| Abstract
| - Factor VIIa, in complex with tissue factor (TF), is the serine protease responsible for initiatingthe clotting cascade. This enzyme complex (TF/VIIa) has extremely restricted substrate specificity,recognizing only three previously known macromolecular substrates (serine protease zymogens, factorsVII, IX, and X). In this study, we found that TF/VIIa was able to cleave multiple peptide bonds in thecoagulation cofactor, factor V. SDS−PAGE analysis and sequencing indicated the factor V was cleavedat Arg679, Arg709, Arg1018, and Arg1192, resulting in a molecule with a truncated heavy chain and an extendedlight chain. This product (FVTF/VIIa) had essentially unchanged activity in clotting assays when comparedto the starting material. TF reconstituted into phosphatidylcholine vesicles was ineffective as a cofactorfor the factor VIIa cleavage of factor V. However, incorporation of phosphatidylethanolamine in the vesicleshad little effect over the presence of 20% phosphatidylserine. FVTF/VIIa was as sensitive to inactivation byactivated protein C (APC) as thrombin activated factor V as measured in clotting assays or by the appearanceof the expected heavy chain cleavage products. The FVTF/VIIa could be further cleaved by thrombin torelease the normal light chain, albeit at a significantly slower rate than native factor V, to yield a fullyfunctional product. These studies thus reveal an additional substrate for the TF/VIIa complex. They alsoindicate a new potential regulatory pathway of the coagulation cascade, i.e., the production of a form offactor V that can be destroyed by APC without the requirement for full activation of the cofactor precursor.
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