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)

We have previously shown that membranes from the retinal pigment epithelium can transform added all-trans-retinol into a mixture of 11-cis-retinoids, demonstrating the "missing reaction" in the visual cycle for the first time (Bernstein, P. S., Law, W. C., and Rando, R. R. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 1849-1853). In this article, this isomerase activity is further characterized. Double-label experiments with [15-3H]- and [15-14C]all-trans-retinol as the substrate show that the tritium label is retained in the 11-cis-retinol and 11-cis-retinyl palmitate products. This requires that isomerization occur at the alcohol level of oxidation. All-trans-retinyl esters, such as the palmitate, acetate, butyrate, and hexanoate esters, are not directly transformed into their 11-cis counterparts by the membranes. The data are consistent with the presence of an all-trans-retinol isomerase enzyme system or enzyme complex, which produces 11-cis-retinol. Other isomeric retinols were tested for substrate activity. Neither 9-cis-retinol(al) nor 13-cis-retinol were processed by the isomerase. Since the membranes containing the isomerase possess other retinol metabolizing activities, such as retinyl ester synthetase and dehydrogenase activities, further purification was attempted. Appreciable quantities of all detergents tested led to the disappearance of isomerase activity, and high salt or EDTA did not dissociate isomerase activity from the membranes. However, extensive sonication of the membranes did produce a 100,000 x g supernatant fraction of light membranes depleted of other all-trans-retinol processing activities. The isomerase activity in these membranes was saturable with all-trans-retinol, as required for a biologically significant process, and showed a Vmax of 5 pmol/h/mg of protein, a KM of 0.8 microM, and a pH optimum of 8. The isomerase was destroyed by proteinase K, by phospholipase C, by heating, or by ethanol at concentrations greater than 1%. The addition of high energy compounds, such as MgATP, MgGTP, or palmitoyl-CoA, did not appear to stimulate isomerase activity in the 100,000 x g supernatant.
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PMID:Biochemical characterization of the retinoid isomerase system of the eye. 350 Jan 73

Guanine nucleotides are thought to mediate the interaction of the receptors for calcium-mobilizing hormones and phosphoinositide-specific phospholipase C. In the present study the characteristics of guanine nucleotide-dependent phospholipase C activation were studied in [3H]inositol-labeled permeabilized hepatocytes. The nonhydrolyzable GTP analogs guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) and guanyl-5'-yl imidodiphosphate stimulated the production of inositol phosphates by phospholipase C. The effect was concentration-dependent with half-maximal and maximal stimulation occurring with 0.6 and 10 microM GTP gamma S, respectively. The guanine nucleotide-induced stimulation of phosphoinositide breakdown was selective for phosphatidylinositol (4,5)-bisphosphate over phosphatidylinositol (4)-phosphate. The individual inositol phosphates formed after maximal GTP gamma S exposure were analyzed by high-performance liquid chromatography. Inositol 1,4,5-trisphosphate was rapidly produced, followed by the formation of inositol 1,3,4,5-tetrakisphosphate and inositol 1,3,4-trisphosphate. Ethanol is known to activate hormone-sensitive phospholipase C in intact rat hepatocytes. Ethanol (0.3 M) was ineffective in altering the characteristics of GTP gamma S-stimulated phospholipase C activation, in both digitonin-treated and sonicated hepatocytes. The metabolism of the various inositol phosphate isomers was unaffected by ethanol. The findings demonstrate the potential for the use of permeabilized hepatocytes in the analysis of phospholipase C activation by guanine nucleotides. Ethanol does not activate phospholipase C by altering this process.
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PMID:Ethanol does not stimulate guanine nucleotide-induced activation of phospholipase C in permeabilized hepatocytes. 360 26

