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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)

1. Phospholipase C [EC 3.1.4.3] found in the growth medium of Streptomyces hachijoensis was purified about sixty-fold by dialysis and column chromatography on Sephadex G-50. 2. The active fraction was separated by isoelectric focusing into two fractions, phospholipase C-I (pI 6.0) and phospholipase C-II (pI 5.6). 3. Both purified phospholipases C were homogeneous by immunodiffusion and were not differentiated as regards antigencity. 4. Phospholipase C-I had maximal activity at pH 8.0 and the optimal temperature was 50degree. Phospholipase C-I was stable at 50degrees for 30 min and was stable at neutral pH. 5. The activity of phospholipase C-I was inhibited by high concentrations of various detergents such as Triton X-100, sodium, cholate, SDS and was also inhibited by Ca2+, Ba2+, Al3+, and EDTA, but was stimulated by Mg2+, and ethyl ether. 6. The Km value of phospholipase C-I was 0.9 mM, using phosphatidylcholine as a substrate. 7. By the gel filtration procedure, the molecular weights of phospholipase C-I and -II were both determined to be 18,000. 8. Phosphatidylcholine, phosphatidylinositol, cardiolipin, sphingomyelin, and lysophosphatidylcholine were hydrolyzed by phospholipase C-I, but phosphatidylethanolamine and phosphatidylserine were hydrolyzed with difficulty under the same conditions, Phospholipase C-I also hydrolyzed phosphatidic acid.
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PMID:Studies on phospholipases from Streptomyces. III. Purification and properties of Streptomyces hachijoensis phospholipase C. 0 11

1. Guanylate cyclase of washed particles and plasma membranes showed S-shaped progress curves when titrated with either GTP or Mn2+ ions; similar results were obtained with Triton X-100-solubilized enzyme preparation from washed particles. Hill plots of these data revealed multiple metal-nucleotide and free-metal binding sites. 2. Guanylate cyclase of supernatant fractions displayed typical Michaelis-Menten properties when enzyme required excess of (free) Mn2+ (over GTP) for maximal activities; Ka (free Mn2+) was about 0.15-0.25 mM at subsaturating concentrations of GTP. 4 MnATP, MnADP, and MnGDP were found to increase the activities of both particulate and superantant enzyme, when MnGTP concentration was below saturation and free Mn2+ ion concentration was low (less than 100 muM); MnATP (50muM-1 mM) inhibited both these activities at high free Mn2+ concentration (1.5 mM) and inhibition of the particulate enzyme was greater than that of supernatant enzyme. 5. Ca2+ ions stimulated supernatant-enzyme activity; the stimulatory concentration of Ca2+ ions depended on the concentration of Mn2+ and GTP. 6. A modest stimulation of particulate guanylate cyclase by pyrophosphate (0.02-1 mM) was observed; the pyrophosphate effect appeared to be competitive with respect to GTP. At a higher concentration (2 mM), pyrophosphate produced a marked inhibition of particulate enzyme; the nature of inhibitory effect appeared complex. 7. Inorganic salts (e.g. NaCl, KCl, LiBr, NaF) produced inhibition of particulate enzyme; the degree of inhibition of Triton X-100-stimulated activity was less than that of unstimulated activity. 9. Treatment of sarcolemmal or microsomal membranes with either phospholipase C or trypsin decreased, whereas phospholipase A increased, the activity of guanylate cyclase.
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PMID:Properties of particulate, membrane-associated and soluble guanylate cyclase from cardiac muscle, skeletal muscle, cerebral cortex and liver. 1 Aug 91

Intact microsomes isolated from rat liver showed no hexose-6-phosphate dehydrogenase activity, but the enzyme was activated by Triton X-100, deoxycholate, NH4OH, glycine/NaOH, lysophosphatidylcholine, phospholipases A and C, pancreatic lipase and cholesterol esterase, and also by sonic treatment. The enzyme activation by deoxycholate, NH4OH and sonic treatments was solely due to solubilization, while that by phospholipase A appeared to be due to the detergent action of the hydrolysis products. On the other hand, the primary effects of phospholipase C, cholesterol esterase and pancreatic lipase might be accounted for by the partial removal of membrane lipids. The results of washing and trypsin digestion experiments suggested that hexose-6-phosphate dehydrogenase is one of the most firmly bound enzymes among the microsomal proteins. The catalytic properties were the same in the solubilized and the membrane-bound, activated enzymes. Feeding the rats on a high carbohydrate diet altered the extent of enzyme activation by sonication and phospholipase C treatment, suggesting that the microsomal membrane would actually undergo changes in the conformation and/or chemical composition under certain circumstances.
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PMID:Latency of microsomal hexose-6-phosphate dehydrogenase activity. 1 59

