Fibrinogen binds to integrin alpha(5)beta(1) via the carboxyl-terminal RGD site of the Aalpha-chain.
Fibrinogen interactions with vascular endothelial cells are implicated in various physiological and pathophysiological events, including angiogenesis and wound healing. We have shown previously that integrin alpha(5)beta(1) is a fibrinogen receptor on endothelial cells [Suehiro, K., Gailit, J., and Plow, E.F. (1997) J. Biol. Chem. 272, 5360-5366]. In the present study, we have characterized fibrinogen interactions with purified alpha(5)beta(1) and have identified the recognition sequence in fibrinogen for alpha(5)beta(1). The binding of fibrinogen to immobilized alpha(5)beta(1) was selectively supported by Mn(2+). Fibrinogen bound to purified alpha(5)beta(1) in a time-dependent, specific, and saturable manner in the presence of Mn(2+), and the binding was blocked completely by Arg-Gly-Asp (RGD)-containing peptides and by anti-alpha(5) and anti-alpha(5)beta(1) monoclonal antibodies. A monoclonal antibody directed to the C-terminal RGD sequence at Aalpha572-574 significantly inhibited the binding of fibrinogen to alpha(5)beta(1), whereas monoclonal antibodies directed to either the N-terminal RGD sequence at Aalpha95-97 or the C-terminus of the gamma-chain did not. Furthermore, substituting RGE for RGD at position Aalpha95-97 in recombinant fibrinogen had a minimal effect on binding, whereas substituting RGE for RGD at position Aalpha572-574 decreased binding by 90%. These results demonstrate that the C-terminal RGD sequence at Aalpha572-574 is required for the interaction of fibrinogen with alpha(5)beta(1).
Suehiro
K
K
Blood Transfusion Division, Miyazaki Medical College, Miyazaki 889-1692, Japan. suehirok@post1.miyazaki-med.ac.jp
Mizuguchi
J
J
Nishiyama
K
K
Iwanaga
S
S
Farrell
D H
DH
Ohtaki
S
S
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D001667:Binding, Competitive; D002413:Cations, Divalent / Q000378:metabolism / Q000494:pharmacology; D005340:Fibrinogen / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D008345:Manganese / Q000378:metabolism / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation; D009842:Oligopeptides / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D017469:Receptors, Fibronectin / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
D000911:Antibodies, Monoclonal; D002413:Cations, Divalent; D009842:Oligopeptides; D010446:Peptide Fragments; D017469:Receptors, Fibronectin; D011994:Recombinant Proteins; C112658:arginyl-glycyl-glutamic acid; D008345:Manganese; C047981:arginyl-glycyl-aspartic acid; D005340:Fibrinogen