title: Identification of critical residues for ligand binding in the integrin beta3 I-domain by site-directed mutagenesis.
abstract: To define the structural basis of ligand recognition by alphaIIb beta3, we conducted site-directed mutagenesis of residues located on the top surface of the beta3 I-domain that is homologous to the I-domain of several alpha subunits and contains a putative ligand binding site. Here we identify D158 and N215 in beta3 as novel residues critical for ligand binding. Alanine substitution of D158 or N215 abolished binding of a ligand-mimetic antibody and fibrinogen to alphaIIb beta3 induced by different types of integrin activation. CHO cells expressing recombinant alphaIIb beta3 bearing D158A or N215A mutation did not adhere to fibrinogen. These mutations had the same effect on ligand binding to another beta3 integrin, alphaVbeta3. Compared to the alphaI-domain structure, the betaB-betaC loop containing D158 in the beta3 I-domain is quite different in length and sequence. These results suggest that the structure for ligand recognition is different in the betaI- and alphaI-domains.
authors: Department of Internal Medicine 1, Ehime University School of Medicine, Shigenobu, Japan.
authors: Jun
authors: J
authors: Yamanouchi
authors: Takaaki
authors: T
authors: Hato
authors: Tatsushiro
authors: T
authors: Tamura
authors: Shigeru
authors: S
authors: Fujita
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001665:Binding Sites; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000235:genetics / Q000378:metabolism; D039661:Integrin beta3 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008024:Ligands; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D039302:Integrin alphaVbeta3; D039661:Integrin beta3; D008024:Ligands; D011993:Recombinant Fusion Proteins; D005340:Fibrinogen