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

Endosulfan (E) is an organochloric insecticide, which is quickly metabolized and eliminated from the body system. Toxic effects of E and its metabolites have been reported. The influence of E and its metabolites, viz. endosulfan sulfate (ES), endosulfan diol (ED) and endosulfan lactone (EL), has been examined on rat liver mitochondria in vitro. Endosulfan stimulated state-4 respiration at lower concentrations and inhibited it at higher ones, whereas state-3 respiration was inhibited at all the concentrations used, i.e. 5-100 micrograms/ml. A maximal 25-fold activation of latent Mg2+-ATPase was achieved at a concentration that caused maximal stimulation of state-4 respiration. Activities of the respiratory chain-linked enzymes were inhibited at levels which corresponded to the concentrations of endosulfan used in vitro. Both the respiratory control ratio (RCR) and the ADP:O ratio fell sharply at endosulfan concentrations above 10 micrograms/ml. ES and ED exerted similar effects on mitochondrial oxidation of beta-hydroxybutyrate, but at more than double the concentration of the parent compound, while EL proved least effective. The effects of the latter compound on mitochondrial enzyme activities were negligible. Our results suggest that endosulfan possesses dual properties, that of an uncoupler of oxidative phosphorylation and of an inhibitor of the electron transport chain, and that the in vivo cytotoxic/insecticidal effects of endosulfan and its metabolites might, therefore, be the consequence of impaired mitochondrial bioenergetics.
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PMID:Effects of endosulfan and its metabolites on rat liver mitochondrial respiration and enzyme activities in vitro. 614 50

Sarcoplasmic reticulum vesicles were preloaded with either 45Ca2+ or unlabeled Ca2+ X 45Ca2+ efflux and influx, together with phosphorylation of the membrane-bound Ca2+, Mg2+-ATPase, were determined in the presence of either ATP and ADP or acetyl phosphate. ATP induced 45Ca2+ efflux. This ATP-induced 45Ca2+ efflux depended on ADP, external Ca2+, and Mg2+. The Ca2+ concentration dependence of the efflux was quite similar to the Ca2+ concentration dependence of the ATP-induced 45Ca2+ influx and the enzyme phosphorylation. The rate of the efflux was proportional to the steady level of the phosphoenzyme. The affinity for free ADP in this efflux was extremely high, being in good agreement with the affinity for free ADP in the transphosphorylation from the phosphoenzyme to ADP. These results show that the ATP-induced 45Ca2+ efflux represents the Ca2+-Ca2+ exchange (between the external medium and the internal medium) mediated by the phosphoenzyme. In this exchange, Mg2+ was essential for the exposure of the bound Ca2+ of the phosphoenzyme to the internal medium. 45Ca2+ efflux and influx were also activated by acetyl phosphate. This acetyl phosphate-induced efflux required the external Ca2+. The Ca2+ concentration dependence of the efflux agreed closely with that of the enzyme phosphorylation by acetyl phosphate. Furthermore, the rate of the efflux was proportional to the steady level of the phosphoenzyme. These and other findings show that the acetyl phosphate-induced 45Ca2+ efflux represents the Ca2+-Ca2+ exchange mediated by the phosphoenzyme and further demonstrate the direct dissociation of Ca2+ from the Ca2+-bound phosphoenzyme to the external medium in this Ca2+-Ca2+ exchange. The rate of the acetyl phosphate-induced, phosphoenzyme-mediated Ca2+ efflux was much slower than that of the ATP-, ADP-induced, phosphoenzyme-mediated Ca2+ efflux. This is consistent with our previous conclusion that the Ca2+ binding site is partially occluded upon the phosphorylation of the enzyme.
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PMID:Ca2+-Ca2+ exchange catalyzed by the membrane-bound Ca2+, Mg2+-ATPase of sarcoplasmic reticulum vesicles. 614 90

Mg2+-ATPase activity was identified in the cytosol of human erythrocytes. A partial purification of this activity was achieved by an initial DEAE-Sephadex column chromatography, followed by gel filtration on Sephadex G-100 and then a second DEAE-Sephadex chromatography procedure. The enzyme appeared in the void volume of the Sephadex G-100 column and was retained on an Amicon XM100A ultrafiltration membrane. The molecular weight of the enzyme was estimated to be 113 000 from SD gels. The above purification protocol yielded an enzyme with an optimal pH between 7.6 and 8.2. The enzyme activity increased linearly between 30 and 44 degrees C. It was stable for several months at -20 degrees C. Magnesium was essential for activity, but the rate attainable with Mn2+ was at least as great as that due to Mg2+. No other divalent cation was able to substitute for Mg2+ or Mn2+. Neither low nor high Ca2+ concentrations significantly affected the enzymatic activity. Substrate specificity studies showed that ATP was the preferred substrate followed by CTP (46% of the rate produced by ATP). Hydrolysis of GTP, UTP, ITP and ADP was less than 10% of the rate seen with ATP. No phosphatase, pyrophosphatase, phosphodiesterase, hexokinase, phosphofructokinase or adenylate cyclase activity could be detected in this enzyme preparation. Calmodulin, which stimulates the (Ca2+ + Mg2+)-ATPase of the human erythrocyte membrane, failed to enhance the Mg2+-ATPase activity. Of considerable interest, the activity of this Mg2+-ATPase was enhanced approximately 5-fold by low concentrations of mercuric ion, p-hydroxymercuribenzoate and DTNB, but was much less sensitive to iodoacetamide.
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PMID:Partial purification and characterization of a novel Mg2+-dependent ATPase present in the cytosol from human erythrocytes. 615 Jul 30

