Regulated unmasking of the cryptic binding site for integrin αMβ2 in the γC-domain of fibrinogen

Fibrinogen is a ligand for leukocyte integrin alpha(M)beta(2) (CD11b/CD18, Mac-1) and mediates adhesion and migration of leukocytes during the immune-inflammatory responses. The binding site for alpha(M)beta(2) resides in gammaC, a constituent subdomain in the D-domain of fibrinogen. The sequence gamma383-395 (P2-C) in gammaC was implicated as the major binding site for alpha(m)beta(2). It is unknown why alpha(M)beta(2) on leukocytes can bind to immobilized fibrinogen in the presence of high concentrations of soluble fibrinogen in plasma. In this study, we have investigated the accessibility of the binding site in fibrinogen for alpha(M)beta(2). We found that the alpha(M)beta(2)-binding site in gammaC is cryptic and identified the mechanism that regulates its unmasking. Proteolytic removal of the small COOH-terminal segment(s) of gammaC, gamma397/405-411, converted the D-100 fragment of fibrinogen, which contains intact gammaC and is not able to inhibit adhesion of the alpha(M)beta(2)-expressing cells, into the fragment D-98, which effectively inhibited cell adhesion. D98, but not D100, bound to the recombinant alpha(M)I-domain, and the alpha(M)I-domain recognition peptide, alpha(M)(Glu(253)-Arg(261)). Exposure of the P2-C sequence in fibrinogen, D-100, and D-98 was probed with a site-specific mAb. P2-C is not accessible in soluble fibrinogen and D-100 but becomes exposed in D-98. P2-C is also unmasked by immobilization of fibrinogen onto a plastic and by deposition of fibrinogen in the extracellular matrix. Thus, exposure of P2-C by immobilization and by proteolysis correlates with unmasking of the alpha(M)beta(2)-binding site in the D-domain. These results demonstrate that conformational alterations regulate the alpha(M)beta(2)-binding site in gammaC and suggest that processes relevant to tissue injury and inflammation are likely to be involved in the activation of the alpha(M)beta(2)-binding site in fibrinogen.