Factor VII autoactivation proceeds via interaction of distinct protease-cofactor and zymogen-cofactor complexes....

Tissue factor (TF), an integral membrane protein, enhances the feedback activation of factor VII by factor VIIa (factor VII autoactivation). We found that, in contrast to the other known membrane-dependent coagulation activation reactions, TF-dependent factor VII autoactivation occurred preferentially on neutral phospholipid vesicles relative to negatively charged vesicles containing phosphatidylserine. This reaction was best described by a novel mechanism in which the enzyme and substrate are each bound to separate cofactor (TF) molecules. This unusual mechanism of substrate presentation to a membrane-bound protease predicts that the reaction rate will be directly dependent on the surface density, and hence lateral diffusion, of factor VII.TF and factor VIIa.TF complexes, obeying obligatorily two-dimensional enzyme kinetics. This prediction was confirmed, yielding a two-dimensional second-order rate constant (k2D) of 4.9 (+/- 0.8) x 10(6) m2 mol-1 s-1. Since intact cells normally sequester acidic phospholipids away from the outer leaflet of the plasma membrane, this reaction mechanism should permit factor VII autoactivation to predominate on unactivated/undamaged cell surfaces when other clotting reactions are dormant.