Thrombin- and nucleotide-activated phosphatidylinositol 4,5-bisphosphate phospholipase C in human platelet membranes.
Thrombin, nucleotides, and chelators elicited a phosphatidylinositol 4,5-bisphosphate (PtdIns-P2) phospholipase C activity that was associated with human platelet membranes. Both alpha- and gamma-thrombin enhanced phospholipase C activity, whereas active site-inhibited alpha-thrombin did not stimulate PtdIns-P2 hydrolysis. PtdIns-P2 phospholipase C was also activated by nucleoside triphosphates, citrate, EDTA, and NaF. Magnesium was an inhibitor of PtdIns-P2 hydrolysis stimulated by nucleotides and chelators. Only PtdIns-P2 was degraded by the phospholipase C activated by alpha-thrombin, nucleotides, and chelators. The soluble fraction phospholipase C activity was also stimulated at low protein concentrations by nucleotides; however, soluble fraction phospholipase C activity cleaved both PtdIns-P2 and phosphatidylinositol 4-phosphate and was inhibited by chelators, suggesting the presence of a different enzyme in this compartment. The pH optimum for the membrane-associated phospholipase C in the presence of alpha-thrombin or nucleotides was 6.0, and the PtdIns-P2 phospholipase C was inhibited by neomycin and high detergent concentrations. Guanine nucleotides did not synergistically activate phospholipase C in the presence of alpha-thrombin. The characteristics of the membrane-associated PtdIns-P2 phospholipase C suggest that this enzyme is involved in platelet activation by the low-affinity alpha- or gamma-thrombin-dependent pathway.
D001792:Blood Platelets / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002458:Cell Fractionation; D002462:Cell Membrane / Q000201:enzymology / Q000648:ultrastructure; D004492:Edetic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000097:blood*; D012265:Ribonucleotides / Q000494:pharmacology*; D013873:Thionucleotides / Q000494:pharmacology*; D013917:Thrombin / Q000502:physiology*
D012265:Ribonucleotides; D013873:Thionucleotides; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D004492:Edetic Acid; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D013917:Thrombin; D008274:Magnesium; D002118:Calcium