Binding of recombinant human protein S to phospholipid vesicles.

PE/PC/PS 40:40:20 phospholipid vesicles were coated onto the wells of microtiter plates and blocked with 3% (wt/vol) BSA. (A) Dialyzed recombinant protein S (0 to 26 nM) was incubated in the wells for 2 hours at 37°C in a buffer containing 3 mM CaCl₂, and bound protein S was detected with a polyclonal anti–human protein S peroxidase-conjugated antibody (see text). (B) Dialyzed recombinant protein S (10 nM) was incubated in the wells for 2 hours at 37°C in a buffer containing various concentrations of CaCl₂ (0 to 5mM), and bound protein S was detected with a polyclonal anti–human protein S peroxidase-conjugated antibody (see text). In both panels, results are expressed as mean A_492nm of 3 experiments ± SD (in many cases the SDs are so small they cannot be visualized) as a proportion of the maximum (designated 100%). ▪ indicates wild-type protein S; ●, protein S with Gly11Asp; ▴, protein S with Thr37Met; ⋄, wild-type protein S assessed in a buffer that contained no Ca²⁺ and was instead supplemented with 5 mM EDTA.