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)

Anion dependence of (Ca2+ + K+)-stimulated Mg2+-dependent transport ATPase and its phosphorylated intermediate have been characterized in both "intact" and "broken" vesicles from endoplasmic reticulum of rat pancreatic acinar cells using adenosine 5'-[gamma-32P] triphosphate ([gamma-32P]ATP). In intact vesicles (Ca2+ + K+)-Mg2+-ATPase activity was higher in the presence of Cl- or Br- as compared to NO3-, SCN-, cyclamate-, SO4(2-) or SO3(2-). Incorporation of 32P from [gamma-32P]ATP into the 100-kDa intermediate of this Ca2+ATPase was also higher in the presence of Cl-, Br-, NO3- or SCN- as compared to cyclamate-, SO4(2-) or SO3(2-). When the membrane permeability barrier to anions was abolished by breaking vesicle membrane with the detergent Triton X-100 (0.015%) (Ca2+ + K+)-Mg2+ATPase activity in the presence of weakly permeant anions, such as SO4(2-) and cyclamate-, increased to the level obtained with Cl-. However, 32P incorporation into 100-kDa protein was still higher in the presence of Cl- as compared to cyclamate-, indicating a direct effect of Cl- on the Ca2+ATPase molecule. The anion transport blocker 4,4-diisothiocyanostilbene-2,2-disulfonate (DIDS) inhibited (Ca2+ + K+)-Mg2+ATPase activity to about 10% of the Cl- stimulation level, irrespective of the sort of anions present in both intact and broken vesicles. This indicates a direct effect of DIDS on (Ca2+ + K+)-Mg2+ATPase. K+ ionophore valinomycin influenced (Ca2+ + K+)-Mg2+ATPase activity according to the actual K+ gradient: Ko+ greater than Ki+ caused inhibition, Ko+ less than Ki+ caused stimulation. From these results we conclude that Ca2+ transport into endoplasmic reticulum is coupled to ion movements which must occur to maintain electroneutrality.
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PMID:Anion dependence of Ca2+ transport and (Ca2+ + K+)-stimulated Mg2+-dependent transport ATPase in rat pancreatic endoplasmic reticulum. 295 52

In a previous paper [Gould, East, Froud, McWhirter, Stefanova & Lee (1986) Biochem. J. 237, 217-227] we presented a kinetic model for the activity of the Ca2+ + Mg2+-activated ATPase of sarcoplasmic reticulum. Here we extend the model to account for the effects on ATPase activity of Mg2+, cations and anions. We find that Mg2+ concentrations in the millimolar range inhibit ATPase activity, which we attribute to competition between Mg2+ and MgATP for binding to the nucleotide-binding site on the E1 and E2 conformations of the ATPase and on the phosphorylated forms of the ATPase. Competition is also suggested between Mg2+ and MgADP for binding to the phosphorylated form of the ATPase. ATPase activity is increased by low concentrations of K+, Na+ and NH4+, but inhibited by higher concentrations. It is proposed that these effects follow from an increase in the rate of dephosphorylation but a decrease in the rate of the conformational transition E1'PCa2-E2'PCa2 with increasing cation concentration. Li+ and choline+ decrease ATPase activity. Anions also decrease ATPase activity, the effects of I- and SCN- being more marked than that of Cl-. These effects are attributed to binding at the nucleotide-binding site, with a decrease in binding affinity and an increase in 'off' rate constant for the nucleotide.
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PMID:Effects of Mg2+, anions and cations on the Ca2+ + Mg2+-activated ATPase of sarcoplasmic reticulum. 295 78

