title: Effect of L-alpha-phosphatidylinositol on a vascular smooth muscle Ca2+-dependent protease. Reduction of the Ca2+ requirement for autolysis.
abstract: Vascular smooth muscle contains large amounts of a Ca2+-dependent protease. Similar to a Ca2+-dependent protease previously purified from chicken gizzard smooth muscle (Hathaway, D. R., Werth, D. K., and Haeberle, J. R. (1982) J. Biol. Chem. 257, 9072-9077), the mammalian vascular muscle protease is a heterodimer consisting of 76,000- and 30,000-dalton subunits (IIa). The enzyme can undergo autolysis in the presence of Ca2+ to produce a smaller species consisting of 76,000- and 18,000-dalton subunits (IIb). Autolysis greatly reduces the Ca2+ dependence of catalytic activity. The autolytic species, IIb, was approximately 23-fold more sensitive to Ca2+ (K0.5 = 39 microM) than the native enzyme, IIa (K0.5 = 891 microM). In this communication, we report that phosphatidylinositol and to a lesser extent one metabolic derivative, dioleoylglycerol, stimulate autolysis of the vascular Ca2+-dependent protease by reducing the Ca2+ for autolysis from K0.5 = 680 microM in the absence of lipid to K0.5 = 87 microM in the presence of both phosphatidylinositol and dioleoylglycerol. Moreover, the reduction in the Ca2+ requirement for autolysis produced by the phosphatidylinositol was antagonized by the phospholipid-binding drug, trifluoperazine. In addition, the effect of phosphatidylinositol was specific for autolysis, and none of several phospholipids or derivatives tested altered the Ca2+ dependence or maximal rate for protein degradation of the autolytic product, IIb. Our results suggest that autolysis may be an important initial step in the activation of the Ca2+-dependent protease in vascular smooth muscle and that this step may be regulated by a combination of Ca2+ and phosphatidylinositol.
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D002154:Calpain; D002417:Cattle; D004075:Diglycerides / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D010716:Phosphatidylinositols / Q000494:pharmacology*
chemical_list: D004075:Diglycerides; D010716:Phosphatidylinositols; D010450:Endopeptidases; D002154:Calpain; D002118:Calcium; C013965:diolein