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

Photophosphorylating activity of chloroplasts rapidly prepared from preilluminated spinach leaves was higher than the activity of chlorplasts from leaves kept in the dark. Higher Vmax values were obtained with the former when either ADP or Pi concentrations were varied. The rate of decay of the in vivo light-activated Mg2+-ATPase was highly dependent on temperature, increasing with it. At 0 degree C it was stable for 40 min or more. The decay at 25 degrees C was prevented by 5 mM ATP or 50 mM dithioerythritol while ADP or Pi did not affect it. Gramicidin or iodosobenzoate induced a very rapid decay even at 0 degree C. Coupling factor 1 with a manifest and stable Ca2+-ATPase activity was solubilized from chloroplasts activated by light in vivo. Incubation of chloroplasts from preilluminated leaves with N-[3H]ethylmaleimide resulted in an inhibition of Ca2+-ATPase activity and in the incorporation of radioactivity into the gamma subunit of coupling factor 1 that was larger than that of chloroplasts from leaves kept in the dark. The results show that activation in vivo of the proton ATPase was manifested by higher phosphorylating and Mg2+-ATPase activities and requires both an electrochemical proton gradient and a redox change of at least one disulfide bond of its gamma subunit.
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PMID:Changes in activity and structure of the chloroplast proton ATPase induced by illumination of spinach leaves. 622 27

In isolated and purified cardiac myofibrillar and sarcolemmal preparations, the route of movement of ADP produced in the Mg2+-ATPase reactions was studied by investigating the efficiency of competition between the endogenous creatine kinase and exogenous pyruvate kinase reactions. In the homogeneous control system composed of hexokinase and glucose as ATPase, soluble creatine kinase rapidly rephosphorylated ADP produced in the presence of 1 mM ATP, but the addition of pyruvate kinase in an increasing amount inhibited the reaction of creatine release from phosphocreatine and symmetrically increased the rate of pyruvate production from phosphoenol pyruvate. At a pyruvate-kinase/creatine-kinase activity ratio (PK/CK) of 50, all ADP was used by the pyruvate kinase. In myofibrillar and sarcolemmal preparations containing particulate creatine kinase, the creatine kinase reaction was much less efficiently suppressed by pyruvate kinase, and at PK/CK = 50 half-maximal release of creatine was still observed. The rate of immediate myofibrillar MgADP rephosphorylation in the endogenous creatine-kinase reaction was observed to be governed by the concentration of phosphocreatine in accordance with the kinetics of this enzyme. The physiological significance of these findings is discussed.
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PMID:Creatine kinase in regulation of heart function and metabolism. I. Further evidence for compartmentation of adenine nucleotides in cardiac myofibrillar and sarcolemmal coupled ATPase-creatine kinase systems. 623 Oct 56

The mechanism of transport of basic amino acids into vacuoles of cells of the yeast Saccharomyces cerevisiae was investigated in vitro. Right-side-out vacuolar membrane vesicles were prepared from purified vacuoles. Arginine was taken up effectively by the vesicles only in the presence of ATP, not in the presence of ADP or AMP-adenosyl-5'-yl imidodiphosphate. It was exchangeable and was released completely by a protonophore, 3,5-di-tert-butyl-4-hydroxybenzilidenemalononitrile (SF6847). The transport required Mg2+ ion but was inhibited by Cu2+, Ca2+, or Zn2+ ions. The transport activity was sensitive to the ATPase inhibitor N,N'-dicyclohexylcarbodiimide (DCCD), but not to oligomycin or sodium vanadate. SF6847 or nigericin blocked arginine uptake completely, but valinomycin had no effect. ATP-dependent formation of a delta pH across the membrane vesicles was shown by quenching of 9-aminoacridine fluorescence. These results indicate that DCCD-sensitive, Mg2+-ATPase of vacuolar membranes is essential as an energy-donating system for the active transport, and that an electrochemical potential difference of protons is a driving force of this basic amino acid transport. Arginine transport showed saturation kinetics with a Km value of 0.6 mM and the mechanism was well explained by an H+/arginine antiport.
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PMID:Active transport of basic amino acids driven by a proton motive force in vacuolar membrane vesicles of Saccharomyces cerevisiae. 645 Jul 64

