Determination of the kinetic constants of tissue factor factor VII factor VIIIA and antithrombin/heparin using surface plasmon resonance

Association of factor Wa (FVIIa) with tissue factor (TF) is generally believed to be a critical step in the initiation of blood coagulation. In this study the rate constants for the complex formation and dissociation between FVII/VIIa and TF were studied in real time using surface plasmon resonance, the BIAcore(TM) technique. Employing this technique k(a) and k(d) were determined and yielded a K-D of 1 nM for FVII and 0.5 nM for FVII, respectively. The association and dissociation between antithrombin (AT) and the FVIIa/TF complex was also studied. In the presence of heparin, AT was bound to the FVIIa/TF complex in a dose-dependent manner with k(a) of 2 x 10(3) M(-1) s(-1). The binding of AT to FVIIa/TF increased the dissociation of FVIIa from TF about 20-fold. Copyright (C) 1997 Elsevier Science Ltd.