The effects of various treatments, which affect membrane structure, on microsomal thiamine diphosphatase and thiamine triphosphatase activities of rat brain, were examined. The treatment of micorosomes at alkaline pH caused a 2-fold activation of the thiamine diphosphatase, this being related to a change in membrane structure which was evidenced by a decrease of the turbidity of the microsomal suspension. Repeated freezing and thawing after hypo-osmotic treatment also increased the activity of microsomal thiamine diphosphatase. In addition, the thiamine diphosphatase activity was enhanced by treatment of the microsomes with phospholipase C or acetone. This lipid depletion resulted in a marked reduction in the apparent Km value of the thiamine diphosphatase with a corresponding loss in heat stability of the enzyme. We found further that brain thiamine diphosphatase was solubilized by Triton X-100. This decreased the phospholipid content in the preparation, but did not affect the apparent Km value and heat stability of the enzyme. In contrast with thiamine diphosphatase, thiamine triphosphatase was inactivated by treatment at alkaline pH or with acetone. However, treatment with phospholipase C did not affect the activity of thiamine triphosphatase.
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PMID:Possible regulation of thiamine diphosphatase activity in rat brain microsomes by lipids. 1 55

The enzyme sphingomyelinase (sphingomyelin phosphorylcholine phosphohydrolase E.C.3.1.4.12) which hydrolyzes sphingomyelin to ceramide (N-acylsphingosine) and phosphorylcholine was identified in the subcellular fractions of pig and human epidermis. The enzyme has an optimum pH of 4.5 to 5 and is activated by Triton X-100 (0.1% w/v). Approximately two-thirds of the enzyme activity in both the pig and human epidermal homogenates was in the soluble subcellular fraction and more than half of the enzyme activity in the subcellular particulate fraction was solubilized by freeze-thawing. The pH optimum suggests that epidermal sphingomyelinase is probably a lysozomal enzyme. The enzymes in both pig and human epidermis exhibited Michaelis-Menten kinetics. The soluble sphingomyelinase in pig epidermis had an apparent Km, 4.5 X 10(-5) M and that in human epidermis an apparent Km 7.7 X 10(-5) M. The pig epidermal sphingomyelinase had no special requirement for either divalent or heavy metal ions and was not inhibited by sulfydryl group-blocking agents but it was moderately inhibited by dithiothreitol. No evidence was found in either pig or human epidermis for the presence of a phospholipase C (E.C.3.1.4.3) which hydrolyzes phosphatidylcholine to diglyceride and phosphorylcholine but there was suggestive evidence of another catabolic pathway for phosphatidylcholine.
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PMID:Sphingomyelinase in pig and human epidermis. 2 35

The effects of a variety of agents on guanylate cyclase activity were tested in broken cell preparations of mammary glands from midpregnant mice. Of the agents tested, only phospholipase A, triton X-100, and an impure egg lysolecithin preparation enhanced the activity of guanylate cyclase in mammary gland homogenates; other agents, including sodium azide and phospholipase C, and purified egg lysolecithin had no effect. Phospholipase A increased the activity of guanylate cyclase in the 150,000 g pellet fractions of mammary gland homogenates, bud did not consistently enhance guanylate cyclase in the 150,000 g supernatant fractions. Phospholipase A did not appear to enhance guanylate cyclase activity by solublizing the enzyme from the 150,000 g pellet. Triton X-100, in contrast, appeared to act by solubilizing guanylate cyclase from the material present in the 150,000 g pellet. Triton X-100 increased by several fold guanylate cyclase activity in the tissue homogenates and the 150,000 g pellets, but did not consistently enhance enzyme activity in the 150,000 g supernatant. Triton X-100 had no effect on the apparent Km of guanylate cyclase.
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PMID:Effects of phospholipase A and triton x-100 on guanylate cyclase activity in mammary gland homogenates from mice. 2 72