Actin binding to myosin-S1 modulates the limited tryptic cleavage of the COOH-terminal region of the 95K heavy chain at the joint connecting the 75K and 20K peptide units; concomitantly actin affords total protection against the resulting loss of acto-S1 Mg2+-ATPase activity. The specificity of the actin effect is illustrated by the fact that it exerts itself not only on free S1 but also on the intact myosin molecule. Mg2+-ATP and Mg2+-ADP impair the protective action of actin to an extent closely related to their respective affinity for the acto-S1 complex. Tryptic fragmentation of S1 heavy chain under highly controlled conditions, using trypsin to S1 weight ratios in the range 1:1000 - 1:1500 led us to establish that peptide bond cleavage at the 75K-20K junction is a sequential process giving rise first to a 22K peptide intermediate which is subsequently converted to the stable 20K fragment. Most importantly, it is also demonstrated that the loss of S1 activation by actin is not due to the initial scission of the 75K-22K linkage but is intimately associated with the breakdown of the 22K precursor into its 20K moiety. Three trypsin-modified S1 derivatives, the heavy chain of which is a complex of two or three fragments, were purified. A detailed analysis of the C-termini of these fragments, as compared to the C-terminal structure of the intact heavy chain, indicated that the 20K fragment is formed mainly through the degradation of a NH2-terminal 2K segment in the 22K precursor and that this proteolytic event is the only one accounting for the acto-S1 ATPase loss. Cross-linking experiments exploiting the reaction of a carbodiimide reagent with rigor complexes containing either fluorescent actin or fluorescent fragmented S1 revealed unequivocally the attachment of the actin monomer to recognition sites on the 20K and 50K units of S1 heavy chain. Specific interactions between the C-terminal 20K domain and light chain LC2 are proposed as being part of the molecular mechanism of the myosin-linked regulation of actomyosin interaction.
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PMID:Structural aspects of actomyosin interaction. 645 9

A vesicular fraction isolated from bovine aorta and enriched in fragmented sarcoplasmic reticulum (FSR) exhibited active calcium transport and ATPase activity. By use of a hypotonic NaHCO3 extraction solution, an active preparation was isolated that retained activity for up to 4 days. A small but significant (P less than 0.05) Ca2+-stimulated, Mg2+-dependent ATPase associated with calcium transport was demonstrated with a specific activity of 0.33 mumol inorganic phosphate (Pi).mg-1.min-1. The basal Mg2+ ATPase demonstrated Michaelis-Menten kinetics [Km(Mg2+-ATP) = 0.44 +/- 0.01 X 10(-3) M; Vmax = 2.22 +/- 0.01 mumolPi.mg-1.min-1]. The Ca2+-stimulated, Mg2+-ATPase demonstrated apparent substrate inhibition (Ks approximately 10 mM) with no evidence for end-product (ADP) or excess added Ca2+ contributing to this inhibition. Oxalate-supported active calcium uptake velocities also exhibited quantitatively similar substrate inhibition. These results suggest that FSR from vascular smooth muscle contains either two enzymes or one enzyme with two isomeric forms, one of which is associated with the calcium uptake activity of this structure and the other of unknown function.
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PMID:Influence of ATP on sarcoplasmic reticulum function of vascular smooth muscle. 646 Dec 58

Covalently cross-linked actin dimer was isolated from rabbit skeletal muscle F-actin reacted with phenylenebismaleimide (Knight, P., and Offer, G. (1978) Biochem. J. 175, 1023-1032). The UV spectrum of the purified cross-linked actin dimer, in a nonpolymerizing buffer, was very similar to that of native F-actin and not to the spectrum of G-actin. Cross-linked actin dimer polymerized to filaments that were indistinguishable in the electron microscope from F-actin made from native G-actin and that were similar to native F-actin in their ability to activate the Mg2+-ATPase of myosin subfragment-1. The critical concentrations of polymerization of cross-linked actin dimer in 0.5 mM and 2.0 mM MgCl2, 2 to 4 microM, and 1 to 2 microM, respectively, were similar to the values for native G-actin. Cross-linked actin dimer contained 2 mol of bound nucleotide/mol of dimer. One bound nucleotide exchanged with ATP in solution with a t 1/2 of 55 min and with ADP with a t 1/2 of 5 h. The second bound nucleotide exchanged much more slowly. The more rapidly exchangeable site contained 10 to 15% bound ADP.Pi and 85 to 90% bound ATP while the second site contained much less, if any, bound ADP.Pi. Cross-linked actin dimer had an ATPase activity in 0.5 mM MgCl2 that was 7 times greater than the ATPase activity of native G-actin and that was also stimulated by cytochalasin D. These data are discussed in relation to the possible role of ATP in actin polymerization and function with the speculation that the cross-linked actin dimer may serve simultaneously as a useful model for each of the two different ends of native F-actin.
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PMID:Isolation and characterization of covalently cross-linked actin dimer. 689 21


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