title: Mapping early conformational changes in alphaIIb and beta3 during biogenesis reveals a potential mechanism for alphaIIbbeta3 adopting its bent conformation.
abstract: Current evidence supports a model in which the low-affinity state of the platelet integrin alphaIIbbeta3 results from alphaIIbbeta3 adopting a bent conformation. To assess alphaIIbbeta3 biogenesis and how alphaIIbbeta3 initially adopts the bent conformation, we mapped the conformational states occupied by alphaIIb and beta3 during biogenesis using conformation-specific monoclonal antibodies (mAbs). We found that alphaIIbbeta3 complex formation was not limited by the availability of either free pro-alphaIIb or free beta3, suggesting that other molecules, perhaps chaperones, control complex formation. Five beta3-specific, ligand-induced binding site (LIBS) mAbs reacted with much or all free beta3 but not with beta3 when in complex with mature alphaIIb, suggesting that beta3 adopts its mature conformation only after complex formation. Conversely, 2 alphaIIb-specific LIBS mAbs directed against the alphaIIb Calf-2 region adjacent to the membrane reacted with only minor fractions of free pro-alphaIIb, raising the possibility that pro-alphaIIb adopts a bent conformation early in biogenesis. Our data suggest a working model in which pro-alphaIIb adopts a bent conformation soon after synthesis, and then beta3 assumes its bent conformation by virtue of its interaction with the bent pro-alphaIIb.
authors.affiliation: Department of Pediatrics, Mount Sinai School of Medicine, and New York Blood Center, 310 E. 67th Street, New York, NY 10021, USA. bmitchell@nybloodcenter.org
authors.forename: W Beau
authors.initials: WB
authors.lastname: Mitchell
authors.forename: Jihong
authors.initials: J
authors.lastname: Li
authors.forename: Marta
authors.initials: M
authors.lastname: Murcia
authors.forename: Nathalie
authors.initials: N
authors.lastname: Valentin
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Newman
authors.forename: Barry S
authors.initials: BS
authors.lastname: Coller
mesh_terms: D002460:Cell Line; D006801:Humans; D039661:Integrin beta3 / Q000096:biosynthesis* / Q000737:chemistry; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000096:biosynthesis* / Q000737:chemistry; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000096:biosynthesis* / Q000737:chemistry; D040201:Platelet Membrane Glycoprotein IIb / Q000096:biosynthesis* / Q000737:chemistry; D025941:Protein Interaction Mapping; D020836:Protein Structure, Quaternary
chemical_list: C061248:ITGB3 protein, human; D039661:Integrin beta3; D046912:Multiprotein Complexes; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D040201:Platelet Membrane Glycoprotein IIb