Identification of a novel binding site for platelet integrins αIIbβ3 (GPIIbIIIa) and α5β1 in the γC-domain of fibrinogen

The interactions of platelets with fibrinogen mediate a variety of responses including adhesion, platelet aggregation, and fibrin clot retraction. Whereas it was assumed that interactions of the platelet integrin alpha(IIb)beta(3) with the AGDV sequence in the gammaC-domain of fibrinogen and/or RGD sites in the Aalpha chains are involved in clot retraction and adhesion, recent data demonstrated that fibrinogen lacking these sites still supported clot retraction. These findings suggested that an unknown site in fibrinogen and/or other integrins participate in clot retraction. Here we have identified a sequence within gammaC that mediates binding of fibrinogen to platelets. Synthetic peptide duplicating the 365 - 383 sequence in gammaC, designated P3, efficiently inhibited clot retraction in a dose-dependent manner. Furthermore, P3 supported platelet adhesion and was an effective inhibitor of platelet adhesion to fibrinogen fragments. Analysis of overlapping peptides spanning P3 and mutant recombinant gammaC-domains demonstrated that the P3 activity is contained primarily within gamma370-383. Integrins alpha(IIb)beta(3) and beta(5)beta(1) were implicated in recognition of P3, since platelet adhesion to the peptide was blocked by function-blocking monoclonal antibodies against these receptors. Direct evidence that alpha(IIb)beta(3) and alpha(5)beta(1) bind P3 was obtained by selective capture of these integrins from platelet lysates using a P3 affinity matrix. Thus, these data suggest that the P3 sequence in the gammaC-domain of fibrinogen defines a previously unknown recognition specificity of alpha(IIb)beta(3) and alpha(5)beta(1) and may function as a binding site for these integrins.