A number of biologically important proteolytic enzymes, particularly those involved in the reactions of blood coagulation and fibrinolysis, have the ability to hydrolyze synthetic esters of the basic amino acids, arginine, and lysine. Though synthetic amino acid esters have been used for the study of the reaction kinetics of these enzymes and for assay purposes, relatively little has been done to characterize these enzymes on the basis of their differing abilities to hydrolyze synthetic substrates. Utilizing a variety of comparably substituted arginine and lysine esters, observations were made on the reaction kinetics of several basic amino acid esterases; significant differences were noted among these enzymes, sufficient to establish their separate identity. Further characterization of the reaction of thrombin with synthetic substrates revealed that this enzyme readily hydrolyzes various substituted lysine esters as well as arginine esters. The highest Vmax was observed for the throm-bin-carbobenzoxy-lysine methyl ester reaction but the greatest affinity was observed with benzoyl-arginine methyl ester. The data also clearly establish enzymatic differences between human and bovine thrombin, and between the latter and bovine plasma thrombokinase.
