthesis reaction, with UMP increasing Kia 15-fold and ornithine decreasing Kia by 18-fold. By contrast, only UMP substantially affects the Kia for MgATP in the ATPase reaction, increasing it by 5-fold.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Quantifying the allosteric properties of Escherichia coli carbamyl phosphate synthetase: determination of thermodynamic linked-function parameters in an ordered kinetic mechanism. 153 67

The uptake, retention and uptake antagonism of 201Tl-DDC, 201Tl-Cl, 123I-IMP, 99mTc-HMPAO and 99mTc-O4- were compared in rat neocortex cultures. 201Tl-DDC and 123I-IMP revealed the highest uptake of radioactivity in the cultures. 99mTc-HMPAO and 123I-IMP showed the highest retention of radioactivity within the tissue in washout experiments. Blocking of bioelectric activity by tetrodotoxin did not significantly affect the uptake of the radiopharmaceuticals (RPHA). Inhibition of Na-K-ATPase by ouabain inhibited the uptake of 201Tl-Cl (77%) and 201Tl-DDC (27%). Imipramine showed a significantly stronger inhibitory effect on 123I-IMP uptake in comparison with the effect on other RPHA. 99mTc-O4- was not concentrated within the cultured tissue. Under the in vitro conditions used in this study, the various RPHA were characterised by distinct differences in their interaction with cortical brain tissue.
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PMID:Uptake of SPECT radiopharmaceuticals in neocortical brain cultures. 253 15

A simple and fast ion pair reversed-phase high-performance liquid chromatographic method has been developed for the simultaneous determination of ATP, ADP, AMP, GTP, GDP, IMP, NADP+, NADPH+, NAD+, NADH, ADP-ribose, inosine, adenosine, hypoxanthine, and xanthine. This method allows us to have a complete picture of the most important nucleotides present in fresh human erythrocytes. Furthermore it is particularly useful in the study of the erythrocyte adenine nucleotide catabolism allowing the detection of degradation products such as IMP, inosine, adenosine, hypoxanthine, and xanthine. The separation of the compounds under investigation is achieved in less than 15 min using a reversed-phase 3-micron Supelcosil LC-18 column and adding tetrabutylammonium, as ion-pair agent, to the buffers. The short time of analysis, the high reproducibility of the system, and the accurate evaluation of the compounds of interest make this method particularly suitable for routine analysis. Finally it is possible to use this assay as an alternative method of measuring activities of enzymes which catalyze reactions involving some of these compounds, as in the case of Na+-K+ ATPase, AMP deaminase, and adenosine deaminase.
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PMID:A very fast ion-pair reversed-phase HPLC method for the separation of the most significant nucleotides and their degradation products in human red blood cells. 282 56

To investigate mechanisms whereby oxidant injury of cells results in cell dysfunction and death, cultured endothelial cells or P388D1 murine macrophage-like cells were exposed to oxidants including H2O2, O2-. (generated by the enzymatic oxidation of xanthine), or to stimulated polymorphonuclear leukocytes (PMN). Although Trypan Blue exclusion was not diminished before 30 min, cellular ATP was found to fall to less than 30% of control values within 3 min of exposure to 5 mM H2O2. Stimulated PMN plus P388D1 caused a 50% fall in cellular ATP levels. During the first minutes of oxidant injury, total adenylate content of cells fell by 85%. Cellular ADP increased 170%, AMP increased 900%, and an 83% loss of ATP was accompanied by a stoichiometric increase in IMP and inosine. Calculated energy charge [(ATP + 1/2 AMP)/(ATP + ADP + AMP)] fell from 0.95 to 0.66. Exposure of P388D1 to oligomycin plus 2-deoxyglucose (which inhibit oxidative and glycolytic generation of ATP, respectively) resulted in a rate of ATP fall similar to that induced by H2O2. In addition, nucleotide alterations induced by exposure to oligomycin plus 2-deoxyglucose were qualitatively similar to those induced by the oxidant. Loss of cell adenylates could not be explained by arrest of de novo purine synthesis or increased ATP consumption by the Na+-K+ ATPase or the mitochondrial F0-ATPase. These results indicate that H2O2 causes a rapid and profound fall in cellular ATP levels similar to that seen when ATP production is arrested by metabolic inhibitors.
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PMID:Alterations in adenosine triphosphate and energy charge in cultured endothelial and P388D1 cells after oxidant injury. 299 79

