title: Swing-out of the β3 hybrid domain is required for αIIbβ3 priming and normal cytoskeletal reorganization, but not adhesion to immobilized fibrinogen.
abstract: Structural and functional analyses of integrin αIIbβ3 has implicated swing-out motion of the β3 hybrid domain in αIIbβ3 activation and ligand binding. Using data from targeted molecular dynamics (TMD) simulations, we engineered two disulfide-bonded mutant receptors designed to limit swing-out (XS-O). XS-O mutants cannot bind the high Mr ligand fibrinogen in the presence of an activating mAb or after introducing mutations into the αIIb subunit designed to simulate inside-out signaling. They also have reduced capacity to be "primed" to bind fibrinogen by pretreatment with eptifibatide. They can, however, bind the small RGD venom protein kistrin. Despite their inability to bind soluble fibrinogen, the XS-O mutants can support adhesion to immobilized fibrinogen, although such adhesion does not initiate outside-in signaling leading to normal cytoskeletal reorganization. Collectively, our data further define the biologic role of β3 hybrid domain swing-out in both soluble and immobilized high Mr ligand binding, as well as in priming and outside-in signaling. We also infer that swing-out is likely to be a downstream effect of receptor extension.
authors.affiliation: Allen and Frances Adler Laboratory of Blood and Vascular Biology, The Rockefeller University, New York, New York, United States of America.
authors.forename: Ming
authors.initials: M
authors.lastname: Cheng
authors.affiliation: Allen and Frances Adler Laboratory of Blood and Vascular Biology, The Rockefeller University, New York, New York, United States of America.
authors.forename: Jihong
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Structural and Chemical Biology, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
authors.forename: Ana
authors.initials: A
authors.lastname: Negri
authors.affiliation: Allen and Frances Adler Laboratory of Blood and Vascular Biology, The Rockefeller University, New York, New York, United States of America.
authors.forename: Barry S
authors.initials: BS
authors.lastname: Coller
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001665:Binding Sites; D001792:Blood Platelets / Q000166:cytology / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D000077542:Eptifibatide; D005340:Fibrinogen / Q000737:chemistry* / Q000378:metabolism; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D055767:Immobilized Proteins / Q000737:chemistry* / Q000378:metabolism; D008024:Ligands; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000737:chemistry; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D012911:Snakes
chemical_list: D000911:Antibodies, Monoclonal; D055767:Immobilized Proteins; D008024:Ligands; D010455:Peptides; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D011994:Recombinant Proteins; C067623:kistrin; D005340:Fibrinogen; D000077542:Eptifibatide