Characterization of phosphoinositide-specific phospholipase C from human platelets.
Phosphoinositide-specific phospholipase C (PI-PLC) from human platelet cytosol was purified 190-fold to a specific activity of 0.68 mumol of phosphatidylinositol (PI) cleaved/min per mg of protein. It hydrolyses PI and phosphatidylinositol 4,5-bisphosphate (PIP2), but not phosphatidylcholine, phosphatidylserine or phosphatidylethanolamine. The enzyme exhibits an acid pH optimum of 5.5 and has a molecular mass of 98 kDa as determined by Sephacryl S-200 gel filtration. It required millimolar concentrations of Ca2+ for PI hydrolysis, whereas micromolar concentrations are optimal for PIP2 hydrolysis. Mg2+ could substitute for Ca2+ when PIP2, but not PI, was used as the substrate. EDTA was more effective than EGTA in inhibiting the basal PI-PLC activity towards PIP2. Sodium deoxycholate strongly inhibits the purified PI-PLC activity with either PI or PIP2 as substrate. Ras proteins, either alone or in the form of liposomes, have no effect on PI-PLC activity.
Manne
V
V
Department of Molecular Oncology, Roche Institute of Molecular Biology, Nutley, NJ 07110.
Kung
H F
HF
D001792:Blood Platelets / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002852:Chromatography, Ion Exchange; D003840:Deoxycholic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000494:pharmacology*; D013379:Substrate Specificity; D010738:Type C Phospholipases / Q000097:blood* / Q000302:isolation & purification
D010716:Phosphatidylinositols; D003840:Deoxycholic Acid; D008345:Manganese; D010738:Type C Phospholipases; D008274:Magnesium; D002118:Calcium