Figure 7.

Purified FVa Leiden degradation and clotting activity of APC derivatives in protein S-deficient plasma. (A) The degradation of FVa Leiden (2.5 nM) by increasing concentrations of APC-WT (O) and APC-G74S (●) was carried out on PC/PS vesicles (25 μM) in TBS/Ca²⁺ in a 96-well assay plate. Following 10 min incubation at room temperature, the remaining cofactor activity of FVa Leiden was determined by a prothrombinase assay (0.5 nM FXa and 500 nM prothrombin for 1 min) as described in Materials and methods. (B) The same as A, except that the APC concentration dependence of FVa Leiden degradation was carried out in the presence of protein S (110 nM) for 1 min. (C) The anti-clotting activities of APC-WT (O) and APC-G74S (●) were determined in normal plasma by an aPTT assay as a function of increasing concentrations of APC at 37 °C as described in Materials and methods. (D) The same as panel C except that the anticoagulant activities of the proteases were evaluated in protein S-deficient plasma.