BINDING OF FIBRINOGEN A-ALPHA-1-50-BETA-GALACTOSIDASE FUSION PROTEIN TO THROMBIN STABILIZES THE SLOW FORM

The interaction of fibrinogen A alpha 1-50-beta-galactosidase fusion p rotein with the slow and fast forms of thrombin was studied and compar ed to thrombin-fibrinogen interaction under identical solution conditi ons. At equilibrium, the affinity of the fusion protein for the slow f orm of thrombin is 3 times higher than its affinity for the fast form. The fusion protein and fibrinogen have the same affinity for the fast form. On the other hand, the affinity of the fusion protein for the s low form of thrombin is 40 times tighter than that of fibrinogen. In t he transition state, binding of the fusion protein has the same proper ties as fibrinogen, with the fast form showing higher specificity. The N-terminal fragment of the fibrinogen A alpha chain thus contains res idues that are responsible for the preferential binding of the fusion protein to the slow form at equilibrium and to the fast form in the tr ansition state. If this fragment binds to thrombin in a similar way fo r fibrinogen and the fusion protein, then the N-terminal domains of th e B beta and gamma chains of fibrinogen, that are not present in the f usion protein, must play a key role in the binding of fibrinogen to th rombin at equilibrium. These chains may destabilize binding to the slo w form by nearly 2.4 kcal/mol, thereby favoring binding of fibrinogen to the fast form. We propose that the three chains of fibrinogen play different roles in the thrombin-fibrinogen interaction, with the A alp ha chain containing residues for preferential binding to the fast form in the transition state and the B beta and gamma chains containing re sidues that destabilize binding to the slow form at equilibrium.