Intact synaptosomes isolated from the electric organ of the electric ray Torpedo marmorata contain, at their surface, enzyme activities for the hydrolysis of externally applied nucleoside phosphates. The diazonium salt of sulfanilic acid, as a low-molecular-weight, slowly permeating, covalent inhibitory agent, selectively blocks these enzyme activities and leaves intracellular lactate dehydrogenase intact. The ectoenzymes comprise both a nucleoside triphosphate and diphosphate phosphohydrolase, as well as a 5'-nucleotidase. Activity of nonspecific ectophosphatases is absent. The nucleoside triphosphatase hydrolyzes almost equally well ATP, GTP, CTP, UTP, and ITP and is activated to a similar degree by Mg2+ or Ca2+. It has a high affinity for ATP (Km for ATP in the presence of Mg2+, 75 microM; in the presence of Ca2+, 66 microM). Maximal rates in the presence of Mg2+ and Ca2+ were very similar (34.8 and 32.5 nmol of Pi/min/mg of synaptosomal protein, respectively). Either Mg-ATP or Ca-ATP can act as a true substrate. ADP inhibits hydrolysis of ATP, but AMP is without effect. The nucleoside triphosphatase is not inhibited significantly by a number of inhibitors of mitochondrial Mg2+-ATPase or of Ca2+ + Mg2+-ATPases. It is, however, considerably inhibited by filipin and quercitin. The capacity of intact synaptosomes to hydrolyze also extracellular ADP, GDP, AMP, GMP, and IMP suggests that the nucleoside triphosphatase is part of an enzyme chain that causes complete hydrolysis of the respective nucleoside triphosphate to the nucleoside. We conclude that the cholinergic nerve terminals of the Torpedo electric organ can hydrolyze ATP released on coexocytosis with acetylcholine via an ectonucleoside triphosphatase activity that is different from known endogenous nerve terminal ATPases. The final product of the hydrolysis, adenosine, can then be salvaged by the nerve terminal for resynthesis of ATP. Other possible physiological functions of the ectonucleotidases are discussed.
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PMID:Ectonucleotidase activities associated with cholinergic synaptosomes isolated from Torpedo electric organ. 301 88

Fatigue--or decrease in force generation--is a reduction of simultaneously attached cross-bridges in the force generating state. Two processes are necessary for the force generation: Firstly Ca++ release from the sarcoplasmic reticulum to the sarcoplasm and the binding of Ca++ by the troponin molecule and secondly the turnover of myosin-actin cross-bridges. These processes require energy in at least three different ATPase reactions and can consequently be inhibited when ATP hydrolysis is decreased, i.e. when ATP content is to low or when the reaction products (ADP, Pi and H+) reach inhibiting levels or when muscle pH has decreased to values inhibiting actomyosin ATPase activity (22). Low pH will also decrease Ca++ release and Ca++ affinity by troponin (23). In isometric contraction the force is well preserved as long as ADP phosphorylation can be provided by both PCr degradation and anaerobic glycolysis. When the PCr store is exhausted the force starts to decline and if muscle activation is maintained the force will continue to decrease along with falling glycolytic rate. ADP phosphorylation rate decreases successively and ATP content falls with an at least transient increase in ADP. The ATP decrease, apart from the minor increase in ADP, is balanced by an equimolar increase in IMP. Lactate accumulation produces an increasing acidity with muscle pH values down to 6.25. Early changes in free ADP content cannot be excluded as reason for the initial decrease in force production followed by more pronounced inhibition of ATPase activity during continued contraction due to both substrate lack and product inhibition together with pH effect on the excitation--contraction mechanism. In dynamic exercise with supramaximum work intensity the relation between fatigue development and metabolism is similar. In prolonged dynamic exercise relying on oxidative metabolism without lactate formation the point of fatigue is reached when the glycogen store is exhausted. Again ADP phosphorylation rate is decreased when the energy substrate is changed from carbohydrate to fat with lower maximum rate of ATP resynthesis.
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PMID:Biochemistry of muscle fatigue. 396 54

By using a reconstituted glycolytic system and a highly active adenosine triphosphatase (ATPase), the metabolism during muscular tetanic contraction was simulated and observed. With an ATPase activity somewhat greater than can be maintained in muscle tissue, phosphocreatine was rapidly and completely utilized, lactate production commenced about 5s after the ATPase was added and after 15s adenine nucleotides were lost through deamination to IMP. By 40s, all metabolism ceased because of complete loss of adenine mononucleotides. With a lower ATPase activity, glycolytic regeneration of ATP was capable of maintaining the ATP concentration at its initial value and even by 80s, only one-half of the phosphocreatine had been utilized. No deamination occurred in this time. It is suggested that the metabolic events observed in the simulated system are basically the same as occur in muscle doing heavy work.
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PMID:Studies with a reconstituted muscle glycolytic system. The anaerobic glycolytic response to simulated tetanic contraction. 427 6

