Extensive structural and functional studies have revealed molecular details of the tissue factor (TF)-mediated regulation of Factor (F)VIIa activity required for the initiation of blood coagulation. In contrast, the structural basis of TF binding to zymogen factor (F)VII remains limited. Here we present, for the first time, the crystal structure of full-length zymogen FVII (S195A) in complex with soluble TF (sTF), devoid of any inhibitors. In addition, we have employed isothermal titration calorimetry (ITC) and molecular dynamics (MD) simulations to elaborate on the findings from the crystal structures.
The crystal structure reveals several unanticipated features with the 170-loop, activation loops 1-3, and the activation peptide showing a high degree of mobility or completely missing electron density (2Fo-Fc) of key amino acids, despite the presence of sTF. The high degree of disorder in FVII, in presence of sTF, was confirmed by ITC experiments as the entropy penalty term for sTF binding to FVII was 2-fold lower compared to that of FVIIa. Interestingly, the stark difference in the underlying thermodynamic basis of TF binding to the two forms, leads to a 10-fold compromise in sTF binding affinity for FVII. The MD simulations indicated that the different surface loops are indeed very flexible and no clear conformity between the electron density and simulated density trace could be found.
The data presented here indicates that sTF binding does not constrict FVII in a single conformation which may be a key requirement in the activation of FVII. Since the binding epitope of TF in zymogen FVII and FVIIa is similar, it is likely that the N-terminus insertion in FVIIa provides further favourable interactions leading to a 10-fold improved affinity for FVIIa. Our combined studies fill an important gap in our understanding of the interaction between zymogen FVII and TF.