Activation of blood coagulation factor XIII and interactions of factor XIII with fibrinogen and fibrin.

The kinetics of activation of platelet factor XIII $(a\sb2)$ were characterized by determining rate constants for activation peptide (AP) release, generation of activity, and exposure of the active-site thiol group. Platelet factor XIII activity was generated at a rate approximately twice the rate of AP release. This difference in rates was shown to be consistent with a reaction pathway for activation of platelet factor XIII wherein full factor XIIIa activity is generated after only one AP is removed from the dimeric zymogen. In the presence of calcium, cleavage of one subunit (formation of an $a\sp\prime a$ heterodimer) resulted in exposure of the active-site thiol groups on both the cleaved and uncleaved subunits, whereas in the absence of calcium, cleavage of one subunit only resulted in exposure of the active-site thiol group on the cleaved subunit. Hence, calcium is an allosteric effector in the factor XIII activation pathway. The active-site thiol groups on the cleaved and uncleaved subunits of a once-cleaved, $a\sp\prime a$ heterodimer were equally accessible to alkylation by (1-$\sp{14}$C) iodoacetamide. Only one active-site thiol group per dimer reacted with iodoacetamide upon cleavage of factor XIII by $\alpha$-thrombin in the presence or absence of calcium. Thus, either a negative cooperative interaction occurs between the factor XIIIa subunits upon alkylation, or the active-site thiol groups on the individual subunits of the dimer reciprocate between accessible and inaccessible states. In addition, the binding of the carbamylmethyl derivatives of the active-site thiol group of platelet factor XIII (CM$a\sb2)$ and factor XIIIa (CM$a\sb2\sp\prime)$ to fibrin was studied. The enzyme-like derivative (CM$a\sb2\sp\prime)$ bound more tightly to fibrin (K$\sb{\rm d}$ = 2.1 $\mu$M) than did CM$a\sb2$ (K$\sb{\rm d}$ = 14 $\mu$M), but the binding of each was weaker than the binding of plasma factor XIII zymogen $(a\sb2 b\sb2)$ to fibrin (K$\sb{\rm d}$ = 0.20 $\mu$M) under the same conditions. Saturation of fibrin with plasma factor XIII in the presence of CM$a\sb2\sp\prime$ did not affect the binding of CM$a\sb2\sp\prime$ to fibrin, suggesting that plasma factor XIII zymogen and CM$a\sb2\sp\prime$ bind to separate, non-interacting sites on fibrin. The affinity of $a\sb2\sp\prime b\sb2$ for fibrinogen (K$\sb{\rm d}$ = 2.3 $\mu$M) was determined using a kinetic method.PhDBiological ChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/105909/1/9226918.pdfDescription of 9226918.pdf : Restricted to UM users only