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:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gonadotropin receptors associated with plasma membrane fractions were solubilized by detergents, including Triton X-100, Lubrol WX, Lubrol PX and sodium deoxycholate before and after equilibration with 125I-labelled human chorionic gonadotropin. The binding activity remained in solution even after centrifugation at 300 000 X g for 3 h. The solubilized gonadotropin receptor or gonadotropin receptor complex was characterized by gel filtration and sucrose density gradient centrifugation. Sucrose density gradient centrifugation of solubilized gonadotropin-receptor complex in the presence of Triton X-100 had a sedimentation coefficient of 6.5 S whereas the solubilized uncomplexed receptor had a sedimentation coefficient of 5.1 S. In the absence of the detergent, solubilized hormone receptor complex from plasma membrane fractions I and II sedimented with an apparent sedimentation coefficient of 6.6 S and 7.4 S, respectively. Similarly, the free receptor also showed higher sedimentation profile with an apparent sedimentation coefficient of 6.7 S for fraction I and 7.2 S for fraction II. Treatment of plasma membranes with phospholipase A and C inhibited the binding of 125I-labelled human chorionic gonadotropin in a dose dependent manner, whereas phospholipase D was without any effect. Doses of 1.4 mI. U. of phospholipase A or 0.6 mI.U. of phospholipase C were required to produce 50% inhibition of the binding activity. These phospholipases had no effect on the preformed 125I-labelled human chorionic gonadotropin-receptor complex nor on the sedimentation profile of solubilized gonadotropin receptor complex.
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PMID:Sedimentation behavior of solubilized gonadotropin receptor from plasma membranes of bovine corpus luteum. 17 61

The binding of staphylococcal [125I]alpha-toxin to rabbit vagus nerves in vitro was a saturable process. The radiolabeled alpha-toxin binding was reduced by the coaddition of added navive alpha-toxin, indicating that the binding is specific. Sucrose gradient analysis of detergent-extracted complexes of [125I]alpha-toxin-rabbit vagus nerves showed both high and low S-value peaks analogous to those observed with similarly treated alpha-toxin-rabbit erythrocyte preparations (P. Cassidy and S. Harshman, Biochemistry, in press).
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PMID:Specific binding of staphylococcal alpha-toxin to isolated rabbit vagus nerves in vitro. 71 27

F9 mouse teratocarcinoma and PyS-2 cells in culture incubated with monovalent cations in buffered sucrose solution (0.25 M) can secrete as much as 40% of their total lysosomal enzymes into the medium within 30 min. Longer incubation does not lead to further loss of enzyme, suggesting that only a certain fraction of lysosomes is capable of discharge. The simultaneous presence of sucrose and cation, each at the respective optimal concentrations of 0.25 and 0.15 M, is required for lysosomal discharge (i.e. twice isoosmolarity). The cells remain fully viable. Sodium ions are more effective than lithium and potassium ions, whereas amines and divalent cations are less effective. Other sugars including glucose can replace sucrose to varying extents. Secretion is accompanied by a rapid short-lived rise in the level of cAMP. Forskolin as well as agents that activate G protein such as cholera toxin, AlF4-, and vanadate ions also increase the rate of secretion. Sucrose-Na+ stimulation takes place independently of changes in influx or efflux of calcium ions or changes in the levels of extracellular or free intracellular calcium ions. Neomycin, an inhibitor of phospholipase C, has little effect on secretion. Our results suggest that the secretion observed is mediated by a cAMP-dependent mechanism involving G proteins. Calcium ions and phospholipase C appear to play little or no part in the activation process.
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PMID:Stimulated secretion of lysosomal enzymes by cells in culture. 254 92

Human polymorphonuclear leucocytes (PMNL) inactivate Clostridium difficile cytotoxin and C. perfringens phospholipase C, but not C. perfringens enterotoxin. Both whole cells and sonicated suspensions possess activity, but mononuclear cell fractions of peripheral blood do not. Antitoxin activity closely correlates with cell concentration. The highest cell concentrations tested completely inactivated C. difficile cytotoxin by 2 min. Sucrose density gradient fractionation of PMNL showed antitoxin activity to be associated with myeloperoxidase, locating it in the primary or azurophil granules. Toxin inactivation was prevented by protease inhibitors suggesting that it is due to one of the neutral proteases present in these granules. PMNL are more active against C. difficile cytotoxin than purified chymotrypsin. PMNL may be a primary defence against certain bacterial exotoxins.
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PMID:Antitoxin activity of human polymorphonuclear leucocytes. 285 49

