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 effects of pH, trypsin, and phospholipase C on the topographic distribution of acidic anionic residues on human erythrocytes was investigated using colloidal iron hydroxide labeling of mounted, fixed ghost membranes. After glutaraldehyde fixation at pH 6.5-7.5, the positively charged colloidal particles were bound to the membranes in small randomly distributed clusters. The clusters of anionic sites were reversibly aggregated by incubation at pH 5.5 before fixation at the same pH. These results correlate with the distribution of intramembranous particles found by Pinto da Silva (J. Cell Biol.53:777), with the exception that the distribution of anionic sites on a majority of the fixed ghosts at pH 4.5 was aggregated instead of dispersed. The randomly distributed clusters could be nonreversibly aggregated by trypsin or phospholipase C treatment of intact ghosts before glutaraldehyde fixation. Previous glutaraldehyde fixation prevented trypsin and pH induced aggregation of the colloidal iron sites. Evidence that N-acetylneuraminic acid groups are the principal acidic residues binding colloidal iron was the elimination of greater than 85% of the colloidal iron labeling to neuraminidase-treated cell membranes. Colloidal iron binding N-acetylneuraminic acid residues may reside on membrane molecules such as glycophorin, a sialoglycoprotein which contains the majority of the N-acetylneuraminic acid found on the human erythrocyte membrane.
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PMID:Anionic sites of human erythrocyte membranes. I. Effects of trypsin, phospholipase C, and pH on the topography of bound positively charged colloidal particles. 412 Dec 89

Fixation of embryonic chick cells (heart, neural retina, and limb bud) in the presence of lanthanum ions shows the presence of an electron-opaque layer, about 50 A thick, external to the cell membrane. This layer, designated LSM (for lanthanum-staining material), is not removable by trypsin, pronase, EDTA, DNase, alpha-amylase, neuraminidase, or N-acetyl-L-cysteine. However, phospholipase C, in concentrations as low as 0.001 mg/ml, succeeds in stripping the LSM from the cell surface. Heating the enzyme preparation does not inhibit this activity, but removal of divalent cations does; both of these results are consistent with the known properties of phospholipase C. The LSM is present at the cell surface in the control tissues and on cells dissociated from the tissues by proteolytic enzymes and EDTA. These results are interpreted to mean that the LSM is probably an integral part of the cell and not an extraneous coat. How this phenomenon bears on the problem of cellular adhesion is discussed, as is the possible chemical composition of the LSM.
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PMID:The removal by phospholipase C of a layer of lanthanum-staining material external to the cell membrane in embryonic chick cells. 416 2

Australia antigen [Au(1)], a particle associated with viral hepatitis, was isolated from the plasma of a patient with chronic anicteric hepatitis and leukemia who had received radioactive phosphorus. We have found that the immunoreactivity and appearance of Au(1) in the electron microscope were not altered by treatment with enzymes including trypsin, pronase, lipase, phospholipase C, ribonuclease, deoxyribonuclease, amylase, and neuraminidase. In contrast, other serum constituents were degraded by these enzymes. Therefore, treatment of the patient's plasma with many enzymes was exploited as an initial step for the isolation of Au(1). Subsequently, Au(1) was purified from the enzyme-treated (32)P-labeled plasma by gel filtration through Sephadex G-200 and centrifugation through sucrose and in cesium chloride gradients. There were no detectable human serum components in the purest fractions, as tested by immunoelectrophoresis and immunodiffusion. The density of the purified Au(1) was 1.21 in CsCl. The particle measured about 200 A in diameter, was predominantly spherical in shape and appeared to be composed of subunits. Nucleic acids were not detected by spectrophotometric, radiochemical, and chemical analyses. Immunoreactivity of purified Au(1) was destroyed by heating for 1 hr at 85 degrees C but was stable at 56 degrees C. Treatment with Carnoy's solution (3 parts ethanol:1 part glacial acetic acid) followed by pronase disrupted the particles as seen with the electron microscope. These findings, combined with other published information on Australia antigen and viral hepatitis, suggest that the bulk of Australia antigen in the blood of this patient is an incomplete virus or virus capsid.
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PMID:Australia antigen (a hepatitis-associated antigen): purification and physical properties. 424 40

