Influence of the gamma-carboxyglutamic acid-rich domain and hydrophobic stack of factor VIIa on tissue factor binding.

Des (1-38)- and des(1-44)-factor VIIa (fVIIa) were inhibited with Phe-Phe-Arg-chloromethyl ketone (FFR-cmk). Des(1-38)-FFR-fVIIa inhibited tissue factor (TF)-enhanced fVIIa amidolytic activity with an IC50 value of 15 nM, whereas 3- and 6-fold higher values were obtained with des(1-44)-FFR-fVIIa using soluble and full-length TF, respectively. The value for FFR-fVIIa was 8-10 nM. Clotting time was prolongated with IC50 values for inactivated des(1-38)- and des(1-44)-fVIIa of 50 nM and 1 mu M, respectively, whereas FFR-fVIIa yielded a value of 10 nM. From binding experiments in a BIA-core instrument, dissociation constants for the complexes between TF1-218 and fVIIa, des(1-38)-fVIIa and des(1-44)-fVIIa of about 3, 20 and 70 nM, respectively, could be estimated.