A novel alkyl-phospholipid with selective antitumor activity was isolated from an anticancer biopreparation cACPL (crude anticancer phospholipids) and from tissues of degenerating chick embryos. The alkyl-phospholipid was isolated and purified by chromatographic methods using silicic acid column chromatography and thin layer chromatography. The chemical structure of the alkyl-phospholipid was characterized by thin-layer chromatographic analysis of the degradation products after enzymatic digestion with phospholipase C from Bacillus cereus and with phospholipase D, by two-dimensional thin-layer chromatography, infrared spectrum, and mass spectrometric analysis. The alkyl-phospholipid was identified as 1-O-alkyl-2-acyl-sn-glycero-3-phospho-(N-acyl)-ethanolamine, i.e. plasmanyl-(N-acyl)-ethanolamine (PNAE), the main molecular species being 1-O-octadecyl-2-oleoyl-sn-glycero-3-phospho-(N-palmitoyl)-ethanol-amine. PNAE exhibits a selective cytolytic effect on human tumor cells HEp-2, HeLa and T24 in tissue cultures at the concentration of 25 micrograms PNAE per ml and 66-98% inhibition of DNA synthesis in the human tumor cells at concentration as low as 2.5 micrograms/ml, but it does not inhibit at a 50-fold higher concentration the DNA synthesis and normal growth of human fibroblasts (cell line LEP). PNAE represents the main biologically active antitumor component of the cACPL biopreparation and exhibits a significant antitumor effect in vivo in B10/An mice bearing Mc11 fibrosarcoma. Possible molecular mechanism of the selective antitumor activity of PNAE is discussed, involving the selective disturbance of phospholipid metabolism in tumor cells, leading to progressive destruction of tumor cell membranes. The fact that PNAE is nontoxic and selectively active against tumor cells at nanomolar concentrations in vitro as well as in vivo, indicates the possibility of its clinical use. PNAE and cACPL biopreparation might provide a very useful new tool for human anticancer chemotherapy.
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PMID:A novel nontoxic alkyl-phospholipid with selective antitumor activity, plasmanyl-(N-acyl)-ethanolamine (PNAE), isolated from degenerating chick embryonal tissues and from an anticancer biopreparation cACPL. 371 23

The role of the gastric nonwettable hydrophobic layer (surfactant) was investigated in the mucosal protection against the damage induced by ethanol in the rat. Although aluminium hydroxide inhibited the development of ethanol-produced gastric hemorrhagic lesions, it did not increase the mucosal phospholipid content. Ambroxol, a known stimulant of pulmonary surfactant production, protected the gastric mucosa against ethanol by increasing the phospholipid content. Surface-active compounds such as dimethyl polysiloxane also inhibited the development of gastric injuries caused by ethanol in a dose-dependent manner. The essential phospholipid-containing drug (Essentiale) also showed a strong and dose-dependent cytoprotective effect. Among the possible constituents of the gastric surfactant, sphingomyelin was totally ineffective. Phosphatidylcholine, phosphatidylinositol and phosphatidylglycerol were able to reduce the extent of mucosal damage in a dose-dependent manner. Gastric mucosal injuries were significantly aggravated by pretreatment with phospholipase C. In conclusion, these results suggest that either the maintenance or the strengthening or even the replacement of the gastric nonwettable hydrophobic lining between the damaging agent and the gastric mucosa may contribute to the cytoprotective mechanism of certain compounds.
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PMID:Cytoprotective role of gastric surfactant in the ethanol-produced gastric mucosal injury of the rat. 376 92

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

A new procedure for the purification of phospholipase C from Clostridium perfringens has been devised that results in essentially pure enzyme. The procedure consists of ammonium sulfate fractionation, ion-exchange chromatography on QAE-Sephadex, and affinity chromatography on phosphatidylcholine linked to Sepharose. The molecular weight of the enzyme, determined by sodium dodecyl sulfate-gel electrophoresis, amino acid analysis, and gel filtration, is 43,000; and the isoelectric point is pH 5.4. The enzyme was optimally active with phosphatidylcholine dispersed in sodium deoxycholate, although appreciable activity was observed with either phosphatidylcholine or sphingomyelin dispersed with ethanol. The requirement for metal ions in the assay could be met by a number of different ions. The pure enzyme was found to contain 2 mol zinc per mol enzyme, thus implicating it as a zinc metalloenzyme.
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PMID:Phospholipase C from Clostridium perfringens: preparation and characterization of homogeneous enzyme. 632

An extracellular bactericidal substance was isolated from the supernatant of Streptococcus mutans Rm-10 culture fluid and partially purified with 60% ammonium sulfate precipitation, differential centrifugation, and gel filtration on Sephadex G-200. There was a good correlation of the sensitivity profiles of indicator strains whether assayed on solid medium or with purified material from cell-free culture fluid, indicating that the same inhibitory substance is produced on solid medium and in broth. Vapor from organic solvents such as chloroform, acetone, ethanol, and ether as well as heat treatment at 100 degrees C for 30 min had little effect on the bactericidal factor. It was sensitive to trypsin and pronase and resistant to deoxyribonuclease, ribonuclease, lysozyme, and phospholipase C. The inhibitor was not infective, and electron microscopic studies failed to reveal phage or phage-like particles in concentrated solutions of the bactericidal material. The results indicate that the extracellular bactericidal substance is indeed a bacteriocin. Activity in broth cultures reached a maximum only after exponential growth had ceased. It was active against other streptococcal strains as well as strains of Actinomyces naeslundii, A. viscosus, Bacillus subtilis and Staphylococcus aureus, but not against strains of Fusobacterium nucleatum and Escherichia coli.
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PMID:Isolation, partial purification and preliminary characterization of a bacteriocin from Streptococcus mutans Rm-10. 641 23

