Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Pure or impure C-type phospholipases hydrolysed rat liver microsomal phosphatides in situ at 5 degrees or 37 degrees C. At 5 degrees C mean hydrolysis of total phospholipids was 90% by Bacillus cereus and 75% by Clostridium perfringens (Clostridium welchii) C-type phospholipases. 2. Four degrees of inhibition of glucose 6-phosphatase (D-glucose 6-phosphate phosphohydrolase; EC 3.1.3.9) resulted. (a) At 37 degrees C inhibition was virtually complete and apparently irreversible. (b) At 5 degrees C phospholipase C inhibited 50-87% of the activity expressed by intact control microsomal fractions. (c) Bovine serum albumin present during delipidation alleviated most of this inhibition: at 5 degrees C phospholipase C plus bovine serum albumin inhibited by 0-35% (mean 18%):simultaneous stimulation by the destruction of its latency seems to offset glucose 6-phosphatase inhibition, sometimes completely. (d) If latency was first destroyed, phospholipase C plus bovine serum albumin inhibited 30-50% of total glucose 6-phosphatase activity at 5 degrees C. Only this inhibition is likely largely to reflect the lower availability of phospholipids, essential for maximal enzyme activity, as it is virtually completely reversed by added phospholipid dispersions. Co-dispersions of phosphatidylserine plus phosphatidylcholine (1:1, w/w) were especially effective but Triton X-100 was unable effectively to restore activity. 3. Considerable glucose 6-phosphatase activity survived 240min of treatment with phospholipase C at 5 degrees C, but in the absence of substrate or at physiological glucose 6-phosphate concentrations the delipidated enzyme was completely inactivated within 10min at 37 degrees C. However, 80mM-glucose 6-phosphate stabilized it and phospholipid dispersions substantially restored thermal stability. 4. It is concluded that glucose 6-phosphatase is at least partly phospholipid-dependent, and complete dependence is not excluded. For reasons discussed it is impossible yet to be certain which phospholipid class(es) the enzyme requires for activity.
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PMID:Inhibition of glucose 6-phosphatase by pure and impure C-type phospholipases. Reactivation by phospholipid dispersions and protection by serum albumin. 16 86

Pathogenic staphylococci secrete a number of exotoxins, including alpha-toxin. alpha-Toxin induces lysis of erythrocytes and liposomes when its 3S protein monomers associate with the lipid bilayer and form a hexomeric transmembrane channel 3 nm in diameter. We have used alpha-toxin to render rat hepatocytes 93-100% permeable to trypan blue with a lactate dehydrogenase leakage less than or equal to 22%. Treatment conditions included incubation for 5-10 min at 37 degrees C and pH 7.0 with an alpha-toxin concentration of 4-35 human hemolytic U/ml and a cell concentration of 13-21 mg dry wt/ml. Scanning electron microscopy revealed signs of swelling in the treated hepatocytes, but there were no large lesions or gross damage to the cell surface. Transmission electron microscopy indicated that the nucleus, mitochondria, and cytoplasm were similar in control and treated cells and both had large regions of well-defined lamellar rough endoplasmic reticulum. Comparisons of the mannose-6-phosphatase and glucose-6-phosphatase activities demonstrated that 5-10 U/ml alpha-toxin rendered cells freely permeable to glucose-6-phosphate, while substantially preserving the selective permeability of the membranes of the endoplasmic reticulum and the functionality of the glucose-6-phosphatase system. Thus, alpha-toxin appears to have significant potential as a means to induce selective permeability to small ions. It should make possible the study of a variety of cellular functions in situ.
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PMID:Permeabilization of rat hepatocytes with Staphylococcus aureus alpha-toxin. 298 73

