Residues flanking scissile bonds in Factor VIII modulate rates of cleavage and proteolytic activation catalyzed by Factor Xa.

Factor Xa (FXa) proteolytically activates Factor VIII (FVIII) by cleaving P1 residues Arg(372), Arg(740), and Arg(1689). The Arg(372) site represents the rate-limiting step for procofactor activation, whereas cleavage at Arg(740) is a fast step. FXa also catalyzes inactivating cleavages that occur on a slower time scale than the activating ones.

Variable contributions of basic residues forming an APC exosite in the binding and inactivation of factor VIIIa.

Basic residues contained in the 39-, 60-, and 70-80-loops of activated protein C (APC) comprise an exosite that contributes to the binding and subsequent proteolytic inactivation of factor (F) VIIIa. Surface plasmon resonance (SPR) showed that WT APC bound to FVIII light chain (LC) and the FVIIIa A1/A3C1C2 dimer with equivalent affinity (Kd = 525 and 546 nM, respectively). These affinity values may reflect binding interactions to the acidic residue-rich a1 and a3 segments adjacent to A1 domain in the A1/A3C1C2 and A3 domain in LC, respectively.