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)

Phthalate esters have shown to stimulate the latent ATPase [EC 3.6.1.3] activity and induce mitochondrial swelling. Among the tested phthalate esters, di-n-butyl phthalate (DBP) exhibited the greatest activity, and the activity decreased progressively as the alkyl chain was lengthened or shortened. The DBP-stimulated ATPase was oligomycin-sensitive. The degree of stimulation of the ATPase was proportional to the extent of mitochondrial swelling induced by DBP in 0.1 M Tris-HCl (pH 7.2) containing 0.25 M sucrose. However, the swelling was dependent on the tonicity of the solution or the concentration of chloride ions, while the stimulation of ATPase was independent of these factors. Swelling was strongly induced by DBP at slightly acidic rather than neutral or alkaline pH. The pH-activity curve of DBP-stimulated ATPase was in inverse correlation with that of swelling, which showed a rather flat maximum at pH 8.0. When bovine serum albumin (BSA) was added to a solution containing mitochondria before addition of DBP, swelling was no longer caused by DBP, though the latent ATPase was stimulated to the same extent in the absence of added BSA.
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PMID:Effects of phthalate esters on the latent ATPase and swelling of mitochondria. 1 58

In the search for gastric ATPases that might be related to the mechanism of HCl secretion, an interesting and rather unique K+-stimulated ATPase has been discovered. This enzyme is isolated from oxyntic cells and has been associated with the apical plasma membrane and/or tubulovesicular system. Membrane vesicles containing the K+-stimulated ATPase transport H+ into the vesicular lumen under the appropriate conditions of ATP, Mg2+, and KCl. This process can be measured by pH electrode, binding of certain metachromatic dyes to "energized" sites, or accumulation ratios of substances with appropriate pK values. Vesicular interior can be acidified to pH 3.5 or below. At the present time, it is difficult to distinguish between an electrogenic H+ pump and an electroneutral H+/K+ exchange mechanism. A hypothetical scheme for the gastric H+ secretory mechanism is proposed which fits much of the data from studies on the K+-ATPase, vesicular transport, and intact gastric mucosa.
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PMID:Gastric adenosine triphosphatases: a review of their possible role in HCl secretion. 2 Mar 86

Fluorescent amines, 9-aminoacridine, acridine orange and quinacrine, were used as probes for a pH gradient (deltapH) across gastric microsomal vesicles. Analysis of probe uptake data indicates that 9-aminoacridine distributes across the membrane as a weak base in accordance with the deltapH. On the other hand, acridine orange and quinacrine show characteristics of binding to membrane sites in addition to the accumulation in response to deltapH. A discussion of the advantages and limitations of the probes is presented. Application of these probes to pig gastric microsomal vesicles indicates that that K+-stimulated ATPase is responsible for the transport of H+ into the vesicles and thus develops a deltapH across the membrane. The deltapH generated by the K+-ATPase has a definite requirement for internal K+. The proton gradient can be discharged slowly after ATP depletion or rapidly either by detergent disruption of the vesicles or by increasing their leakiness using both H+ and K+ ionophores. On the other hand, the sole use of the K+ ionophore, valinomycin, stimulates the ATP-induced formation of deltapH by increasing the availability of K+ to internal sites. This stimulation by valinomycin requires the presence of permeable anions like Cl-. Analysis of the Cl- requirement indicates that in the presence of valinomycin the net effect is the accumulation of HCl inside the gastric vesicles. With an external pH of 7.0, the ATP-generated deltapH was calculated to be from 4 to 4.5 pH units. The results are consistent with the hypothesis that the K+-stimulated ATPase drives a K+/H+ exchange across the gastric vesicles. Since other lines of evidence suggest that these gastric microsomes are derived from the tubulovesicular system of the oxyntic cell, the participation of the ATP-driven transport processes in gastric HCl secretion is of interest.
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PMID:A study of H+ transport in gastric microsomal vesicles using fluorescent probes. 2 82

The phospholipid and fatty acid composition and role of phospholipids in enzyme and transport function of gastric (H+ + K+)-ATPase vesicles was studied using phospholipase A2 (bee venom). The composition (%) was phosphatidyl-choline (PC) 33%; sphingomyelin (sph) 25%; phosphatidylethanolamine (PE) 22%; phosphatidylserine (PS) 11%; and phosphatidylinositol (PI) 8%. The fatty acid composition showed a high degree of unsaturation. In both fresh and lyophilized preparations, even with prolonged incubation, only 50% of phospholipids were hydrolyzed, but the amount of PE and PS disappearing was increased following lyophilization. There was a marked decrease in K+-ATPase activity (75%) but essentially no loss of the associated K+ p-nitrophenyl phosphatase was found. ATPase activity could be largely restored by various phospholipids (PE greater than PC greater than PS). There was also an increase in Mg2+-ATPase activity, partially reversed in fresh preparations by the addition of phospholipids (PE greater than PS greater than PC). Proton transport activity of the preparation was rapidly inhibited, initially due to a large increase in the HCl permeability of the preparation. Associated with these enzymatic and functional changes, the ATP-induced conformational changes, as indicated by circular dichroism spectra were inhibited.
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PMID:Effect of phospholipase A2 on purified gastric vesicles. 4 34

