Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A procedure has been developed for the separation of intrinsic proteins of plasma membranes from the electric organ of Torpedo marmorata. (Na+ + K+)-ATPase, nicotinic acetylcholine receptor and acetylcholinesterase remained active after solubilization with the nonionic detergent dodecyl octaethylene glycol monoether (C12E8). These components could be separated by ion exchange chromatography on DEAE-Sephadex A-25. Fractions enriched in ouabain-sensitive K+-phosphatase or (Na+ + K+)-ATPase activity showed two bands in sodium dodecyl sulphate polyacrylamide gel electrophoresis corresponding to the alpha- and beta-subunits. The (Na+ + K+)-ATPase was shown to have immunological determinants in common with a 93 kDa polypeptide which copurified with the nicotinic acetylcholine receptor, also after solubilization in Triton X-100 and chromatography on Naja naja siamensis alpha-toxin-Sepharose columns. The data suggest that the alpha-subunit of (Na+ + K+)-ATPase associates with the acetylcholine receptor in the membranes of the electric organ.
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PMID:Fractionation of protein components of plasma membranes from the electric organ of Torpedo marmorata. 629 54

The adenylate cyclase and Na+ -K+ ATPase activities decreased on storage at 4 degrees C as well as on freezing and thawing of the rat heart sarcolemma. Treatment of the sarcolemmal fraction with phospholipase C and trypsin also depressed the adenylate cyclase and Na+ -K+ ATPase activities; the Na+ -K+ ATPase was more sensitive to these treatments than the adenylate cyclase. When the sarcolemmal enzyme activities were determined in the presence of different concentrations of some cations the adenylate cyclase activity was enhanced and the Na+ -K+ ATPase activity was depressed by monovalent cations (Na+, K+, Rb+, Cs+, Li+, and NH+4). Divalent cations such as Sr2+, Ba2+, Co2+, and Mn2+ had biphasic or no effects on the adenylate cyclase activity but inhibited the Na+ -K+ ATPase activity. Although Ca2+, Ni2+, Cd2+, Cu2+, Hg2+, and Zn2+ depressed both Na+ -K+ ATPase and adenylate cyclase activities, the degree of inhibition of these enzymes was different. These results reveal the role of membrane integrity for full expression of the adenylate cyclase and Na+ -K+ ATPase activities, whereas both monovalent and divalent cations appear to regulate sarcolemma-bound enzyme activities.
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PMID:Role of membrane integrity and cation interaction for heart sarcolemmal adenylate cyclase and Na+-K+ ATPase. 630 75

1 The effect of (+/-)-, (+)- and (-)-verapamil on the Ca2+-binding, Ca2+-transporting activity, and Ca2+-dependent adenosine triphosphatase (ATPase) activity of isolated cardiac sarcolemmal preparations was studied. Enzymatic treatment was used to establish the nature of the sites facilitating [14C]-(+/-)-verapamil binding. 2 (+/-)-Verapamil 1 microM inhibited the passive binding of 45Ca2+. The (+/-)- and (-)-isomers were equiactive. 3 (+/-)-Verapamil 1 microM inhibited the ATP-dependent transport of 45Ca2+ and the associated activation of the Ca2+-sensitive ATPase. The activity resided in the (-)-isomer. 4 Lineweaver-Burk plots for the initial rates of ATP-dependent transport showed that the inhibition induced by the (-)-isomer was accompanied by a reduced Km and Vmax. 5 Enzymatic removal of N-acetyl neuraminic acid and galactose residues increased [14C]-(+/-)-verapamil binding; removal of N-acetylglucosamine and treatment with phospholipase C and trypsin decreased the binding. 6 These results have been interpreted to mean that (-)-verapamil interferes with the ATP-dependent Ca2+-transporting properties of the sarcolemma, and that this effect is accompanied by an altered activity of the intrinsic Ca2+-sensitive ATPase. N-acetylneuramic acid and galactose residues do not provide binding sites for verapamil at the cell surface.
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PMID:The effect of verapamil on the Ca2+-transporting and Ca2+-ATPase activity of isolated cardiac sarcolemmal preparations. 645 Dec 52

