title: Regulated unmasking of the cryptic binding site for integrin alpha M beta 2 in the gamma C-domain of fibrinogen.
abstract: Fibrinogen is a ligand for leukocyte integrin alpha(M)beta2 (CD11b/CD18, Mac-1) and mediates adhesion and migration of leukocytes during the immune-inflammatory responses. The binding site for alpha(M)beta2 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)beta2. It is unknown why alpha(M)beta2 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)beta2. We found that the alpha(M)beta2-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 D100 fragment of fibrinogen, which contains intact gammaC and is not able to inhibit adhesion of the alpha(M)beta2-expressing cells, into the fragment D98, 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)(Glu253-Arg261). Exposure of the P2-C sequence in fibrinogen, D100, and D98 was probed with a site-specific mAb. P2-C is not accessible in soluble fibrinogen and D100 but becomes exposed in D98. 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)beta2-binding site in the D-domain. These results demonstrate that conformational alterations regulate the alpha(M)beta2-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)beta2-binding site in fibrinogen.
authors.affiliation: Joseph J. Jacobs Center for Thrombosis and Vascular Biology, Department of Molecular Cardiology, Lerner Research Institute, Cleveland, Ohio 44195, USA.
authors.forename: Valeryi K
authors.initials: VK
authors.lastname: Lishko
authors.forename: Bohdan
authors.initials: B
authors.lastname: Kudryk
authors.forename: Valentin P
authors.initials: VP
authors.lastname: Yakubenko
authors.forename: Vivien C
authors.initials: VC
authors.lastname: Yee
authors.forename: Tatiana P
authors.initials: TP
authors.lastname: Ugarova
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002448:Cell Adhesion; D002460:Cell Line; D010450:Endopeptidases / Q000378:metabolism; D005109:Extracellular Matrix / Q000737:chemistry / Q000378:metabolism; D005338:Fibrin Fibrinogen Degradation Products / Q000737:chemistry / Q000378:metabolism; D005340:Fibrinogen / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D016177:Macrophage-1 Antigen / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D005338:Fibrin Fibrinogen Degradation Products; D016177:Macrophage-1 Antigen; D011994:Recombinant Proteins; C015214:fibrinogen D fragment; D005340:Fibrinogen; D010450:Endopeptidases