Broth cultures of Clostridium perfringens (ATCC 10543) were fractionated by ammonium sulfate precipitation and Sephadex G-150 chromatography. Components isolated, as well as some enzymes present in the culture, were assayed for toxicity by feeding to white mice. Early work indicated that when a meat-fat-starch slurry, infected with C. perfringens, was fed to mice, the intestinal passage time was reduced. By using large numbers of mice as test animals and analyzing the data statistically, we found that C. perfringens and several fractions from the culture supernatant significantly affected the mice. A toxic material present in the supernatant was not identifiable as phospholipase C. Phospholipase C and physphorylcholine affected the intestinal passage time of the mice only when large amounts were given. The enzyme, neuraminidase, and another unidentified compound present in the supernatant affected the passage time when very small amounts were fed to mice.
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PMID:Response of white mice to cells and culture constituents of Clostridium perfringens. 430 35

The structure and function of the platelet surface was probed by phospholipase C (Clostridium perfringens) which hydrolyzes membrane phospholipids, particularly phosphatidylcholine. Platelet phospholipids were susceptible to phospholipase C, and extent of hydrolysis was dependent on concentration of phospholipase C and Ca(++). Phospholipase C (0.15 U/ml) with Ca(++) (0.55 mM) hydrolyzed 15.6% phospholipids during 5 min. Phospholipase C released platelet serotonin (5HT), ADP, and platelet factor 4. Hydrolysis of 5% phospholipids resulted in release of 70% 5HT. Platelet 5HT release was rapid, occurring within 2 min. Phospholipase C (0.2 U/ml) with Ca(++) (0.55 mM) also released 10.35 nmol sotrage pool ADP/10(9) platelets and 63% platelet factor 4 during 3 min. Phospholipase C did not cause leakage of cytoplasmic metabolic pool ADP, since only 6.6% [(3)H]ADP was released. Ultrastructural analysis of phospholipase C-modified platelets showed that platelets were intact. After 2% phospholipid hydrolysis, centralization of granules and contraction of microtubules were evident. After 18% phospholipid hydrolysis, there were morphological indications of degranulation. Phospholipase C-induced phospholipid hydrolysis caused the release of ADP and 5HT since: (a) Phospholipase C purified by heating was shown to be free of protease and neuraminidase activity and capable of inducing the platelet release reaction. (b) Antitoxin (Cl. perfringens) neutralized phospholipase C-induced 5HT release which rules out a contaminant. (c) Phosphorylcholine, the hydrolysis product, did not induce platelet 5HT release. This study demonstrates that minimal hydrolysis of platelet phospholipids triggers the release reaction. Our hypothesis is that phospholipids, presumably phosphatidylcholine, are situated at or near active site or "receptor" on the platelet surface and function as the modulator for the release reaction.
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PMID:The role of platelet membrane phospholipids in the platelet release reaction. 437 63

Leukocytes can generate procoagulant (tissue factor) activity when incubated with endotoxin. These studies were undertaken to determine whether platelets could influence the procoagulant activity generated by leukocytes. Intact or disrupted platelets (rabbit or human) enhanced the clot-promoting properties of rabbit leukocytes. The enhancing effect of human platelets on human leukocytes required the presence of human serum (devoid of factor VII and X activities). When platelets were incubated with endotoxin in the absence of leukocytes, no increase in their clot-promoting properties was discernible. However, a mixture of platelets, leukocytes, and endotoxin generated procoagulant activity which appeared rapidly and was fivefold greater than that produced by leukocytes incubated with endotoxin alone. The enhancement produced by platelets was even more pronounced if homogenates were used. The platelet effect was examined in more detail by the substitution of membranes, granules, and the "soluble" fraction for whole platelets in the test system. The stimulating activity was localized to the particulate fractions, i.e., membranes and granules. Prior treatment of platelet membranes with phospholipase C or gangliosides or by extraction of lipid resulted in loss of enhancing activity, whereas no inhibition was observed after exposure to neuraminidase or trypsin. It is proposed that platelets contribute a membrane lipoprotein surface which enhances the procoagulant activity generated by leukocytes in the presence of endotoxin. This mechanism may be involved in some of the clinical and pathologic manifestations of gram-negative sepsis with disseminated intravascular coagulation.
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PMID:The stimulatory effect of platelets and platelet membranes on the procoagulant activity of leukocytes. 461 59

