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 phosphatidylinositol phosphodiesterase from the culture broth of Bacillus cereus, was purified to a homogeneous state as indicated by polyacrylamide gel electrophoresis, by ammonium sulfate precipitation and chromatography with DEAE-cellulose and CM-Sephadex. The enzyme (molecular weight: 29000 +/- 1000) was maximally active at pH 7.2-7.5, AND NOT INFLUENCED BY EDTA, ophenanthroline, monoiodoacetate, p-chloromercuribenzoate or reduced glutathione. The enzyme specifically hydrolyzed phosphatidylinositol, but did not act on phosphatidylcholine, phosphatidylethanolamine and sphingomyelin, under the conditions examined. The products from phosphatidylinositol of enzyme reaction were diacylglycerols and a mixture of myoinositol 1- and 1, 2-cyclic phosphates, suggesting that the enzyme was a phosphatidylinositol-specific phospholipase C. The enzyme released alkaline phosphatase quantitatively from rat kidney slices. A kinetic analysis was made on the release of alkaline phosphatase. The results suggest that phosphatidylinositol-specific phospholipase C can specifically act on plasma membrane of rat kidney slices.
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PMID:Studies on phosphatidylinositol phosphodiesterase (phospholipase C type) of Bacillus cereus. I. purification, properties and phosphatase-releasing activity. 1 Sep 86

After alkaline extraction, purified subsynaptic fragments isolated from Torpedo electric tissue exhibit on sodium dodecyl sulfate/polyacrylamide gel electrophoresis predominant peptides of apparent Mr 41,000, 50,000, and 65,000 (i.e., the peptides characteristic of the nicotinic receptor purified and isolated in detergent solutions). The peptide of Mr 43,000 that is also found in the isolated postsynaptic membranes is recovered in the supernatant after alkaline extraction. The alkaline-extracted membranes were functionally intact, as demonstrated by the following criteria. The kinetics of binding of [3H]acetylcholine in the presence and absence of 30 micron carbamoylcholine to occupy acetylcholine binding sites, [14C]-meproadifen [2-(diethylmethylaminoethyl)-2,2-diphenylvalerate iodide ] was bound with a dissociation constant, KD, of 0.3 +/- 0.1 micron to 0.3 +/- 0.1 site per [3H]alpha-toxin site. This binding was displaced by perhydrohistrionicotoxin. The carbamoylcholine-stimulated efflux of 22Na+ from the Torpedo vesicles were preserved after alkaline extraction. It is concluded that not only the acetylcholine binding site, but also the local anesthetic binding site, must be associated with the peptides of the cholinergic receptor itself and not that of Mr 43,000. Those peptides remaining after alkaline extraction are also sufficient for permeability control.
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PMID:Acetylcholine and local anesthetic binding to Torpedo nicotinic postsynaptic membranes after removal of nonreceptor peptides. 3 54

The ultrastructural study of liver tissues from 38 patients with type B viral hepatitis consistently showed the presence of hepatitis B core antigen of 21-25 nm size in the liver cell nuclei and to a lesser extent in the cytoplasm. This finding and the demonstration of the tubular form of hepatitis B surface antigen in the proliferative degranulated endoplasmic reticulum constituted the etiologic criterion for the diagnosis of the disease. The double-shelled Dane-like particles were frequently found in association with the tubular form of the surface antigen. The core particles were found in the protoplasmic processes of hepatocytes and this correlated with the immunofluorescent microscopic findings that the antigen may be shed into circulation with the protoplasm. The core antigen was found to resist digestion by various enzymes such as protease, DNase, RNase, phospholipase C, lipase, lysozyme, diastase, neuraminidase and hyaluronidase, all of which did not destroy the immunoreactivity as demonstrated by immunoelectron and immunofluorescent microscopy. Similarly, sodium dodecyl sulfate, Tween 80 and mercaptoethanol also had no effect. The formalin-fixed paraffin-embedded liver tissue sections could be treated with protease to facilitate the immunofluorescent staining for the core antigen in tissue.
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PMID:Structural and immunoreactive characteristics of hepatitis B core antigen. 5 6

