Enhancement of TFPIα in the inhibition of FXa by WT protein S and protein S alanine composite variants. FXa activity (0.5 nM) was followed in real-time through cleavage of S-2765 (200 µM) at 405 mm in the presence of 25 μM phospholipids, presence or absence of TFPIα (2.5 nM), and increasing concentrations (0-160 nM) of WT protein S (A), LG1A (B), LG1B (C), or LG1C (D). Results from representative experiments are shown (n = 3-4). (E) The initial velocity (V₀) was calculated for each curve and plotted against protein S concentration. Results are given as mean ± SD and are expressed as a percentage of the V₀ for TFPIα alone (n = 3-4). The EC₅₀ values of TFPIα enhancement by WT protein S and its variants were derived and are presented as part of the figure. N.M., not measurable; PS, protein S.