title: EM structure of the ectodomain of integrin CD11b/CD18 and localization of its ligand-binding site relative to the plasma membrane.
abstract: One-half of the integrin α-subunit Propeller domains contain and extra vWFA domain (αA domain), which mediates integrin binding to extracellular physiologic ligands via its metal-ion-dependent adhesion site (MIDAS). We used electron microscopy to determine the 3D structure of the αA-containing ectodomain of the leukocyte integrin CD11b/CD18 (αMβ2) in its inactive state. A well defined density for αA was observed within a bent ectodomain conformation, while the structure of the ectodomain in complex with the Fab fragment of mAb107, which binds at the MIDAS face of CD11b and stabilizes the inactive state, further revealed that αA is restricted to a relatively small range of orientations relative to the Propeller domain. Using Fab 107 as probe in fluorescent lifetime imaging microscopy (FLIM) revealed that αA is positioned relatively far from the membrane surface in the inactive state, and a systematic orientation search revealed that the MIDAS face would be accessible to extracellular ligand in the inactive state of the full-length cellular integrin. These studies are the first to define the 3D EM structure of an αA-containing integrin ectodomain and to position the ligand-binding face of αA domain in relation to the plasma membrane, providing new insights into current models of integrin activation.
authors.affiliation: Structural Biology Program, Massachusetts General Hospital and Harvard Medical School, Charlestown, Massachusetts, United States of America.
authors.forename: Brian D
authors.initials: BD
authors.lastname: Adair
authors.forename: Jian-Ping
authors.initials: JP
authors.lastname: Xiong
authors.forename: José Luis
authors.initials: JL
authors.lastname: Alonso
authors.forename: Bradley T
authors.initials: BT
authors.lastname: Hyman
authors.forename: M Amin
authors.initials: MA
authors.lastname: Arnaout
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry; D001665:Binding Sites; D039481:CD11b Antigen / Q000737:chemistry* / Q000378:metabolism*; D018821:CD18 Antigens / Q000737:chemistry* / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D006801:Humans; D021621:Imaging, Three-Dimensional; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism; D008024:Ligands; D008670:Metals / Q000378:metabolism; D008854:Microscopy, Electron; D008958:Models, Molecular; D017434:Protein Structure, Tertiary
chemical_list: D000911:Antibodies, Monoclonal; D039481:CD11b Antigen; D018821:CD18 Antigens; D007140:Immunoglobulin Fab Fragments; D008024:Ligands; D008670:Metals