The action of factor Xa, thrombin and trypsin on human factor II

The proteolytic action of human and bovine Factor Xa, bovine thrombin and bovine pancreatic trypsin Factor II at pH 7.5 and 25°C was monitored by sodium dodecylsulfate gel electrophoresis and thrombin assays. Purified human and bovine Factor Xa, and trypsin, were found to activate Factor II to thrombin. The conversion of Factor II to thrombin by either Factor Xa or trypsin was found to proceed through two thrombogenic intermediates. The reaction pathway appears to be sequential in that the Factor II (75 000 daltons) is first cleaved to a 55 000-dalton thrombogenic product (Intermediate 1) and a 25 000-dalton non-thrombogenic product (Fragment 1). Intermediate 1 is subsequently converted to an inactive 37 000-dalton thrombogenic protein (Intermediate 2) and a 16 000-dalton protein (Fragment 2). Intermediate 2 is finally converted to an active 37 000-dalton thrombin (α-thrombin). Purified bovine thrombin readily converted Factor II to Intermediate 1 and Fragment 1, but possessed little capacity to catalyze subsequent cleavages to produce active thrombin. The ability of thrombin to cleave Factor II was entirely obviated in the presence of hirudin. Under the conditions of the incubation, the maximum thrombin yield obtainable by Factor Xa or trypsin activation was 50% when compared to the two-stage potential thrombin.