Subviral cores have been prepared from the oncornavirus-like particle found in human milks with the use of phospholipase C and ether or Sterox SL. The major protein of these cores has a molecular weight of 27,000 daltons, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This protein is found in the core fractions of reverse transcriptase-positive milks and is absent in negative milks. It is distributed in sucrose gradients only in those fractions containing cores and reverse transcriptase activity. The major core protein of the human milk oncornavirus-like particle is electrophoretically identical to the major core protein of the mouse mammary tumor virus.
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PMID:Identification and isolation of the major core protein from the oncornavirus-like particle in human milk. 6 99

A preparation enriched in junctional complexes, as judged by marker enzymes and electron microscopy, was prepared from rat cerebellum. The junctional complexes were incubated with gamma-amino [14C]butyric acid at 25degreesC for 10 min, using [3H]sucrose as a marker for entrapped space, Total binding was determined in the absence of, and non-specific binding in the presence of, and excess of unlabelled gamma-aminobutyric acid. The difference bewteen the two binding values, i.e. the specific binding, was saturable and reversible, and showed positive cooperativity with a Hill number of about 2. The specific binding was inhibited by N-methylbicuculline, picrotoxinine and imidazole-4-acetic acid, but not by curare, strychnine or L-2,4-diaminobutyric acid. The above compounds had little effect on the non-specipic binding, but addition of ethylenediaminetetraacetic acid decreased non-specific binding by 80%. Trypsin, pronase, phospholipase A2 (EC 3.1.1.4), lysolecithin and sodium dodecyl sulfate decreased binding. Phospholipase C (EC 3.1.4.3) increased the specific binding by 260%. Phospholipids competed with gamma-aminobutyric acid for binding, with phosphatidylethanolamine being more potent than phosphatidylcholine. These results lend support for Watkins' hypothesis that phosphatidylethanolamine competes with gamma-aminobutyric acid for binding to the receptor protein.
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PMID:The effect of phospholipases and proteases on the binding of gamma-aminobutyric acid to junctional complexes of rat cerebellum. 13 18

A method using phospholipase C (PL-C) for removing nonspecific inhibitors (NSI) of rubella virus hemagglutinin is described. PL-C was found to hydrolyze NSI without altering the hemagglutination inhibition (HI) activity of the specific antibody and could be used to remove NSI in the rubella HI test by using formalinized erythrocytes, which resisted the enzymatic action; fresh erythrocytes were lysed by PL-C. The HI test using PL-C treated sera gave true measurements of actual rubella antibody content, and HI titers of PL-C treated sera were identical or equivalent (+/-1 dilution) to those of sera treated with dextran sulfate and CaCl2 (DS-C). Thus, the PL-C method gave results as reproducible and reliable as the DS-C method and was more convenient.
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PMID:Improved rubella hemagglutination inhibtion test: inactivation of non-immunoglobulin hemagglutination inhibitors by phospholipase C. 18 17

Protein composition of cardiac sarcolemmal membranes was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Membranes were observed to contain about 20 polypeptide bands ranging from 18000 to 200 000 dalton mass. Out of these, six bands were prominent and together comprised 57% of the membrane protein. When sarcolemmal membranes, phosphorylated by [gamma-(32)P] ATP in the presence of Ca(2+) or Na+ with and without K+, were fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis at pH 2.4, the band III region (Mr 105 000) of gels was found to contain active sites of monomeric Ca-ATPase and (Na,K)ATPase. Bands I (Mr greater than 200 000), II (Mr 150 000), III (Mr 105 000), and VI (Mr 47 000) were accesible to trypsin; the extent of proteolysis was dependent on the time of exposure to, and the concentration of, trypsin (i.e, ratio of sarcolemmal protein/trypsin). Addition of molar sucrose protected sarcolemmal proteins from the tryptic proteolysis. Calcium transport was reduced by the action of trypsin; the degree of reduction was influenced by the time of exposure of membranes to trypsin as well as the concentration of trypsin. (Mg,Ca)ATPase activity, on the other hand, was elevated moderately at lower concentration and reduced at higher concentration of trypsin. Treatment with phospholipase C cium transport and (Mg,Ca)ATPase activity; electrophoretic patterns were unaffected by this treatment. Addition of lecithin to phospholipase C treated membranes produced a moderate increase in calcium transport. Exposure to Triton X-100 (1%) specifically solubilized three protein bands (Mr90 000, 67 000, and 57 000), whereas exposure to deoxycholate (1%) preferentially solubilized high-molecular-weight proteins, including band III (Mr 105 000); Lubrol-PX (1%) caused nonspecific solubilization of proteins, although the extent of solubilization with Lubrol-PX was considerably less than with either Triton or deoxycholate.
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PMID:Protein analysis of cardiac sarcolemma: effects of membrane-perturbing agents on membrane proteins and calcium transport. 21 4