The role of phosphatidylinositol-specific phospholipase C (PIase C) in a) the enigmatic phosphatidylinositol (PI) turnover and b) in our understanding of membrane enzyme-PI interactions is the subject matter of this article. PIase C is present in both procaryotes and eukaryotes. This enzyme is considered to be involved in the cells PI breakdown which occurs in response to several external stimuli. Recent information on the physical properties, Ca2+ requirement, cellular localization and modulation of the activity of PIase C of mammalian systems can help to evaluate the PI turnover from a new angle. Existing evidence suggests that Ca2+-dependent PI breakdown is probably mediated through the cytosolic and particulate PIase C while a Ca2+ independent pathway is catalyzed by a lysosomal enzyme. Apparently PI turnover may be operating through more than one mechanism. The association of this phenomenon with a membrane receptor event linked with "Ca2+ gating" may have to be reconsidered. Modulation of the PIase C activity by unsaturated amphiphiles or the presence of this enzyme in different physico-chemical forms could be a potential regulatory feature. Hydrolysis of membrane PI of a number of cells and tissues by the bacterial PIase C has been shown to cause substantial release of acetylcholinesterase, alkaline phosphatase and 5'-nucleotidase in free, soluble form. Other membrane enzymes, e.g., alkaline phosphodiesterase I, L-leucyl-beta naphthyl amidase and Ca2+ or Mg2+ ATPase are not affected. These results indicate a specific interaction between PI and certain enzymes in membranes. The chemical nature of this linkage, whether it is covalent or non-covalent, has also been explored and has provided intriguing insight into this phenomenon. New findings also indicate that hydrolysis of PI by PIase C also can cause modifications in membrane-enzyme activities, e.g., adenylate cyclase.
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PMID:Minireview. Phosphatidylinositol specific phospholipases C. 708 67

We used thapsigargin (TG), 2,5-di-tert-butyl-1,4-benzohydroquinone (BHQ) and cyclopiazonic acid (CPA), each of which inhibits microsomal Ca(2+)-ATPase, to evaluate the effects of this inhibition on cytoplasmic free calcium ([Ca2+]i) and secretagogue-stimulated enzyme secretion in rat pancreatic acini. Using single-cell microspectrofluorimetry of fura-2-loaded acini we found that all three agents caused a sustained increase in [Ca2+]i by mobilizing calcium from inositol-(1,4,5)-trisphosphate-sensitive intracellular calcium stores and by promoting influx of extracellular calcium. Concentrations of all three agents that increased [Ca2+]i potentiated the stimulation of enzyme secretion caused by secretagogues that activate adenylate cyclase but inhibited the stimulation of enzyme secretion caused by secretagogues that activate phospholipase C. With BHQ, potentiation of adenylate cyclase-mediated enzyme secretion occurred immediately whereas inhibition of phospholipase C-mediated enzyme secretion occurred only after several min of incubation. In addition, the effects of BHQ and CPA on both [Ca2+]i and secretagogue-stimulated enzyme secretion were reversed completely by washing whereas the actions of TG could not be reversed by washing. Concentrations of BHQ in excess of those that caused maximal changes in [Ca2+]i inhibited all modes of stimulated enzyme secretion by a mechanism that was apparently unrelated to changes in [Ca2+]i. Finally, in contrast to the findings with TG and BHQ, CPA inhibited bombesin-stimulated enzyme secretion over a range of concentrations that was at least 10-fold lower than the range of concentrations over which CPA potentiated VIP-stimulated enzyme secretion.
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PMID:Effect of inhibition of microsomal Ca(2+)-ATPase on cytoplasmic calcium and enzyme secretion in pancreatic acini. 750 54

