INTERACTIONS OF FACTOR-XIII WITH FIBRIN AS SUBSTRATE AND COFACTOR

Factor XIIIa (a2') is a homodimeric transglutaminase that is formed via limited alpha-thrombin-catalyzed proteolysis of the platelet (a2) or plasma (a2b2) factor XIII zymogen in a reaction that results in proteolytic removal of a 37-aminoacyl residue peptide from the N-terminus of the a chains and exposure of the active-site thiol group in the resulting a' chains of factor XIIIa. In this study, we characterized interactions of factor XIII and factor XIIIa with fibrin, a natural substrate for factor XIIIa and a cofactor for the alpha-thrombin-catalyzed activation of plasma factor XIII. The carbamylmethyl derivatives of the active-site thiol group of platelet factor XIII (CMa2) and factor XIIIa (CMa2') were prepared, and their interactions with fibrin were measured. The enzyme-like derivative (CMa2') which contained nicked a' chains bound more tightly to fibrin (K(d) = 2.1-mu-M) than did CMa2 (K(d) = 14-mu-M), the platelet zymogen-like derivative with intact a chains, but the binding of each was weaker than the binding of plasma factor XIII zymogen (a2b2) to fibrin (K(d) = 0.20-mu-M) under the same conditions. Saturation of fibrin with plasma factor XIII zymogen (a2b2) did not affect the binding of CMa2' to fibrin, suggesting that the plasma factor XIII zymogen (a2b2) and the active-site-modified form of factor XIIIa (CMa2') bind to separate, noninteracting sites of fibrin. In contrast to its promoting effect on the activation of plasma factor XIII, fibrin did not promote AP release from platelet factor XIII at concentrations of fibrin high enough to ensure significant binding of platelet factor XIII by fibrin, suggesting that the b chains in plasma factor XIII determine not only the affinity of a2b2 for fibrin but also the geometric disposition of a2b2 in the termolecular thrombin-fibrin-a2b2 complex that is involved in fibrin promotion of alpha-thrombin-catalyzed activation of plasma factor XIII. An analysis of the effects of fibrinogen on the rate of release of b chains and concomitant exposure of the active-site thiol group in alpha-thrombin-cleaved plasma factor XIII (a2'b2) yielded a value of 2.3-mu-M for the equilibrium constant for dissociation of the fibrinogen-a2'b2 complex. The observation that the intact zymogen a2b2 did not antagonize the interaction of a2'b2 With fibrinogen suggested that the proteolytic conversion of a2b2 to a2'b2 results in altered interactions with fibrinogen.