Interrelationships between the catalytic behavior of glucose-6-phosphatase and the structure of rat-liver microsomal membranes were investigated. 2. Rabbit anti-microsomal serum completely inhibited glucose-6-phosphate hydrolysis in detergent-modified microsomes but showed no inhibitory effect on the enzyme activity of intact or mechanically disrupted vesicles. 2. Controlled proteolysis of intact microsomes using carboxypeptidase A and/or aminopeptidase M largely denatured enzymes situated on the outer surface of the microsomal vesicles such as monodehydroascorbate reductase and cytochrome c reductase. However, it did not affect the glucose-6-phosphatase activity at all, which remained in a latent state within the membrane. 3. Temperature studies on glucose-6-phosphatase have revealed that only the enzyme activity of intact microsomes exhibited a nonlinear Arrhenius plot, whereas detergent-modified microsomes showed a linear temperature response. 4. Treatment of microsomes with phospholipase C and toluene-2,4-diisocyanate resulted in an apparent loss of about 65% and 85% of the original glucose-6-phosphatase activity and was closely correlated with hydrolysis and chemical modification of phosphatidylethanolamine, respectively. These apparent inactivations could be reversed by addition of Triton X-114 alone without any phospholipid supplementation. These observations indicate that glucose-6-phosphatase is buried within the microsomal membrane, not exposed on either side. They also suggest that phospholipids are involved in the glucose-6-phosphate transport mechanism.
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PMID:Investigations on the possible involvement of phospholipids in the glucose-6-phosphate transport system of rat-liver microsomal glucose-6-phosphatase. 624 79

The role of phospholipids in the glucose-6-phosphatase system, including glucose-6-P phosphohydrolase and glucose-6-P translocase, was studied in rat liver microsomes by using phospholipases C and detergents. In the time course experiments on detergent exposure, the maximal activation of glucose-6-P phosphohydrolase varied according to the nature of the detergent used. On treatment of microsomes with phospholipase C of C. perfringens, the activity of glucose-6-P phosphohydrolase without detergent (i.e. without rupture of translocase activity) was gradually decreased with the progressive hydrolysis of phosphatidylcholine and phosphatidylethanolamine on the microsomal membrane, and was restored by incubation of these microsomes with egg yolk phospholipids. The extent of decrease in this phosphohydrolase activity in the detergent-exposed microsomes (with rupture of translocase activity) also varied depending on the detergent used (Triton X-114 or taurocholate). When 66% of the phosphatidylinositol on the membrane was hydrolyzed by phosphatidylinositol-specific phospholipase C of B. thuringiensis, the inhibition of glucose-6-P phosphohydrolase activity without detergent was very small. Although the inhibition of enzyme activity with detergent was apparently greater than that without detergent, the enzyme activity was stimulated by the breakdown of phosphatidylinositol when the enzyme activity was measured at lower concentration (0.5 mM) of substrate, glucose-6-P. The latency of mannose-6-P phosphohydrolase, a plausible index of microsomal integrity, remained above 70% after the hydrolysis of phosphatidylcholine, phosphatidylethanolamine, or phosphatidylinositol. The results show that the glucose-6-phosphatase system requires microsomal phospholipids for its integrity, suggesting that there exists a close relation between phosphatidylinositol and glucose-6-P translocase.
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PMID:Studies on the interactions between phospholipids and membrane-bound enzymes in microsomes. Effects of phospholipases C on the glucose-6-phosphatase system of rat liver microsomes. 630 98