In rat jejunal brush-border membranes (BBM), ATP hydrolysis activity was specifically stimulated by CaCl2 and by MgCl2, allowing to identify Ca(2+)-ATPase and Mg(2+)-ATPase activities with a broad pH optimum near 8.0. Nonspecific ATPase activity (in the absence of cations) had a pH optimum above 9.5 as alkaline phosphatase. The effects of Ca2+ and Mg2+ concentrations on ATPase activity evidenced two apparent KA for each cation. At high concentrations, a similar affinity for both cations was recorded (KA: 0.35 mM). At low concentrations, the affinity for Mg2+ was greater than for Ca2+ (KA: 0.02 mM and 0.07 mM respectively). In an attempt to differentially solubilize alkaline phosphatase and ATPase activities, eleven different detergents were assayed. They more or less successfully released Ca(2+)-ATPase and Mg(2+)-ATPase activities from BBM but the more membranes were solubilized by a detergent, the more activities were lost, suggesting a close dependence on integration in BBM. As to alkaline phosphatase and nonspecific ATPase, they almost co-solubilized with Ca(2+)-ATPase and Mg(2+)-ATPase but their total activity was little affected. After treatment of BBM with phosphatidylinositol-specific phospholipase C (E.C. 3.1.4.10), 58% of alkaline phosphatase activity and 45% of nonspecific ATPase activity were released in the supernatant while Ca(2+)-ATPase and Mg(2+)-ATPase activities remained totally incorporated in BBM pellets. These last results definitively demonstrate that Ca(2+)-ATPase and Mg(2+)-ATPase activities are not manifestations of alkaline phosphatase, as earlier suggested, but rather result from the existence of one or several intrinsic membrane enzymes.
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PMID:Ca(2+)-ATPase and Mg(2+)-ATPase activities distinct from alkaline phosphatase in rat jejunal brush-border membranes. 751 33

Rabbit gastric glands were treated with alpha-toxin to test for permeabilization of basolateral membrane and retention of functional activity of parietal cells. Treatment with up to 400 U alpha-toxin/mL resulted in a dose-dependent increase in permeabilization, as judged by nuclear uptake of trypan blue (960 daltons), while causing relatively little loss of cytoplasmic macromolecules in the size range of lactate dehydrogenase (134,000 daltons). In the presence of cAMP and ATP, alpha-toxin-permeabilized resting gastric glands were stimulated to accumulate aminopyrine by approximately 10-fold over glands incubated without added nucleotides. Aminopyrine accumulation in stimulated permeabilized glands was inhibited by specific H+,K(+)-ATPase inhibitors, omeprazole and SCH-28080, and by the selective inhibitor of protein kinase A, H-89 (IC50 = 7.17 +/- 2.05 microM; n = 4). Aminopyrine accumulation in the alpha-toxin-treated glands was dependent on both exogenous ATP and cAMP; however, when no exogenous ATP was present, cAMP-activated aminopyrine accumulation reached approximately 50% of maximum, and at levels of ATP > 0.05 mM, maximal aminopyrine accumulation occurred without exogenous cAMP. In the presence of ATP alone, aminopyrine accumulation in permeabilized glands achieved 61.1 +/- 3.2% (n = 10; range, 50-70%) of the values measured on paired samples of intact glands stimulated with histamine plus isobutylmethylxanthine. These results demonstrate the functional responsiveness of alpha-toxin-permeabilized resting gastric glands. The participation of a protein kinase A dependent pathway during activation of permeabilized parietal cell is proposed.
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PMID:Acid secretion in alpha-toxin-permeabilized gastric glands. 752 Jul 7

