Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.6.1.1 (adenylate cyclase)
 19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ca2+ is a powerful inhibitor (Ki is congruent to 16 muM) of basal and prostaglandin E1 (PGE1)-stimulated adenylate cyclase [ATP pyrophosphate-lyase (cyclizing); EC 4.6.1.1] activity in membranes obtained from homogenized human platelets. Ca2+ (but not the ionophore A23,187) decreased V(max) of the reaction without an effect on the Ks for ATP. Neither ATP nor PGE1 affected Ki for Ca2+. In intact platelets A23,187 induced Ca2+ influx and markedly inhibited PGE1-stimulated rise in adenosine 3':5'-cyclic monophosphate (cAMP) levels. Guanylate cyclase [GTP pyrophosphate-lyase (cyclizing); EC 4.6.1.2] activity was mainly found in the soluble fraction (greater than 90%). Both soluble and membrane bound enzymes were stimulated by Mn2+ and Ca2+ and inhibited by Zn2+. Adenylate and guanylate cyclase activity were both present in a membrane fraction cyclase activity were both present in a membrane fraction which contained Ca2+ activated ATPase activity, and accumulated Ca2+ from the medium in the presence of ATP and oxalate. Other evidence indicates that these membranes originated in large part from the dense tubular system of the platelets. It is proposed that concurrent inhibition of adenylate cyclase and stimulation of guanylate cyclase facilitates the direct initiating effect of Ca2+ on platelet secretion and aggregation.
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PMID:Interrelationships between Ca2+ and adenylate and guanylate cyclases in the control of platelet secretion and aggregation. 0 60

The purpose of this study was to determine whether the previously reported differences in adenylate cyclase activity between the sarcolemma of normal and dystrophic chick muscles are also found in the SR, to search for a possible relationship between the adenylate cyclase changes and the pathophysiology of dystrophy, and to investigate whether the findings can be extended to Duchenne human muscular dystrophy by studying the adenylate cyclase and ATPase activities of erythrocyte ghosts from DMD patients and carriers. Microsomes were separated by standard techniques from the pectoralis muscles of normal and dystrophic ckeckens of various ages. The microsomal yields were significantly larger in dystrophic muscles. Adenylate cyclase activities in dystrophic microsomes were higher than those in matched controls and increased with the progression of the disease. The ratio between the two rose from one at 2 weeks of age to nine at about 9--10 weeks. Kinetic analyses showed that the ks for MgATP2- was about 40 microM (at 3 mM Mg2+ and 0.3 mM Ca2+) both in normal and dystrophic microsomes, that calcium caused umcompetitive inhibition of the enzyme (Ki = 0.2 mM), that the effect of calcium was noncooperative (Hill coefficient, nH = 1), that calcium did not affect the cooperativity for MgATP2-, and that magnesium competitively removed the calcium inhibition and caused additional, cooperative stimulation of the enzymatic activity (ka = 1.5 mM; NH =2). The major difference between normal and dystrophic adenylate cyclase was a higher enzymatic velocity in the latter, suggesting a larger amount of enzyme. We investigated whether altered cAMP levels may effect calcium accumulation. Calcium uptake measured (in the presence of oxalate) at several ages revealed no difference between normal and dystrophic chickens. The extent of calcium binding was also similar, although the kd for Ca2+ was lower in dystrophic microsomes. Binding was enhanced in the presence of exogenous protein kinase, but the responses of normal and dystrophic tissues were similar. We concluded that the elevation of adenylate cyclase in dystrophy was not related to microsomal calcium accumultion. Ivestigation of the localization of microsomal adenylate cyclase supported this view. Separation of calcium-loaded microsomes on a discontinuous sucrose gradient into four fractions demonstrated that adenylate cyclase activity, measured in the presence of Lubrol-PX and EGTA, was inversely related to calcium-accumulating activity. Na+, K+-ATPase comigrated with adenylate cyclase. Highest specific activities were found in the lightest fraction. These observations were confirmed by histochemical studies. The reaction product from adenylate cyclase activity was present predominantly in the terminal cisternae of the SR. In the context of the literature, our findings suggest that the rises in adenylate cyclase and Na+, K+-ATPase in avian dystrophy are compensatory changes, elicited by a defect in ECC at the calcium release step...
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PMID:Adenylate cyclase in muscular dystrophy. 15 10

