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Query: EC:3.6.3.1 (Mg2+-ATPase)
 1,484 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Most biological membranes are functionally asymmetric. To study biochemical control of cardiac transsarcolemmalion fluxes, it would be of obvious advantage to use isolated vesicles of sarcolemma which retains the low passive permeability characteristics of intact sarcolemma because in such vesicles the membrane should exhibit its normal asymmetric character with respect to enzymic activities. The purpose of this investigation was to attempt identify such vesicles in a cardiac microsomal (membrane vesicular) preparation. We studied activation by Na+ and K+ of Na+, K+-ATPase and its associated K+-phosphatase activities, using as substrates ATP or p-nitrophenylphosphate (pNPP) in the presence of Mg2+. Optimal concentrations of K+ alone (10 mM) stimulated p-nitrophenylphosphatase (pNPPase) activity 1.8-fold, and over 80% of the increase could be inhibited by ouabain. Optimal Na+ plus K+ concentrations (100 mM and 10 mM, respectively) stimulated the rate of ATP hydrolysis 2-fold, but only 11 +/- 1.1% of the increased activity was ouabain-sensitive. Optimal pretreatment with sodium dodecyl sulfate (SDS) (0.3 mg/ml) rendered both activities completely sensitive to inhibition by ouabain and reduced the basal Mg2+-ATPase activity by 70-90%. The K+-stimulated pNPPase activity doubled after preincubation in SDS, but the ATPase activity stimulated by Na+ plus K+ fell by 50% under these conditions. A similar pattern of apparent activation was produced by preincubation with deoxycholate (DOC), except that basal Mg2+-dependent activities were resistant to destruction by this detergent. The incremental responses to activation by ions and substrates, and inhibition by oubain, are consistent with the hypothesis that permeability-intact vesicles of sarcolemma are present in the isolated preparation, and that detergent activation renders the vesicles highly permeable to the ions, substrates, and ouabain.
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PMID:Intact vesicles of canine cardiac sarcolemma: evidence from vectorial properties of Na+, K+-ATPase. 18 13

An acid-stable phosphoprotein was formed in a microsomal membrane fraction isolated from bovine aortic smooth muscle in the presence of Mg2+ + ATP and Ca2+. The microsomes also showed Ca2+ uptake activity. The Ca2+ dependence of phosphoprotein formation and of Ca2+ uptake occurred over the same range of Ca2+ concentration (1-10 microM), and resembled similar findings from rabbit skeletal microsomes. The molecular weight of the phosphorylated protein, estimated by SDS-gel electrophoresis, was approximately 105,000. The phosphoprotein was labile at alkaline pH, and its decomposition was accelerated by hydroxylamine. Half-maximum incorporation of 32P in the presence of 10 microM Ca2+ occurred at 60 nM ATP. The calcium-dependent phosphoprotein formation was not affected by 5 mM NaN3, but was inhibited in a dose-dependent fashion by ADP with a 50% inhibition occurring at 180 microM. Fifty mM MgCl2 was required for the maximal phosphorylation. The rate of phosphoprotein decomposition after adding 2 mM EGTA was accelerated by varying the Mg2+ concentration from 10 microM to 3 mM. Alkaline pH (9.0) slowed the rate of phosphoprotein decay. Optimal Ca2+-dependent phosphoprotein occurred at 15 degrees C over a broad pH range (6.4 to 9.0). The activation energy of EGTA-induced phosphoprotein decomposition was 25.6 kcal/mol between 0 and 16 degrees C and 14.6 kcal/mol between 16 and 30 degrees C. The phosphoprotein formed by aortic microsomes was thus quite similar to the acid-stable phosphorylated intermediate of the Ca2+-transport ATPase of sarcoplasmic reticulum from skeletal and cardiac muscle. These data suggest that the Ca2+-dependent phosphoprotein is a reaction intermediate of the Ca2+,Mg2+-ATPase of the aortic microsomes.
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PMID:Ca2+,Mg2+-ATPase of microsomal membranes from bovine aortic smooth muscle. Identification and characterization of an acid-stable phosphorylated intermediate of the Ca2+,Mg2+-ATPase. 615 48