Hemextin AB complex--a snake venom anticoagulant protein complex that inhibits factor VIIa activity

Snake venom is a veritable gold mine of bioactive molecules, capable of binding to a wide variety of pharmacological targets, including the blood coagulation cascade. Here, we report the isolation and characterization of two synergistically acting anticoagulant three-finger proteins, hemextin A and hemextin B, from the venom of Hemachatus haemachatus (African Ringhals cobra). Hemextin A but not hemextin B exhibits mild anticoagulant activity. However, hemextin B interacts with hemextin A and forms a complex (hemextin AB complex), and synergistically enhances its anticoagulant potency. Prothrombin time assay showed that these two proteins form a 1:1 complex. Using a 'a dissection approach', we found that hemextins A and AB complex prolong clotting by inhibiting extrinsic tenase activity. Further studies showed that hemextin AB complex potently inhibits the proteolytic activity of factor VIIa (FVIIa) and its complexes. Kinetic studies showed that hemextin AB complex is a non-competitive inhibitor of FVIIa-soluble tissue factor proteolytic activity with a K(i) of 25 nM. Hemextin AB complex is the first reported natural inhibitor of FVIIa that does not require either tissue factor or factor Xa scaffold to mediate its inhibitory activity. Molecular interactions of hemextin AB complex with FVIIa/tissue factor-FVIIa may provide a new paradigm in the search for anticoagulants inhibiting the initiation of blood coagulation.