Sarcolemmal and sarcoplasmic reticulum membrane vesicle fractions were isolated from cardiac microsomes. Separation of sarcolemmal and sarcoplasmic reticulum membrane markers was documented by a combination of correlative assay and centrifugation techniques. To facilitate the separation, the crude microsomes were incubated in the presence of ATP, Ca2+, and oxalate to increase the density of the sarcoplasmic reticulum vesicles. After sucrose gradient centrifugation, the densest subfraction (sarcoplasmic reticulum) contained the highest (K+,Ca2+)-ATPase activity and virtually no (Na2+,K+)-ATPase activity, even when latent (Na+,K+)-ATPase activity was unmasked. In addition, the sarcoplasmic reticulum fraction contained no significant sialic acid, beta receptor binding activity, or adenylate cyclase activity. Sarcolemmal membrane fractions were of low buoyant density. Preparations most enriched in sarcolemmal vesicles contained the highest level of all the other parameters and only about 10% of the (K+,Ca2+)-ATPase activity of the sarcoplasmic reticulum fraction. The results suggest that (Na+,K+)-ATPase, sialic acid, beta-adrenergic receptors, and adenylate cyclase can be entirely accounted for by the sarcolemmal content of cardiac microsomes. Gel electrophoresis of the sarcolemmal and sarcoplasmic reticulum membrane fractions showed distinct bands. Membrane proteins exclusive to each of the fractions were also demonstrated by phosphorylation. Cyclic AMP stimulated phosphorylation by [gamma-32P]ATP of two proteins of apparent Mr = 20,000 and 7,000 that were concentrated in sarcoplasmic reticulum, but the stimulation was markedly dependent on the presence of added soluble cyclic AMP-dependent protein kinase. Cyclic AMP also stimulated phosphorylation of membrane proteins in sarcolemma, but this phosphorylation was mediated by an endogenous protein kinase activity. The apparent molecular weights of these phosphorylated proteins were 165,000, 90,000, 56,000, 24,000, and 11,000. The results suggest that sarcolemma may contain an integral enzyme complex, not present in sarcoplasmic reticulum, that contains beta-adrenergic receptors, adenylate cyclase, cyclic AMP-dependent protein kinase, and several substrates of the protein kinase.
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PMID:Separation of vesicles of cardiac sarcolemma from vesicles of cardiac sarcoplasmic reticulum. Comparative biochemical analysis of component activities. 21 77

Cell surface membrane fragments were isolated and purified by successive rate zonal and isopycnic centrifugation of calcium oxalate-loaded pigeon heart microsomes in sucrose density gradients. The most highly purified cell membrane fraction sediments at a buoyant density of 1.105 g/ml. Some of the membrane pieces are present as open fragments and leaky vesicles, while others form tightly sealed vesicles of both inside-in and inside-out membrane orientation. The pigeon heart cell membrane preparation exhibits high (Na+ + K+ + Mg2+)-ATPase and adenylate cyclase activities. Additional activity of these enzymes is uncovered by sodium dodecyl sulfate and alamethicin, respectively. Electron microscopic inspection of the cell surface membrane preparation revealed (a) a predominance of thick-walled vesicles with smooth surfaces on negative staining and (b) binding of concanavalin A to the bulk of isolated membrane pieces following their incubation with the lectin.
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PMID:Mass isolation of cell surface membrane fragments from pigeon heart. 22 Oct 21

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

A rat heart sarcolemmal preparation could be obtained in which both 5'-nucleotidase and adenylate cyclase were enriched approx. 9-fold by subjecting a homogenate to a discontinuous sucrose gradient, without the use of a high salt extraction. After incubation of this fraction with Mg[gamma-32P]ATP, the majority of 32P incorporated was present in 24 000- and 9000-dalton protein components. Only when a heart cytosol fraction or a purified cyclic AMP-dependent protein kinase was added, was enhancement of 32P-incorporaton found by addition of cyclic AMP. The 9000- and 24 000-dalton proteins appeared to be interconvertible. The degree of conversion could be affected by changing the temperature during solubilizaion of the membranes in SDS prior to electrophoresis. This suggested that the 24 000-dalton protein does not correspond to phospholamban, first identified by others in canine heart sarcoplasmic reticulum. Moreover, it could be excluded that the 24 000-dalton protein was derived from contaminating myofibrillar troponin I. When the sarcolemmal fraction was preincubated with Ca2+, Mg2+, ATP and oxalate, contaminating sarcoplasmic reticulum vesicles, loaded with calcium oxalate, settled to a greater density in the sucrose gradient. Membrane constituents other than those with enzymatic activity were monitored to confirm the separation between sarcolemmal and sarcoplasmic reticulum membranes: Coomassie blue staining material, sialic acid, cholesterol and phospholipid. The 24 000- and 9000-dalton proteins were equally distributed among the sarolemmal and sarcoplasmic reticulum fractions present in the sucrose gradient. However, the rate of 32P-incorporation in the presence of heart cytosol fraction was much slowr in the sarcoplasmic reticulum than in the sarcolemmal fraction.
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PMID:Phosphorylation of low molecular weight proteins in purified preparations of rat heart sarcolemma and sarcoplasmic reticulum. 625

