Activation of a Proteolytic System by a Membrane Lipoprotein: Mechanism of Action of Tissue Factor

Abstract

One pathway of blood coagulation, the extrinsic system, is initiated by a specific interaction between tissue factor, which is a membrane lipoprotein, and factor VII, one of the plasma coagulation factors. Factor VII was prepared from bovine plasma by adsorption onto and elution from BaSO(4). The eluate was chromatographed on DEAE-Sephadex and purified by preparative disc-gel electrophoresis. Factor VII complexed with purified bovine-brain tissue factor and, when eluted from the complex, factor VII had a greater mobility in acrylamide-gel electrophoresis in the presence of sodium dodecyl sulfate, i.e., it had a reduced molecular weight. Factor VII was also cleaved in the presence of orthophenanthroline, an inhibitor of the peptidase activity of tissue factor. Prior treatment of factor VII with diisopropylphosphorofluoridate, however, completely blocked its cleavage and the development of coagulant activity, although factor VII treated with diisopropylphosphorofluoridates complexed equally well as the native protein with tissue factor. Factor VII in whole bovine plasma was also inhibited by the drug. Factor VII labeled with [(32)P]diisopropylphosphorofluoridate and radioautographed after electrophoresis in gels showed two major components, only one of which was labeled. We conclude that tissue factor initiates blood coagulation by facilitating a proteolytic attack within the factor VII "complex." The most likely mechanism is proteolysis of one form of factor VII by the diisopropylphosphorofluoridate-sensitive enzyme, although additional intramolecular proteolysis may be involved.