In order to study the mechanism of inhibition of gastric acid secretion by omeprazole, its action was investigated in several different in vitro preparations. In preparations from isolated gastric mucosa, isolated rabbit gastric glands and isolated parietal cells, omeprazole was found to inhibit both basal and stimulated acid secretion. These effects were seen irrespective of whether acid formation was stimulated by histamine or by db-cAMP. The inhibitory pattern of omeprazole was found to be of a non-competitive nature against db-cAMP stimulation. Furthermore, in isolated glands, omeprazole was found to inhibit stimulation induced by high medium K+ and low Na+ concentrations. The basal membrane of the intact gland preparation was made permeable to molecules of large size by the use of digitonin, and acid secretion was subsequently initiated by the addition of exogenous ATP. Even under these conditions, omeprazole was found to be inhibitory, with an IC50-value comparable to that of intact glands. SCN- was found to mimic the action of omeprazole in that it counteracted both basal and stimulated acid secretion in the test models described above. In contrast, cimetidine was found to inhibit only histamine stimulation, consonant with its H2-receptor-blocking properties. In the gastric gland preparation, changes in oxygen consumption is closely related to changes in acid formation. When oxygen consumption and acid formation were measured in parallel under histamine stimulation, another benzimidazole, timoprazole (H 83/69) (structurally related to omeprazole), was found to inhibit both parameters. However, under non-stimulated conditions, timoprazole was found to have only a minor effect on the oxygen consumption. The isolated H+K+ATPase preparation was used in order to investigate the effects of omeprazole at the "proton pump level". This enzyme was found to be inhibited by omeprazole in a pH-dependent manner. Under neutral or slightly alkaline conditions, slight inhibition occurred. When the pH of the incubation media was progressively decreased, the inhibitory activity of omeprazole was augmented. Several reactions of the H+K+ATPase enzyme cycle were investigated, i.e., K+-stimulated ATPase- and pNPPase- activities and formation of phosphoenzyme. All three of these reactions were inhibited. The results presented are in agreement with the hypothesis that omeprazole inhibits gastric acid secretion by blocking the gastric H+K+ATPase.
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PMID:The mechanism of action of omeprazole--a survey of its inhibitory actions in vitro. 298 9

Ethacrynic acid (EA) highly sensitive Mg2+-ATPase activity was demonstrated in rat brain microsomes. Marker enzyme studies suggested that the EA highly sensitive Mg2+-ATPase activity originated mainly from plasma membranes, and possibly from synaptic vesicles. Oligomycin did not affect the EA highly sensitive Mg2+-ATPase activity. Sulfhydryl reagents, such as N-ethylmaleimide and 5,5'-dithiobis-(2-nitrobenzoic acid), and anion transport inhibitors, such as 4-acetamide-4'-isothiocyanostilbene-2,2'-disulfonic acid, 4,4'-diisothiocyano-stilbene-2,2'-disulfonic acid and 2,4-dinitro-1-fluorobenzene, completely inhibited the EA highly sensitive Mg2+-ATPase activity with apparent Ki values at 5, 5, 8, 8 and 10 microM respectively. Treatment of microsomes with ethylenediaminetetraacetic acid and ammonium sulfate increased the EA highly sensitive Mg2+ and Na+,K+-ATPase activities, but not EA less sensitive Mg2+- or HCO3-ATPase activity, 2- to 3-fold that in crude microsomes. Relative substrate specificities of ATP much greater than GTP greater than ITP greater than UTP, CTP, a Km for ATP at 0.77 mM, and an optimal pH at pH 7.4 were observed. Among the anions tested (Cl-, Br-, F-, HCO3-, I-, SCN-, NO3-), EA highly sensitive Mg2+-ATPase activity was stimulated significantly by Cl- and reduced by NO3-. These data suggest that a novel, plasma membrane-located and anion-sensitive Mg2+-ATPase activity exists in the brain.
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PMID:Novel microsomal anion-sensitive Mg2+-ATPase activity in rat brain. 298 56

The Na,K-ATPase partially purified from porcine lens fiber cells (Sen and Pfeiffer, 1982) is stimulated fourfold (specific activity) by treatment with sodium thiocyanate. The optimum conditions are 1.5 M NaSCN, 2 mg protein ml-1 reaction mixture, pH 7.0, with incubation continued for 30 min at 23 degrees C. Sodium docecyl sulphate-gel electrophoresis and [3H]ouabain binding studies indicate that the extent of purity is not increased significantly by the procedure. The high-activity preparation has elevated phospholipid:protein and phosphatidylethanolamine:sphingomyelin ratios compared with the deoxycholate-extracted starting material. The cholesterol:phospholipid ratio and phospholipid acyl group composition are not significantly altered by SCN- treatment. Measurements of 1,6-diphenyl-1,3,5-hexatriene fluorescence polarization show that SNC- treatment produces approximately a 5 degrees C decrease in a membrane phase transition temperature. The phase transition also affects the activation energy of the Na,K-ATPase reaction and probably reflects the onset of the gel to liquid crystalline transition rather than the midpoint location of the transition per se. p-Nitrophenylphosphatase activity and Na,K-ATPase activity in the gel state membrane are also increased by SCN- treatment. Increased specific activity may result, in part, from a membrane fluidity-dependent enzyme activation but is also due, in part, to the expression of latent enzyme activity. Using ouabain-binding data and the specific activity of the activated preparation, it can be shown that the turnover number of the fiber cell enzyme is approximately 1% of that observed in most other tissues.
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PMID:Persistent stimulation of lens fiber cell Na,K-ATPase by sodium thiocyanate. 302 21