Hemolymph (blood) of an insect, the tobacco hornworm, Manduca sexta, contains a phospholipase A1. The specificity of this enzyme was demonstrated by the use of substrates with labeled fatty acids in specific positions and by conversion of the enzyme product, a lysophospholipid, to 2-acylglycerol by the action of bacterial phospholipase C. The insect phospholipase A1 hydrolyzes phosphatidylethanolamine and phosphatidylglycerol, but not phosphatidylcholine or triolein. Divalent cation is required, with calcium ion being most effective, although strontium, barium, and magnesium ions also support activity. Only substrates dispersed in buffer from ethanol are hydrolyzed; ether, Triton X-100, and taurodeoxycholate are inhibitory. The enzyme has been purified 90-fold. At that stage, it is still far from homogeneous, but stability problems have hindered further purification. It has an apparent Mr = 155,000 +/- 11,000, estimated by gel permeation chromatography.
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PMID:A phospholipase A1 from the hemolymph of the tobacco hornworm, Manduca sexta. 683 59

By using a suckling mouse assay, heat-stable enterotoxin (ST) was purified from the culture filtrate of Yersinia enterocolitica isolated from a diarrheal patient. The purification procedures involve ultrafiltration with an Amicon HIP-10 hollow fiber, ethanol fractionation, protamine sulfate treatment, diethylaminoethyl-Sephacel and hydroxylapatite column chromatographies, and Sephacryl S-200 superfine gel filtration. About 408-fold purification was achieved, with a yield of 12.0%. The minimal effective dose of purified ST was about 110 ng in the suckling mouse assay. The molecular weight of purified ST was 9,000 by Sephadex G-100 superfine gel filtration. The purified ST was stable to heating (100 degrees C for 20 min, 121 degrees C for 20 min) and did not lose its toxicity after treatment with protease, trypsin, lipase, phospholipase C, ribonuclease, deoxyribonuclease, beta-glucosidase, and neuraminidase. The purified ST was separated by isoelectric focusing into two active fractions, with pI's of 3.29 (ST-1) and 3.00 (ST-2), respectively. Antiserum from guinea pigs immunized with the purified ST neutralized the activity of both Y. enterocolitica ST and Escherichia coli ST.
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PMID:Partial purification and characterization of heat-stable enterotoxin produced by Yersinia enterocolitica. 721 60

The hydrolysis of HeLa non-histone nuclear proteins over 24 h has been monitored in dilute alkali at 4, 15 and 25 degrees C using the standard ninhydrin estimation, dansylation and various electrophoresis techniques. Under conditions (up to 0.2 N NaOH, 4 degrees C) that do not release a significant quantity of ninhydrin-positive material or new N-terminal end group considerable breakdown was observed by two-dimensional electrophoresis analysis. The number of stained spots decreased from approx. 140 to 25--30. No internal protease activity could be found. Labelling studies (14C-labelled amino acids) showed that much of the hydrolysed material was extracted from the gel during normal staining and destaining procedures. Peptides could be extracted from alkali-hydrolysed non-histone protein with acid/ethanol and could be further separated by thin-layer chromatography on silica gel G. Short-term labelling of HeLa cells (14C-labelled amino acids for up to 60 min) revealed that these peptides probably have a high rate of turnover. [14C]Glucosamine studies also indicated the presence of considerable carbohydrate material in the low molecular weight products of this alkaline hydrolysis. Various standard proteins and histones were unaffected by hydrolysis in up to 0.2 N NaOH (4 degrees C, 24 h) as judged by gel electrophoresis. Seven different phosphate-splitting enzymes and an esterase had no effect on the non-histone protein electrophoresis patterns but a preparation of phospholipase C which had no protease activity towards eight standard proteins did produce considerable breakdown in HeLa non-histone proteins similar to that produced by 0.2 N NaOH at 4 degrees C.
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PMID:Studies on the degradation of HeLa non-histone proteins. 735 14


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