Proteolysis of PAR1 and PAR3 cleavage site mutants by FXa on cells. Cleavage of SEAP-PAR1 (A-B) and SEAP-PAR3 (C-D) mutants by FXa in the presence (A,C) and absence (B,D) of wt-EPCR. SEAP released in the media was determined 1 hour after the addition of FXa and corrected for background and spontaneous SEAP release in the absence of protease (∼2%). The activity of released SEAP was expressed as a percentage of total available SEAP activity on the cells. No SEAP activity was detected on HEK-293 or wt-EPCR HEK-293 cells in the absence of SEAP-PAR1 or SEAP-PAR3. Data points represent mean ± SEM of at least 3 independent experiments with 2 to 3 replicates per experiment. K, Lys; Q, Gln; R, Arg.