The binding of ADP to subfragment-1 was investigated by the gel filtration method. The amount of bound ADP was determined as a function of free ADP concentration. Linear Scatchard plots were obtained. The maximum binding number, 0.55 mole of ADP per 10(5) g of protein, and the dissociation constant, 1.6 x 10(-6) M, were obtained, using subfragment-1 prepared by tryptic digestion, in the presence of 0.083 M KCl-10 mM MgCl2-0.02 M Tris-HCl (pH 8), at 25 degrees. Similar maximum numbers, about 0.5 mole per 10(5) g of protein, were obtained with subfragment-1 prepared by chymotryptic digestion of myosin or papain digestion of myofibrils. The maximum number did not depend on the KCl concentration or the temperature, while the dissociation constant decreased on decreasing either the KCl concentration or the temperature. Adenylyl imidodiphosphate binding to subfragment-1 prepared by chymotryptic digestion was also measured by the gel filtration method. The maximum binding number, 0.41 mole per 10(5) g of subfragment-1, and the dissociation constant, less than 10(-7) M, were obtained in the presence of 0.7 M KCl-10 mM MgCl2-0.02 M Tris-HCl (pH 8), at 8 degrees. The difference absorbance at 288 nm of the difference absorption spectrum induced by ADP of subfragment-1 prepared by tryptic digestion was proportional to the amount of bound ADP. The steady-state ATPase rate of subfragment-1 prepared by tryptic digestion was inhibited competitively by ADP in the presence of MgCl2. The extent of the initial burst of ATPase [EC 3.6.1.3] decreased from 0.46 +/- 0.06 to 0.30 +/- 0.09 mole of Pi per 10(5) g of subfragment-1 on adding ADP to a level of 0.6 mM. Subfragment-1 prepared by tryptic digestion bound F-actin with a mole ratio of 1/0.96 of actin monomer. The binding was depressed by the addition of ADP. On the basis of these results, subfragment-1 preparations were assumed to be a half-and-half mixture of two kinds of protein, and properties of each protein are discussed.
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PMID:A study of the binding of adenosine diphosphate to myosin subfragment-1. 12 50

Membrane vesicles isolated from wild-type and dicyclohexylcarbodiimide-resistant strains of Escherichia coli exhibit identical respiration-dependent transport activities, and in both cases, this activity is abolished by extraction of the vesicles with 1.0 M guanidine-HCl. Transport activity of extracted wild-type vesicles is completely restored by exposing the vesicles to lipophilic or water-soluble carbodiimides, while transport activity of the mutant vesicles is not restored by exposure to lipophilic carbodiimides. Strikingly, however, complete reactivation of transport in mutant vesicles is observed with water-soluble carbodiimides. Similarly, the Ca2+, Mg2+-stimulated ATPase activity of wild-type vesicles is inhibited by both classes of carbodiimides, while the ATPase activity of mutant vesicles is inhibited by water-soluble carbodiimides, but resistant to inhibition by lipophilic carbodiimides. The carbodiimide-reactive component of the membraneous Ca2+, Mg2+-stimulated ATPase complex in wildtype vesicles is readily labeled with N,N'-dicyclohexyl[14C]-carbodiimide, while the analogous component in mutant vesicles is not reactive. Alternatively, when vesicles are treated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide [14C]methiodide, a water-soluble carbodiimide, the carbodiimide-reactive component is labeled to a similar degree in both preparations. The results suggest that the altered carbodiimide-reactive proteolipid in the dicyclohexylcarbodiimide-resistant mutant is specifically defective in its ability to react with lipophilic carbodiimides. In addition, these and other findings indicate that the increase in proton permeability observed on extraction of isolated membrane vesicles with chaotropic agents is due exclusively to an effect on the carbodiimide-reactive component of the Ca2+, Mg2+-stimulated ATPase complex.
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PMID:The role of the carbodiimide-reactive component of the adenosine-5'-triphosphatase complex in the proton permeability of Escherichia coli membrane vesicles. 13 63