Two enzymes, alkaline phosphatase and acetylcholinesterase (AChE), have been shown previously to be components of the surface of the trematode parasite Schistosoma mansoni. In this study we report that both these enzymes and other serine hydrolases are susceptible to release from the S. mansoni tegumental membrane by a phosphatidylinositol-specific phospholipase C (PIPLC) of bacterial origin. These data suggest that AChE and alkaline phosphatase of S. mansoni, as in higher organisms, are anchored to the membrane via covalently attached phosphatidylinositol. The release of AChE from the vesicular fraction of the parasite with PIPLC occurs in a concentration-dependent manner. Sucrose gradient centrifugation of the PIPLC-released AChE showed a single 8.3 S molecular form, similar to that observed for AChE solubilized by Triton X-100. PIPLC removed large amounts of AChE from the surface of intact schistosomula in culture, with no impairment of the viability of the parasite. In this case, an increase in the overall levels of AChE in the intact parasite was observed after addition of PIPLC.
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PMID:Acetylcholinesterase in Schistosoma mansoni is anchored to the membrane via covalently attached phosphatidylinositol. 313 66

The susceptibility to phosphatidylinositol-specific phospholipase C of the membrane associated acetylcholinesterase (AChE) forms of Xenopus laevis skeletal muscle was examined. This treatment released almost all the detergent-soluble AChE species from muscle homogenates. Sucrose gradient analysis showed that the released acetylcholinesterase form corresponds to a hydrophilic G2 dimer, indicating that this dimer has a glycolipid anchoring domain which contains phosphatidylinositol.
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PMID:A membrane-associated dimer of acetylcholinesterase from Xenopus skeletal muscle is solubilized by phosphatidylinositol-specific phospholipase C. 341 41

Escherichia coli hemolysin is secreted as a water-soluble polypeptide of Mr 107,000. After binding to target erythrocytes, the membrane-bound toxin resembled an integral membrane protein in that it was refractory towards extraction with salt solutions of low ionic strength. Toxin-induced hemolysis could be totally inhibited by addition of 30 mM dextran 4 (mean Mr, 4,000; molecular diameter approximately 3 nm) to the extracellular medium. Uncharged molecules of smaller size (e.g., sucrose, with a molecular diameter of 0.9 nm, or raffinose, with a molecular diameter of 1.2 to 1.3 nm) did not afford such protection. Treatment of erythrocytes suspended in dextran-containing buffer with the toxin induced rapid efflux of cellular K+ and influx of 45Ca2+, as well as influx of [14C]mannitol and [3H]sucrose. [3H]inulin only slowly permeated into toxin-treated cells, and [3H]dextran uptake was virtually nil. Membranes lysed with high doses of E. coli hemolysin exhibited no recognizable ultrastructural lesions when examined by negative-staining electron microscopy. Sucrose density gradient centrifugation of deoxycholate-solubilized target membranes led to recovery of the toxin exclusively in monomer form. Incubation of toxin-treated cells with trypsin caused limited proteolysis with the generation of membrane-bound, toxin-derived polypeptides of Mr approximately 80,000 without destroying the functional pore. We suggest that E. coli hemolysin may damage cell membranes by partial insertion into the lipid bilayer and generation of a discrete, hydrophilic transmembrane pore with an effective diameter of approximately 3 nm. In contrast to the structured pores generated by cytolysins of gram-positive bacteria such as staphylococcal alpha-toxin and streptolysin O, pore formation by E. coli hemolysin may be caused by the insertion of toxin monomers into the target lipid bilayers.
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PMID:Escherichia coli hemolysin may damage target cell membranes by generating transmembrane pores. 351 65

A specific binding site for somatotropin was solubilized by 1% (v/v) Triton X-100 from a crude particulate membrane fraction of pregnant rabbit liver, partially purified and characterized. The solubilized binding site retained many of the characteristics observed in the original particulate fraction, indicating that extraction of the binding site with Triton X-100 does not cause any major changes in its properties. The binding of human 125I-labelled-somatotropin to the solubilized binding site is a saturable and reversible process, depending on temperature, incubation time, pH and ionic environment. Analysis of the kinetic data revealed a finite number of binding sites with an affinity constant of 0.32 x 10(10)M-1. The binding activity for human 125I-labelled-somatotropin was adsorbed to a concanavalin-A-Sepharose column and was dissociated from the column with alpha-methyl-D-glucoside, suggesting that the binding protein may be a glycoprotein. Using affinity chromatography on concanavalin-A-Sepharose, ion-exchange chromatography on DEAE-cellulose and gel filtration on Sepharose 6B, the binding protein was purified 1000-4000-fold from the original liver homogenate. When the partially purified preparation was chromatographed on Sepharose 6B, the binding protein eluted as a molecule with an apparent molecular weight of 200000, with a Stokes' radius of 4.9 nm. Sucrose-density-gradient centrifugation of the preparation showed that the sedimentation coefficient of the binding protein was 7.2S. Isoelectric focusing experiments revealed that a major part of the protein has an acidic pI (4.2-4.5). Exposure of the protein to trypsin decreased the binding activity for human 125I-labelled-somatotropin or bovine 125I-labelled-somatotropin, whereas ribonuclease, deoxyribonuclease, phospholipase C or neuraminidase had little or no effect.
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PMID:Characteristics of solubilized human-somatotropin-binding protein from the liver of pregnant rabbits. 624 70

