Proteolytic Cleavage of Vitamin K-Dependent Bovine Plasma Protein S by o-Thrombin and Plasma Serine Protease12

It is known that protein S, a vitamin K-dependent plasma protein, isolated from a human source, gives a closely spaced doublet on sodium dodecyl sulfate-polyacryl-amide gel electrophoresis after reduction and that this heterogeneity in molecular size results from a limited proteolysis of protein S mediated by a-thrombin in human species. We found here that a-thrombin also rapidly converted single-chain bovine protein S to a nicked form, which consisted of the NHt-terminal segment containing y-carboxyglutamic acid and the COOH-terminal large segment bridged by a disul-fide linkage(s). These two segments were isolated and chemically characterized after S-alkylation of the nicked protein S. The results suggest that the a-thrombin-catalyzed hydrolysis of protein S probably occurs at a peptide linkage (Arg-Ser) located in the NH,-terminal portion. The conversion of single-chain protein S to the nicked form was also mediated by plasma kallikrein and plasmin, in addition to a-chymotrypsin and trypsin. However, the a-thrombin-catalyzed conversion did not occur when calcium ions were added to the reaction mixture. Protein S is a new vitamin K-dependent protein 4). Although the physiological function of pro