Tropomyosin was found to undergo only limited digestion by trypsin at 0 degrees C and the two segments that accumulated amounted to two-thirds of the original protein. They are referred to as segments A and B. These segments were not resistant to trypsin digestion at 20 degrees C and at the latter temperature no large fragments remained as judged by disc gel electrophoresis. Segments A and B were separated from each other on the basis of solubility differences and were found to have molecular weights of 24600 and 21900 respectively. Each of the segments appeared to retain about 70-75% of the helical conformation as judged by circular dichroism at 20 degrees C. However, the segments did not show any of the inhibitory activity of the parent tropomyosin molecule when mixed with troponin in the Mg2+-actomyosin ATPase system. Amino acid analysis showed that the portion of tropomyosin that was digested by trypsin (EC 3.4.21.4) had a lower content of the helix stabilizing residues Glu and Leu and a higher content of the helix-destabilizing residues Arg and Lys. These differences indicate that the digested portion should be less stable in the helical conformation than the two trypsin-resistant segments. End group determinations along with the results of the amino acid analysis indicated that segment A was probably derived from the central one-third of tropomyosin and segment B from the C-terminal one-third. By the process of elimination the N-terminal third appears to have been more liable region that was digested by trypsin. The segments A and B were shown to differ in their stability to denaturation by guanidine-HCl and elevated temperature. All of these observations indicate that tropomyosin is not a uniform structure and is composed of regions of different stability.
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PMID:The structure and stability of trypsin-resistant segments from rabbit tropomyosin. 13 16

Studies conducted into the activity of adenosine triphosphatase (ATPase) in homogenate of several tissues of sheep and against the background of pH 7.5 (tris-HCl buffer) have shown highest enzyme activity to develop in renal cortex and cerebral cortex followed, in declining order of quotation, by liver, myocardium, and mucous membrane of small intestine. ATPase activities were studied also in the presence of pH-values between 7.2 and 8.95 (tris-HCl buffer) and between 8.6 and 11 (piperazine buffer), with the pH optimum of ATPase in the above tissues having been found to lie at approximately 9.0. Different concentrations of Mg ions were added, and maximum ATPase activity of 2 mMol ATP was obtained by adding 2 mMol Mg. Decline in ATPase activity should be expected in the case of hypomagnesaemia. Addition of different concentrations of sodium and potassium ions gave in most of the tissues tested maximum activity in response to 10 mMol potassium and 68 mMol sodium. Na-K ATPase could be inhibited by oubain particularly in cerebral and renal cortex.
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PMID:[Activities and properties of adenosine triphosphatase (ATPase) in homogenates of renal cortex, liver, myocardium and small-intestine mucosa in sheep]. 13 80

Specific [3H]ouabain binding to rat and guinea pig skeletal muscle (musculus soleus) was studied using a rapid centrifugation and a filtration method. Both assays gave identical results: the incubation of the cell membranes in 50 mM imidazole/HCl buffer pH 7.25 or 7.4 MgCl2, Pi caused a time dependent loss of (Na+ +K+)-ATPase activity indicating an alteration of the membrane preparation. Ouabain binding properties were changed concomitantly. If ouabain binding was allowed to proceed until equilibrium was reached (3 min in rat and 10 min in guinea pig) at 37 degrees C the data plotted according to Scatchard followed a straight line. The dissociation constants of the ouabain-receptor-complexes of the rat cell membrane preparation as calculated from the slope of the plot (KD = 134 nM) and from the ratio of the dissociation and association rate constants (KD = 175 nM) agreed within experimental error with that determined by Clausen and Hansen [(1974) Biochim. Biophys. Acta 345, 387-404] in intact soleus muscles (KD = 210 nM). If ouabain binding was allowed to proceed for a longer period, however, nonlinear Scatchard plots resulted with an identical maximal number of binding sites but inconstant and decreased affinity for the cardiac glycoside. Experimental evidence is presented that nonlinear Scatchard plots often obtained in hormone (drug)-receptor binding experiments may (among other things) be the result of damaged cell membrane particles in vitro.
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PMID:Ouabain-receptor interactions in (Na+ + K+)-ATPase preparations. A contribution to the problem of nonlinear Scatchard plots. 13 90

F-Actin (FA) and pyruvate kinase (PK) [EC 2.7.1.40] were immobilized on PAB-cellulose. HMM-Subfragment-1 (S-1) was applied to a column of immobilized FA and PK, and eluted with 1-1.5 muM ATP and 1 mM PEP in 50 mM KCl, 2 mM MgCl2, and 10 mM Tris-HCl at pH 7.8 and 4 degrees. The size of the initial burst of Pi liberation of S-1 applied to the column was 0.5 mole/mole S-1. The burst size of S-1 decreased with increase in the fraction number, and S-1 in later fractions showed a burst size of 0.1-0.3 mole/mole. On the other hand, the rate of the ATPase [EC 3.6.1.3] reaction in the steady state was almost independent of the burst size, and increased slightly with increase in the fraction number. The ATPase activity of S-1 with a burst size of less than 0.2 mole/mole was scarcely activated by FA. Usually, the dependence on the burst size of S-1 of its ATPase activity in the presence of FA was sigmoidal, and marked activation by FA was observed when the burst size was larger than 0.3-0.4 mole/mole. Similar results were obtained with S-1 fractions separated by the ultracentrifugation method described in our previous paper ((1976) J. Biochem. 79, 419-434).
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PMID:Structure and function of the two heads of the myosin molecule. II. Separation of the two fractions of subfragment-1 of myosin by affinity column chromatography on immobilized F-actin: direct evidence for acceleration by F-actin of the decomposition of the reactive enzyme-phosphate-ADP complex formed on head B of myosin. 13 78


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