Vesicles isolated from rat heart, particularly enriched in sarcolemma markers, were examined for their sidedness by investigation of side-specific interactions of modulators with the asymmetric (Na+ + K+)-ATPase and adenylate cyclase complex. The membrane preparation with the properties expected for inside-out vesicles showed the highest rate of ATP-driven Ca2+ transport. The Ca2+ pump was stimulated 1.7- and 2.1-fold by external Na+ and K+, respectively, the half-maximal activation occurring at 35 mM monovalent cation concentration. In vesicles loaded with Ca2+ by pump action in a medium containing 160 mM KCl, a slow spontaneous release of Ca2+ started after 2 min. The rate of this release could be dramatically increased by the addition of 40 mM NaCl to the external medium. In contrast, 40 mM KCl exerted no appreciable effect on vesicles loaded with Ca2+ in a medium containing 160 mM NaCl. Ca2+ movements were also studied in the absence of ATP and Mg2+. Vesicles containing an outwardly directed Na+ gradient showed the highest Ca2+ uptake activity. These findings suggested the operation of a Ca2+/Na+ antiporter in addition to the active Ca2+ pump in these sarcolemmal vesicles. A valinomycin-induced inward K+-diffusion potential stimulated the Na+-Ca2+ exchange, suggesting its electrogenic nature. If in the absence of ATP and Mg2+ the transmembrane Nai+/Nao+ gradient exceeded 160/15 mM concentrations, Ca2+ uptake could be stimulated by the addition of 5 mM oxalate, indicating Na+ gradient-induced Ca2+ uptake to be a translocation of Ca2+ to the lumen of the vesicle. A sarcoplasmic reticulum contamination, removed by further sucrose gradient fractionation, contained rather low Na+-Ca2+ exchange activity. This result suggests that the activity can be entirely accounted for by the sarcolemmal content of the cardiac membrane preparation.
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PMID:An electrogenic Na+/Ca2+ antiporter in addition to the Ca2+ pump in cardiac sarcolemma. 721 3

We investigated the effects of age and nafidrofuryl oxalate (Naftidrofuryl), a 5-HT2 antagonist, on neurotransmission and transduction systems in the gerbil hippocampus using quantitative autoradiography. [3H]Quinuclidinyl benzilate (QNB), [3H]cyclohexyl-adenosine (CHA), [3H]MK-801, and [3H]muscimol were used to label muscarinic acetylcholine, adenosine A1, N-methyl-D-aspartate (NMDA), and gamma-aminobutyric acid-A (GABAA) receptors, respectively. [3H]PN200-110 labeled L-type Ca2+ channels. [3H]Forskolin, [3H]cyclic adenosine monophosphate (cAMP), [3H]phorbol 12,13-dibutyrate (PDBu), and [3H]inositol 1,4,5-triphosphate (IP3) were used to label adenylate cyclase, cAMP-dependent protein kinase, protein kinase C (PKC), and IP3 receptors, respectively. Approximately 20% reductions in [3H]QNB, [3H]forskolin, and [3H]PDBu binding were observed in the hippocampus of 9-month-old gerbils in comparison with 5-week-old gerbils. Treatment with Naftidrofuryl (10 mg/kg, i.p., once a day for 7 days) ameliorated these reductions. No changes were found in [3H]CHA, [3H]MK-801, [3H]muscimol, [3H]PN200-110, [3H]cAMP, and [3H]IP3 binding. The results suggest that Naftidrofuryl may have beneficial effects on the age-related alterations in signal transmission and transduction systems in the brain. Because the acetylcholine system, adenylate cyclase, and PKC are considered to be involved in learning and memory processes, the result may have clinical implications.
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PMID:Effects of naftidrofuryl oxalate, a 5-HT2 antagonist, on neurotransmission and transduction systems in the gerbil hippocampus. 806 66