Cyclic AMP-dependent protein kinases from several mammalian sources inhibit Na+-dependent alpha-aminoisobutyric acid transport by membrane vesicles isolated from 3T3 cells. Evidence is provided that phosphorylation of membrane proteins by the enzyme is responsible for the inhibition. Lysis of the vesicles, or a reduction in the intravesicular volume is not the cause of reduced transport. The cyclic AMP-dependent protein kinase and its catalytic subunit phosphorylate a number of membrane proteins. Most of these proteins are phosphorylated, but to a lesser extent in the absence of protein kinase or cyclic AMP. The phosphorylated proteins remain associated with the membranes during hypotonic lysis treatments, which would be expected to release intravesicular contents and loosely associated membrane proteins. 32P-labeled bands detected on sodium dodecyl sulfate polyacrylamide gels after phosphorylation of membranes by the catalytic subunit of the cyclic AMP-dependent kinase are eliminated by treatment with either pronase or 1 N NaOH, but not by ribonuclease nor by phospholipase C. The stability of the incorporated radioactivity to hot acid and hydroxylamine relative to hot base suggests that most of the 32P from [gamma-32P]ATP is incorporated into protein phosphomonoester linkages.
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PMID:Inhibition of alpha-aminoisobutyric acid transport in membrane vesicles from mouse fibroblasts after phosphorylation by cyclic AMP-dependent protein kinase. 22 60

Heat-stable enterotoxin (ST) produced by porcine strains of enterotoxigenic (ENT+) Escherichia coli has been purified to apparent homogeneity by sequential ultrafiltration, acetone fractionation, preparative gel electrophoresis, diethylaminoethyl Bio-Gel A ion-exchange chromatography, and Bio-Gel P-10 gel filtration. The enterotoxin, purified more than 1,500-fold, exhibited a molecular weight of 4,400, as determined by both sodium dodecyl sulfate-gel electrophoresis and gel filtration. A molecular weight of 5,100, representing 47 residues, was calculated from amino acid analysis data. The amino acid content was distinctive, with an unusually high proportion of cystines and few hydrophobic amino acids. A single amino-terminal residue, glycine, was observed. Purified ST was stable to heating (100 degrees C, 30 min) and did not lose biological activity after treatment with Pronase, trypsin, proteinase K, deoxyribonuclease, ribonuclease, and phospholipase C. Periodic acid oxidation and several organic solvents (acetone, phenol, chloroform, and methanol) had no effect on the biological activity of ST. Further, purified ST was stable to acid treatment at pH 1.0 but lost biological activity at pH values greater than 9.0. Neither lipopolysaccharide nor lipid contamination was evident in purified preparations. A characteristic absorption spectrum was observed during the course of the purification, which shifted from a maximum at 260 nm in crude preparations to 270 nm for the purified toxin. Antiserum obtained from rabbits immunized with ST or ST coupled to bovine serum albumin neutralized the action of the enterotoxin in suckling mice; however, passive hemagglutination and hemolysis titer assays suggested that ST is a poor antigen.
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PMID:Purification and chemical characterization of the heat-stable enterotoxin produced by porcine strains of enterotoxigenic Escherichia coli. 34 81

Purified staphylococcal alpha-toxin (molecular weight approximately 36,000) was mildly digested with trypsin, yielding two components by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A fast-moving component (molecular weight 17,000 +/- 5%) which is relatively resistant to tryptic digestion and a slow-moving component (molecular weight 20,000 +/- 5%) which tends to aggregate. The fast-moving component was highly purified by means of combined procedures of column chromatography on Sephadex G-200 with zone electrophoresis on starch. The purified fast-moving component retained a high degree of lethal toxicity for mouse but lacked hemolytic and dermonecrotic activities, whereas the slow-moving component proved to be a nontoxic polypeptide. The lethal toxic fragment was antigenically active showing partial immunological identity with the parent alpha-toxin and stimulated the formation of antibodies capable of neutralizing the lethal action of alpha-toxin in vivo. Some physical properties and the amino acid composition of the purified lethal toxic fragment have been compared with those of native alpha-toxin.
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PMID:Purification and some properties of a lethal toxic fragment of staphylococcal alpha-toxin by tryptic digestion. 67 58


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