The nature of the senktide response of the human NK3 receptor expressed in Chinese hamster ovary cells was characterised using the Ca2+ sensitive dye Fura-2 and imaging methods. Application of the NK3 receptor agonist senktide caused an increase in [Ca2+]i in the cells. The profile for NK3 receptor agonists was that senktide was more potent than [beta-Ala8]neurokinin A-(4-10) which was more potent than [Sar9,Met(O2)11]substance P. SR 48968 was a poor antagonist of the senktide response in intact cells confirming the weak affinity of this agent for the NK3 receptor (IC50 of approximately 1 microM) shown in binding assays. The NK3 receptor mediated increase in intracellular Ca2+ was independent of [Ca2+]o, blocked by the microsomal Ca2+ ATPase inhibitor thapsigargin and the phospholipase C inhibitor U73122 but not by ryanodine. Thus the source of the Ca2+ was probably a ryanodine insensitive, inositol triphosphate sensitive intracellular store.
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PMID:Characterization of tachykinin mediated increases in [Ca2+]i in Chinese hamster ovary cells expressing human tachykinin NK3 receptors. 753 Feb 8

When applied at low concentrations (< 10 micrograms/ml), staphylococcal alpha-toxin generates a small channel in keratinocyte and lymphocyte membranes that permits selective transmembrane flux of monovalent ions. Here we show that a moderate concentration (1-50 micrograms/ml) of alpha-toxin similarly produces a small pore in membranes of human fibroblasts. This process leads to rapid leakage of K+ and to a drop in cellular ATP to 10-20% of normal levels in 2 h. In the presence of medium supplemented with serum and at pH 7.4, the cells are able to recover from toxin attack, so that normal levels of K+ and ATP are reached after 6-8 h at 37 degrees C. The repair process is dependent on the presence of serum in the medium and is very sensitive towards pH. Decreases of pH in the medium to < or = 7.0 as well as increases to > or = 7.8 causes the repair mechanism to fail. The fate of cell-bound toxin molecules was investigated by using a radiolabelled tracer and by immunological detection of toxin exposed at the cell surface. The results indicated that 50-70% of the toxin was shed from cell membranes. However, there was no clear correlation between shedding and recovery, and shedding was also observed in cells that died at pH 7.8. Shedding was not decisive for repair, since cells that had recovered from toxin attack continued to carry 30-40% of initially bound toxin on their cell surface. Blockade of Na+/K(+)-ATPases with ouabain evoked similar kinetics of K(+)-depletion in control cells, compared with cells that had just recuperated from toxin attack and that still carried 30-40% alpha-toxin on their surface. We therefore tentatively concluded that repair of alpha-toxin lesions was due to closure of small pores, rather than from compensation of membrane leaks by up-regulation of Na+/K(+)-ATPase activity. We speculate that repair of small membrane lesions may extend to other agents that produce channels of similar nature in nucleated cells. Larger pores created by E. coli hemolysin or streptolysin O, both of which form larger functional transmembrane lesions, could not be repaired by fibroblasts.
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PMID:Recovery of human fibroblasts from attack by the pore-forming alpha-toxin of Staphylococcus aureus. 753 74

Whether phosphatidylinositol hydrolysis and a subsequent Ca2+ mobilization are responsible for muscarine-induced transient outward currents (IO) and non-selective cation currents (INS) in the guinea-pig chromaffin cell was investigated using the perforated patch method. IO, but not INS, failed to be reproduced in Ca(2+)-free solution and was markedly reduced by prior exposure to caffeine under Ca(2+)-free conditions or by addition to normal solution of cyclopiazonic acid (CPA), a Ca2+ ATPase inhibitor. Application of CPA in Ca(2+)-free solution, however, suppressed INS by about 50% in 73% of the cells tested. Bath application of 1.5 mM neomycin, a phospholipase C inhibitor, induced the time-dependent decline of IO with near abolition at 20 min or less, whereas it produced a time-independent decrease of INS and an inwardly rectifying K+ current. INS in the presence or absence of neomycin was well fitted to rectangular hyperbolas with the same ED50 of 2.17 microM, but with a 33% smaller maximum amplitude in the former, indicating a non-competitive inhibition by neomycin. We conclude that, while phosphatidylinositol hydrolysis mediates the production of IO, it does not mediate that of INS by muscarine.
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PMID:Phosphatidylinositol hydrolysis is involved in production of Ca(2+)-dependent currents, but not non-selective cation currents, by muscarine in chromaffin cells. 754 Jan 39


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