The role of electrostatic interactions in the assembly of the factor X activating complex on both activated platelets and negatively-charged phospholipid vesicles

Factor X was activated by factor IXa on the surface of either activated platelets or artificial lipid vesicles in the presence of different NaCl concentrations. The V-max of reactions using platelets was optimal at physiologic [NaCl] both in the absence and in the presence of factor VIIIa. In contrast, the V,,a, Of reactions using vesicles decreased with increasing [NaCl] in the absence of factor VIIIa, and increased with increasing [NaCl] when cofactor was present, In the absence of factor VIIIa, the EC(50FIX), a although stable to changes in [NaCl] in platelet-supported reactions, was found to increase significantly as [NaCl] increased in vesicle-supported reactions and correlated with the decreased V-max. Thus, in contrast to platelet-supported reactions, enzyme interaction with negatively-charged vesicles was highly dependent upon electrostatic interactions. In the presence of factor VIIIa, the EC(50FIXa) of vesicle-supported reactions decreased with increasing [NaCl], indicating that interactions between FIXa and FVIIIa call increase enzyme affinity when fewer ionic interactions are favored. The EC(50FVIIIa) was insensitive to changes in [NaCl] on both surfaces. The K-mapp derived from platelet-supported titrations of factor X was lowest just above physiological [NaCl], whereas on vesicles K-mapp was minimal at the lowest [NaCl] tested, Thus, the direct interaction of factor X and factor IXa with the artificial lipid surface is highly dependent upon ionic interactions with the negatively-charged polar heads of phospholipids. However, the interaction of factor IXa and factor X with the activated platelet surface must rely both on electrostatic interactions with lipid and on other interactions provided by surface proteins.