Extracellular ATP activates two distinct types of P2 purinoreceptor-mediated signaling pathways in macrophages, 1) the rapid formation of nonselective pores/channels in the plasma membrane and 2) a guanine nucleotide-binding protein-dependent stimulation of phosphotidylinositol-specific phospholipase C, with subsequent mobilization of intracellular Ca2+. Several studies have suggested that the pore-forming, or P2z, purinoreceptor may be involved in the cytolytic effects of ATP on macrophages and other cell types. We have identified 3'-O-(4-benzoyl)benzoyl-ATP (BzATP) and UTP as selective agonists for the P2z purinoreceptor and Ca(2+)-mobilizing nucleotide receptor, respectively, in BAC1.2F5 macrophages. In this paper we demonstrate that BzATP and ATP (but not UTP) activate membrane depolarization in BAC1.2F5 cells and also stimulate appropriate electrophysiological responses, consistent with the expression of the P2z purinoreceptor, in Xenopus oocytes injected with poly(A)+ RNA derived from BAC1.2F5 cells. Micromolar concentrations of BzATP or millimolar concentrations of ATP induced a sustained increase in the membrane holding current in these voltage-clamped oocytes. This response was significantly potentiated in the absence of extracellular divalent cations, consistent with the specificity of the P2z purinoreceptor for tetrabasic nucleotides. The sustained currents induced by BzATP or ATP were distinct from the transient and/or oscillating increases in Ca(2+)-dependent Cl- currents that were stimulated by UTP but not BzATP. UTP-stimulated transient currents and nucleotide-dependent increases in aequorin luminescence in poly(A)+ RNA-injected oocytes were independent of extracellular Ca2+ and were correlated with the mobilization of intracellular Ca2+ stores. Sucrose fractionation of the poly(A)+ RNA from BAC1.2F5 cells resulted in the enrichment of mRNA species encoding the components of the P2z purinoreceptor, as well as the Ca(2+)-mobilizing nucleotide receptor, in fractions containing 2.5-4.0-kilobase species.
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PMID:Expression of the pore-forming P2Z purinoreceptor in Xenopus oocytes injected with poly(A)+ RNA from murine macrophages. 768 70

Sucrose-density flotation analysis of Triton-insoluble membrane domains isolated from highly purified sheep ventricular sarcolemma revealed the presence of two major 120- and 100-kDa proteins. Both species migrated in two-dimensional isoelectric focussing/SDS gels with an apparent pI of approximately 4.3, suggesting that they might be related. Microsequence analysis of peptides derived from the 100-kDa protein yielded amino acid sequences with high homology to T-cadherin, a truncated cadherin lacking a cytoplasmic domain. The similarity was confirmed using antibodies to chicken T-cadherin that reacted with both proteins on immunoblots. T-cadherin was released from the detergent-insoluble sarcolemmal fraction by phospholipase C treatment indicating that it is linked to the membrane by a glycophosphoinositol anchor. T-cadherin could be ADP-ribosylated by a transferase that was also present in the caveolin-enriched Triton-insoluble fraction. T-cadherin-containing membrane fragments cofractionated on sucrose gradients with caveolin-3, a marker protein for myocyte caveolae. However, immunopurified caveolin-3-containing membranes contained no associated T-cadherin. Immunocytochemical analysis of cultured rat atrial myocytes revealed that T-cadherin and caveolin have related but nonoverlapping staining patterns. These results suggest that T-cadherin is a major glycophosphoinositol-linked protein in cardiac myocytes and that it may be located in plasma membrane "rafts" distinct from but possibly adjacent to caveolae.
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PMID:T-cadherin is a major glycophosphoinositol-anchored protein associated with noncaveolar detergent-insoluble domains of the cardiac sarcolemma. 950 99


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