Inhibitors of the initial step (H2-antagonist) and of the final step (thiocyanate, SCN-; and nitrite, NO2-) were used to study the dynamics of acid secretion in isolated frog gastric mucosa. Tissues were mounted in flow-through chambers, and the acid secretion rate (SR) was recorded on a pH-stat microprocessor. Continuous presence of H2-antagonist decreases the SR to a lower steady state, and on removal the SR returns to basal SR, causing a net loss of acid, the nonconservative effect. The amount of lost acid is a unique function of exposure, thus, independent of the patterns (pulses or steps) of inhibition. In contrast, continuous presence of SCN- or NO2- (below 3 mM) results in an undershoot in SR with a return to basal SR, whereas at higher concentrations there is no return. Removal of these inhibitors causes an overshoot in SR with return to basal SR. The rebound acid is equal to acid suppressed by NO2- and low concentration of SCN-, resulting in no net loss of acid, the conservative effect, whereas at high concentrations of SCN- there is an apparent loss of acid. In maximally secreting tissue the overshoot of SR is not observed. However, the acid is not lost, merely delayed. In resting tissue NO2- also merely delays the exit of the acid produced in response to forskolin. The rebound acid is proposed to reside in a sequestered "acid" pool that is stable for at least 120 min. Results with NO2- and SCN- suggest an effect on a saturable exit enzyme, possibly the K+-H+-ATPase.
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PMID:Conservative and nonconservative inhibitors of gastric acid secretion. 311 14

The effect of NH4+ on synaptosomal membrane potential was examined using rat brains. The membrane potential was measured by the rhodamine 6G fluorescence method. Both NH4+ diffusion potential (NH4+-potential) and K+ diffusion potential (K+-potential) were observed in the synaptosomes. Upon replacement of medium Cl- with SCN-, both K+- and NH4+-potentials depolarized. On the other hand, replacement of medium Cl- with gluconate resulted in the hyperpolarization of K+-potential, but not of NH4+-potential. Ethacrynic acid (0.3 mM), a Cl- -ATPase inhibitor, depolarized both K+(Cl-)- and NH4+ (Cl-)-potentials. In the presence of ethacrynic acid, both of the potentials were further depolarized by replacement of medium Cl- with SCN-, but not with gluconate-. Picrotoxin (5 mM), a Cl- channel inhibitor, did not significantly affect either K+- or NH4+-potential. In the presence of picrotoxin, replacement of medium Cl- with SCN- depolarized both K+- and NH4+-potentials with or without ethacrynic acid. Gluconate depolarized the K+-potential with ethacrynic acid and the NH4+-potential with or without ethacrynic acid. These findings suggest that NH4+ forms a diffusion potential in nerve endings, and inhibits the anion-mediated hyperpolarization through mechanisms other than anion channels.
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PMID:[Effect of NH4+ on synaptosomal membrane potential in the rat brain]. 325 Sep 13

Epithelial brush-border membrane vesicles (BBMV) were made from lobster hepatopancreas by using Mg2+ precipitation. Alkaline phosphatase, Na+-K+-ATPase, and cytochrome c oxidase activities in these vesicles were enriched 15.0-, 1.0-, and 0.19-fold, respectively, compared with activities of a washed original homogenate pellet, indicating a relatively pure apical membrane preparation reduced in basolateral or organelle contamination. Complete vesicular closure was confirmed with electron microscopy and equilibrium [3H]D-glucose uptake experiments using various transmembrane osmotic gradients. Glucose uptake was stimulated by a transmembrane Na+ gradient but not by an identical K+ gradient or by a Na+ gradient in the presence of phloridzin. Electrogenicity of Na+-dependent glucose transport was confirmed in two ways. First, an anion permeability sequence indicated glucose uptake was stimulated in the following order: SCN- greater than Cl- greater than gluconate- greater than SO4(2-). Second, an outwardly directed valinomycin-induced K+ diffusion potential, rendering the vesicle interior electrically negative, enhanced glucose uptake compared with K+-loaded vesicles lacking the ionophore. Glucose influx occurred by a combination of carrier-mediated transfer, illustrating Michaelis-Menten kinetics, and nonsaturable "apparent diffusion." pH (same on both sides) strongly influenced Na+-dependent glucose uptake according to the sequence: pH 6.0 greater than pH 7.4 greater than pH 8.0. Increased proton concentration lowered the Michaelis constant for glucose transport and increased the apparent diffusional permeability of the membrane to the sugar. Maximal carrier-mediated glucose transport rate was largely unaffected by pH.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Glucose transport by lobster hepatopancreatic brush-border membrane vesicles. 397 Feb 30

