Decreased PS dependence of FVMTTS/Y activity on vesicles and platelets. (A) Activity on vesicles with high PS content was determined with 5pM CHAPS-washed FVWT or FVMTTS/Y and various concentrations of vesicle of composition 15:20:65 (PS:PE:PC). (B) Activity on vesicles with limiting PS was evaluated with vesicles of composition 2:20:78 (PS:PE:PC) using 10pM FV. (C) Activity on vesicles lacking PS was evaluated on vesicles of composition 20:80 (PE:PC) using 50pM FV. FVMTTS/Y showed increasingly higher relative prothrombinase activity with lower vesicle PS content. Concentrations of FXa and prothrombin were 1nM and 1μM, respectively. Data represent the mean ± SEM for 3 experiments, each performed in duplicate. (D) Prothrombinase activity of FVMTTS/Y on unstimulated platelets or platelets incubated for 10 minutes with 10μM thrombin receptor agonist protein was measured in the presence of FXa and prothrombin. Platelets were suspended at 4 × 107/mL, and concentrations of FV and mutant were 20pM; FXa, 25pM; and prothrombin 1μM. Data represent the mean ± SD for 2 experiments, each performed in duplicate. Probabilities were calculated using the Student t test.