Through kinetic analysis, the relationships between the glucose-6-phosphatase system and constituent phospholipids were studied in rat liver microsomes. When phosphoglycerides such as phosphatidylcholine and phosphatidylethanolamine on the microsomal membrane were hydrolyzed by phospholipase C of C. perfringens, the activities of glucose-6-P phosphohydrolase and glucose-6-P:glucose phosphotransferase both decreased with or without subsequent exposure to taurocholate. In these cases, the Michaelis constants (Km) for glucose-6-P were increased, concomitant with the decrease in the maximal velocities (Vmax) for glucose-6-P hydrolysis. On exposure to taurocholate, the apparent Km for glucose of phosphotransferase was decreased. When phosphatidylinositol was hydrolyzed by phosphatidylinositol-specific phospholipase C of B. thuringiensis, the activities of phosphohydrolase and phosphotransferase were both decreased on exposure to taurocholate. In this case, the value of Vmax of phosphohydrolase was decreased and that of Km for glucose-6-P was slightly decreased, while the apparent Km for glucose of phosphotransferase was increased. Without exposure to detergent, the activities of phosphohydrolase and phosphotransferase both decreased at glucose-6-P concentrations higher than 10 mM. However, at a concentration lower than 1 mM, the activity of phosphohydrolase became higher than that of the control, and Vmax and Km for glucose-6-P were decreased. A similar tendency was also observed in microsomes where membranous phosphatidylinositol was hydrolyzed, when they were treated with DIDS (an anion-transport inhibitor). From these results, it is concluded that the activity of glucose-6-phosphatase is greatly influenced by changes of the phospholipids on the microsomal membrane, and the activity of glucose-6-P translocase is stimulated by the breakdown of phosphatidylinositol.
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PMID:Studies on the interactions between phospholipids and membrane-bound enzymes in microsomes. Effects of phospholipases C on kinetic properties of the glucose-6-phosphatase system in rat liver microsomes. 630 99

Two mutations, R69D and K115E, converted a bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) to a phosphatase with much higher specific activity toward glucose-6-phosphate than inositol-1-phosphate. PI-PLC single mutations R69D and K115E can cleave PI but lack any demonstrable phosphatase activity. The bacterial PI-PLC has no sequence homology with known glucose-6-phosphatase enzymes, which need His, Arg, and negatively charged residues (Asp or Glu) at the active site. The change in chemical reaction and substrate specificity can be rationalized by energy minimization of the mutant with I-1-P or G-6-P bound.
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PMID:Mutation of two active-site residues converts a phosphatidylinositol-specific phospholipase C to a glucose phosphatase. 1474 54

Fatty acids serve vital functions as sources of energy, building materials for cellular structures, and modulators of physiological responses. Therefore, this study examined the effect of linoleic acid on glucose production and its related signal pathways in primary cultured chicken hepatocytes. Linoleic acid (double-unsaturated, long chain) increased glucose production in a dose (> or =10(-4) M)- and time (> or =8 h)-dependent manner. Both oleic acid (monounsaturated, long chain) and palmitic acid (saturated, long chain) also increased glucose production, whereas caproic acid (saturated, short chain) failed to increase glucose production. Linoleic acid increased G protein-coupled receptor 40 (GPR40; also known as free fatty acid receptor-1) protein expression and glucose production that was blocked by GPR40-specific small interfering RNA. Linoleic acid increased intracellular calcium concentration, which was blocked by EGTA (extracellular calcium chelator)/BAPTA-AM (intracellular calcium chelator), U-73122 (phospholipase C inhibitor), nifedipine, or methoxyverapamil (L-type calcium channel blockers). Linoleic acid increased cytosolic phospholipase A(2) (cPLA(2)) phosphorylation and the release of [(3)H]-labeled arachidonic acid. Moreover, linoleic acid increased the level of cyclooxygenase-2 (COX-2) protein expression, which stimulated the synthesis of prostaglandin E(2) (PGE(2)). The increase in PGE(2) production subsequently stimulated peroxisome proliferator-activated receptor (PPAR) expression, and MK-886 (PPAR-alpha antagonist) and GW-9662 (PPAR-delta antagonist) inhibited glucose-6-phosphatase and phosphoenolpyruvate carboxykinase. In addition, linoleic acid-induced glucose production was blocked by inhibition of extracellular and intracellular calcium, cPLA(2), COX-2, or PPAR pathways. In conclusion, linoleic acid promoted glucose production via Ca(2+)/PLC, cPLA(2)/COX-2, and PPAR pathways through GPR40 in primary cultured chicken hepatocytes.
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PMID:Linoleic acid stimulates gluconeogenesis via Ca2+/PLC, cPLA2, and PPAR pathways through GPR40 in primary cultured chicken hepatocytes. 1884 27