Factor VIIIa A2 subunit residues 558-565 represent a factor IXa interactive site.

摘要: Factor VIIIa is a non-covalent heterotrimer of A1, A2, and A3-C1-C2 subunits. Previously, we speculated that the central portion A2 subunit, in around activated protein C-sensitive bond at Arg562-Gly (Fay, P. J., Smudzin, T.M., Walker, F.J. (1991) J. Biol. Chem. 266, 20139-20145), important for macromolecular interactions within factor Xase enzyme complex. A peptide corresponding to VIII residues 558-565, SVDQRGNQ designated FVIII558-565, was chemically synthesized inhibited Xa generation purified system with an apparent KI 105 microM. Tryptic cleavage FVIII558-565 eliminated its inhibitory activity, whereas scrambled sequence version possessed < 30% activity native version. Overlapping peptides FVIII556-564 FVIII561-569 were also confirmed importance scissile functional Xase. Kinetic analysis revealed peptide-mediated inhibition non-competitive respect X. However, increasing IXa concentration overcame observed inhibition. Furthermore, IXa-dependent enhancement reconstituted from isolated A1/A3-C1-C2 dimer plus subunit. Isolated heavy chain (contiguous A1-A2 domains) cleaved Arg336 by equimolar reaction phospholipid-independent. No proteolysis A1 subunit similar reaction. These results indicate delineated 558-565 contributes interaction cofactor protease this essential intrinsic activity. blocks both capacity stabilize labile suggest latter property physiologic significance.