ATP-dependent 45Ca2+ uptake was investigated in purified plasma membranes from rat pancreatic acinar cells. Plasma membranes were purified by four subsequent precipitations with MgCl2 and characterized by marker enzyme distribution. When compared to the total homogenate, typical marker enzymes for the plasma membrane, (Na+,K+)-ATPase, basal adenylate cyclase and CCK-OP-stimulated adenylate cyclase were enriched by 43-fold, 44-fold, and 45-fold, respectively. The marker for the rough endoplasmic reticulum was decreased by fourfold compared to the total homogenate. Comparing plasma membranes with rough endoplasmic reticulum, Ca2+ uptake was maximal with 10 and 2 mumol/liter free Ca2+, and half-maximal with 0.9 and 0.5 mumol/liter free Ca2+. It was maximal at 3 and 0.2 mmol/liter free Mg2+ concentration, at an ATP concentration of 5 and 1 mmol/liter, respectively, and at pH 7 for both preparations. When Mg2+ was replaced by Mn2+ or Zn2+ ATP-dependent Ca2+ uptake was 63 and 11%, respectively, in plasma membranes; in rough endoplasmic reticulum only Mn2+ could replace Mg2+ for Ca2+ uptake by 20%. Other divalent cations such as Ba2+ and Sr2+ could not replace Mg2+ in Ca2+ uptake. Ca2+ uptake into plasma membranes was not enhanced by oxalate in contrast to Ca2+ uptake in rough endoplasmic reticulum which was stimulated by 7.3-fold. Both plasma membranes and rough endoplasmic reticulum showed cation and anion dependencies of Ca2+ uptake. The sequence was K+ greater than Rb+ greater than Na+ greater than Li+ greater than choline+ in plasma membranes and Rb+ greater than or equal to K+ greater than or equal to Na+ greater than Li+ greater than choline+ for rough endoplasmic reticulum. The anion sequence was Cl greater than or equal to Br greater than or equal to 1 greater than SCN greater than NO3 greater than isethionate greater than cyclamate greater than gluconate greater than SO2(4) greater than or equal to glutarate and Cl- greater than Br greater than gluconate greater than SO2(4) greater than NO3 greater than 1 greater than cyclamate greater than or equal to SCN, respectively. Ca2+ uptake into plasma membranes appeared to be electrogenic since it was stimulated by an inside-negative K+ and SCN diffusion potential and inhibited by an inside-positive diffusion potential. Ca2+ uptake into rough endoplasmic reticulum was not affected by diffusion potentials. We assume that the Ca2+ transport mechanism in plasma membranes as characterized in this study represents the extrusion system for Ca2+ from the cell that might be involved in the regulation of the cytosolic Ca2+ level.
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PMID:Electrogenic calcium transport in plasma membrane of rat pancreatic acinar cells. 399 24

Anionic (NO(3) (-), Br(-), I(-), and SCN(-)) and cationic (Zn(++) and Cd(++)) potentiators of the twitch output of skeletal muscle depress the active binding of Ca by sarcoplasmic reticulum isolated from rabbit skeletal muscle. Zinc and Cd exchange for Ca and Mg at the binding sites of the reticular membranes, whereas the anions effectively induce a replacement by Mg of Ca bound actively in the presence of ATP. In the absence of ATP, the passive binding of both Ca and Mg is increased by the anions tested. Furthermore, the anions increase the total capacity of the membrane fragments for passive cation binding. The Ca-stimulated ATPase activity of the membranes is inhibited by Zn and Cd, but not by the anions. Shifts in cations bound to muscle membrane systems caused by agents that increase the force of contraction developed during the twitch are considered to be the primary event modifying excitation-contraction coupling, and thus leading to potentiation.
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PMID:Effects of potentiators of muscular contraction on binding of cations by sarcoplasmic reticulum. 423 Nov 48


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