Trinitrophenyl derivatives of adenine nucleotides (TNP-nucleotides: 2',3'-O-2,4,6-trinitrocyclohexadienylidene complexes at neutral or basic pH) are potent inhibitors of (Na,K)-ATPase activity. The inhibitory potency of the derivatives tested followed the sequence: TNP-ADP greater than TNP-ATP greater than TNP-AMP much greater than TNP-IMP greater than TNP-adenosine. In the presence of Na+ plus K+, high and low affinity activation of ATPase activity by ATP was observed. Under these conditions, TNP-ATP inhibited (Na,K)-ATPase activity competitively with respect to ATP at the kinetically defined "low affinity ATP site." In the presence of Na+ alone, only high affinity activation by ATP was observed. Under these conditions, TNP-ATP inhibited (Na)-ATPase and enzyme phosphorylation by competing with ATP at the kinetically defined "high affinity ATP site." The Ki values for inhibition were similar to the KD values determined by direct TNP-ATP binding measurements, indicating that the same TNP-ATP site is involved in the inhibition of (Na,K)-ATPase and (Na)-ATPase activities. We conclude that high and low affinity ATP "sites" are interconvertible (i.e. they represent two forms of the same site) and do not co-exist independently. TNP-ATP also inhibited competitively the K+-stimulated p-nitrophenyl phosphatase activity and enzyme phosphorylation by Pi, suggesting that the catalytic site for these substrates is associated with the TNP-ATP site. A kinetic model for (Na,K)-ATPase turnover based on a single ATP site which changes affinity during turnover is presented. The model was analyzed by the King-Altman (1956) J. Phys. Chem. 60, 1375-1378) method to obtain the steady state equation for the rate of ATP hydrolysis as a function of ATP concentration. Computer simulations using published values of the rate constants of intermediate steps suggest that the model is adequate to describe the observed dependence of enzyme activity on ATP concentration and the inhibition by TNP-ATP. The implications of these results on the structure and mechanism of the (Na,K) pump are discussed.
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PMID:Inhibition of sodium and potassium adenosine triphosphatase by 2',3'-O-(2,4,6-trinitrocyclohexadienylidene) adenine nucleotides. Implications for the structure and mechanism of the Na:K pump. 625 16

Mixed anhydrids of AMP, ADP, ATP and IMP and mesitylene carboxylic acid (AMP-MC, ADP-MC, ATP-MC and IMP-MC) are efficient irreversible inhibitors of the Ca-ATPase activity of myosin and heavy meromyosin. The highest rate of inhibition is observed in the case of AMP-MC: at AMP-MC concentration of 1,5.10(-3) M the half inactivation time for heavy meromyosin varies in different protein preparations from 10 to 20 min. The rates of inhibition in the presence of ADP-MC and ATP-MC are roughly the same and are far lower than those for AMP-MC (half inactivation time is 1,5-2 hrs). However, in the latter case the inhibition is complete, the time of the analogs interaction with the protein being increased up to several hours. In the presence of IMP-MC the inhibition is also time-dependent but is never complete. A necessary condition for the manifestation of irreversible inhibition of the Ca-ATPase activity of TMM by phosphorylating analogs of the substrate is the presence of bivalent cations. No inhibition occurs in the presence of EDTA. An addition of ADP or ATP to the preincubation medium causes a sharp decrease of the inhibition rate (a protective effect), which suggests a specific interaction of the analogs with TMM at the substrate binding site.
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PMID:[Inhibition of the Ca-ATPase activity of heavy meromyosin by phosphorylating analogs of the substrate]. 644 61

A (2'-5')An synthetase activity was isolated from human placental extracts by affinity chromatography on poly(rI) . poly(rC)-agarose. The oligonucleotide (2'-5')An was identified by (1) chromatography on PEI-cellulose and DEAE-cellulose, (2) inhibition of polypeptide synthesis in lysed rabbit reticulocytes (3) competition of the binding of pppA(pA)3,3'-[32P]pCp to rabbit reticulocyte lysates, and (4) alkaline phosphatase digestion. The synthetase activity in most placental preparations is activated by natural or synthetic dsRNA. However, in a few placental synthetase preparations, dsRNA is only marginally stimulatory and only becomes effective by prior treatment of the enzyme preparations with the calcium-dependent micrococcal nuclease. This suggests that there is an endogenous placental dsRNA contaminant in the enzyme preparations. In some synthetase preparations, a second dsRNA-stimulated product, tentatively identified as the nucleotide 5'-IMP, is also observed. Because the specific AMP deaminase inhibitor coformycin (10 microM) blocks the formation of IMP from ATP and causes a quantitative accumulation of AMP, and because the formation of IMp becomes independent of dsRNA when ADP or AMP is used in place of ATP, the presence of a dsRNA-stimulated ATP phosphohydrolase (ATPase) activity in human placenta is suggested.
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PMID:Double-stranded RNA-stimulated enzyme activities isolated from human placental extracts. 662 76


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