Interaction of prothrombin with factor Va-phospholipid complexes.

Abstract

The effects of factor Va and the phospholipid-binding fragment of factor Va [factor Va light chain (LC), Mr 80000] on the binding of prothrombin, factor X, and factor Xa to phospholipid vesicles are reported. Equilibrium binding experiments were performed that utilized large-volume vesicles, which can be removed from the bulk solution by centrifugation. Factor Va decreased the dissociation constant of the prothrombin-phospholipid complex 50-fold, from 2.0 X 10(-7) M to 4.0 X 10(-9) M. For the factor X-phospholipid complex the decrease was 60-fold (1.8 X 10(-7) M to 3.0 X 10(-9) M) and for factor Xa, 160-fold (1.6 X 10(-7) M to 1.0 X 10(-9) M). The ratios of moles of protein bound to moles of total added factor Va at saturation of phospholipid-bound factor Va indicate an 1:1 stoichiometric complex of either factor Xa, factor X, or prothrombin and phospholipid-bound factor Va. In the presence of factor Va LC, the dissociation constants of factor Xa- and prothrombin-phospholipid complexes were increased, while the maximal protein-binding capacities of the vesicles were not affected by factor Va LC. The data suggest a competitive interaction between factor Xa and factor Va LC binding as well as between prothrombin and factor Va LC binding at the phospholipid surface. From this, it is concluded that the phospholipid-binding fragment of factor Va alone does not serve as the binding site for interactions of factor Xa and prothrombin with factor Va.