Phospholipase C (EC 3.1.4.3.) from Clostridium perfringens (crude extracts) was used to study the role of phospholipids in the osmotic permeability of the urinary bladder of the toad. When added to the serosal bath (430 mU/ml) it inhibited the effects of antidiurectic hormone (ADH) and exogenous cyclic AMP. Under the same conditions the increase in osmotic flow produced by serosal hypertonicity (SH) was slightly enhanced by the lipase. The hydroosmotic effect of SH was greatly potentiated by the lipase by decreasing 10-fold the Ca2+ concentration. The SH-induced flow was inhibited by the lipase if the Ca2+ or the H+ concentration was increased 10-fold, but not if the increase in positive charges was produced by a concentration of Mg2+. Phospholipase C had no effect on the action of either ADH or SH if added to the mucosal bath. Serosal neuraminidase or phospholipase A2 could not mimic the effect of phospholipase C on SH. The effect of phospholipase C on the response to SH was not modified if fatty acid-free bovine serum albumin was added to the bath. Therefore, the release of products of lipolysis into the bath do not seem to be responsible for the effects of phospholipase C on SH-induced water flow. The results suggest that the effects of the enzyme on the composition and rearrangement of lipids at the basolateral membrane produce modifications of the water flow. Ca2+ and H+ may modify the enzyme-substrate interaction, suggesting that different phospholipids may be differentially involved in the control of water permeability of the basolateral membrane.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Ca2+- and H+-dependent effects of crude bacterial phospholipase C on the hydroosmotic response of toad urinary bladder to serosal hypertonicity. 608 40

[3H]prostaglandin E2 (PGE2) binding receptors exist in rabbit alveolar bone cell membranes. The presence of high (Kd = 3.9 X 10(-9) M) and low (Kd = 8.8 X 10(-8) M) affinity binding sites of [3H]PGE2 was demonstrated. The saturation values of [3H]PGE2 for high and low affinity binding sites were 0.13 pmol/mg protein and 1.22 pmol/mg protein, respectively. The digestion of the membranes with pronase, phospholipase C, D and neuraminidase led to a decrease of [3H]PGE2 binding but phospholipase A2 did not.
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PMID:Some properties of prostaglandin E2 receptors in rabbit alveolar bone cell membranes. 613 25

The intestinal epithelium of Ascaris suum consists of a single layer of tall columnar epithelial cells that rest on a thick basal membrane in contact with the pseudocoelomic cavity. Experiments were conducted on glutaraldehyde-fixed tissue to ascertain the nature of the electronegative charges associated with both the apical microvillar surface and basal membrane. A strong electronegative charge was demonstrated on the microvillar surface and basal membrane with ruthenium red and cationic ferritin staining. The ionic nature of ferritin binding was demonstrated with poly-L-lysine, a polycation that interacts with anionic groups on the membrane and thus blocks the subsequent binding of ferritin. Tissue thus treated was devoid of reaction product. Methylation with diazomethane completely abolished staining. Since the stronger acidic groups of sulfates or phosphates would not be protonated under the conditions employed in this study, and therefore susceptible to methylation, staining by ferritin is thought to be due to its interaction with carboxyl groups. Prior enzymatic treatment of tissue with neuraminidase or phospholipase C had no effect on subsequent ferritin binding. Tissue exposed to colloidal iron at various pH values showed maximal reactivity at a pH of 2.5 or above. Above pH 2.5, the dissociation of protons from free carboxyl groups of protein-bound amino-acid residues with pK's of 3.8 and 4.2 would be maximal, and the ionized carboxyl groups are then available to interact with iron micelles. These results suggest the presence of weaker acidic groups, such as the carboxyl groups of acidic amino acids or uronic acid residues. The stronger acidic groups of sialic acid and the esterified sulfate groups, if present, contribute only minimally to overall staining. These results demonstrate that a high electronegative charge density exists, despite the apparent lack of sialic acid. Staining is believed to be due to carboxyl groups of acidic amino acids and/or carboxyl groups or uronic acid residues.
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PMID:Ultrastructural observations on the cell surface of the intestinal epithelium of the nematode, Ascaris suum. Nature of the electronegative charge. 615 29

Colloidal iron staining, calcium binding and enzyme activities were studied in the isolated rat heart sarcolemma. Colloidal iron staining of the sarcolemma revealed a high density of negatively charged sites associated with the cell surface. This membrane fraction was found to have calcium binding activity at both low (0.1 mM) and high (1.25 mM) concentrations of calcium. Pretreatment of the sarcolemma with either trypsin, phospholipase C or neuraminidase, was associated with a reduction in colloidal iron staining as well as decreased calcium-binding activity at high concentrations of calcium. Calcium binding at low concentrations was decreased by both trypsin and neuraminidase. Mg2+ ATPase, Ca2+ ATPase, and Na+-K+ ATPase activities were altered by neuraminidase and trypsin treatments, whereas phospholipase C treatment altered Na+-K+ ATPase only. It is concluded that both surface negative charge and calcium-binding sites associated with the isolated rat heart sarcolemma are contributed by a mosaic of biomolecules including proteins, phospholipids and glycoproteins, and alterations in the surface charge may influence the activities of membrane-bound enzymes.
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PMID:Negatively charged sites and calcium binding in the isolated rat heart sarcolemma. 616 50


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