Cyclic AMP phosphodiesterase (PDE) in membrane fraction from rat cerebral cortex was activated by Triton X-100, and treatment at alkaline pH and with phospholipase C. These results suggest that membrane PDE exists in a latent form and is influenced by microenvironmental changes within the membrane. Furthermore, the PDE, unlike soluble enzyme, is not influenced by a protein activator and Ca++.
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PMID:Possible regulation of membrane-associated cyclic AMP phosphodiesterase in rat cerebral cortex by lipids. 3 Dec 95

Some parameters of the receptor element from the rat olfactory epithelium are evaluated; it is characterized by high affinity for camphor (KD = 1.5. x 10(-9) M). Triton X-100 has no marked effect on the binding of [3H]camphor. Neither RNAase nor phospholipase C affected [3H]camphor-binding activity. Pronase and trypsin abolished [3H]camphor binding activity by 65 and 40%, respectively. Sulfhydryl reagents decrease the binding of [3H]camphor by a factor of 5--8. The isoelectric point of the receptor solubilized with Triton X-100 is 4.8, as determined by isoelectric focusing. The molecular weight of the receptor as determined by gel electrophoresis is about 120 000. It is proposed that the camphor receptor is a membrane protein containing sulfhydryl groups and playing a key role in olfactory reception.
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PMID:Molecular mechanisms of olfactory reception. IV. Some biochemical characteristics of the camphor receptor from rat olfactory epithelium. 4 3

Benzalkonium chloride (BAC) is a mixture of quaternary benzyldimethylalkylammonium chlorides which was found to inhibit histamine release induced by polyamines (48/80, ATP, bradykinin, curare, guanethidine, polylysine, polymyxin B, poly-THIQ, protamine, stilbamidine or substance P), but not that caused by antigens, concanavalin A, dextran, lonophores (A23187 or X-537A), enzymes (chymotrypsin or phospholipase C), monoamines (dextromethorphan, meperidine or chlorpromazine) or detergents (decylamine, Triton X-100 or a fire ant venom alkylpiperidine). Inhibition by 1.5 and 3 microgram of BAC per ml caused parallel shifts of the 48/80 dose-response curves to the right with no loss of efficacy, indicating that the antagonism was surmountable. Phospholipase C was partially inhibited by BAC, but Triton X-100 also inhibited phospholipase C (but not 48/80), indicating that the inhibition of phospholipase C by BAC was probably a nonspecific, detergent effect. BAC caused histamine release by itself at concentrations over 5 microgram/ml. Heat inactivation (50 degrees C for 15 min) of the mast cells did not prevent this release, suggesting a lytic mechanism for this action. Structure-activity relations studies on various members of the BAC family for their ability to inhibit 48/80-induced histamine release indicated that benzyldimethyltridecylammonium chloride was the most potent.
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PMID:Benzalkonium chloride: selective inhibitor of histamine release induced by compound 48/80 and other polyamines. 9 63

Phospholipase C (phosphatidylcholine choline-phosphohydrolase, EC 3.1.4.E) from Bacillus cereus (IAM-1208) was adsorbed to palmitoyl cellulose from a crude enzyme solution at pH 5--9. The adsorption was not influenced by ionic strength up to 2 M NaCl. The adsorbed enzyme was eluted almost completely by washing the cellulose with a suitable detergent, such as Triton X-100, Adekatol SO-120, Cation DT-205, or sodium deoxycholate. The enzyme was then purified by column chromatography on a palmitoylated textile (palmitoylated gauze) with an overall recovery of 91% and a 467-fold increase in specific activity over that of enzyme in the crude culture supernatant. Subsequent fractionation with acetone and chromatography on a Sephadex G-75 column separated two nearly homogeneous phospholipase C's. The enzyme adsorbed on palmitoyl cellulose was active, although its activity was about one-fourth that of free phospholipase C. Therefore, the enzyme appeared to be adsorbed to the cellulose through a hydrophobic site that was distinct from the catalytic site on the enzyme molecule.
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PMID:Purification of phospholipase C from Bacillus cereus by hydrophobic chromatography on palmitoyl cellulose. 11 Aug 96


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