Characterization of a conformationally sensitive murine monoclonal antibody directed to the metal ion-dependent adhesion site face of integrin CD11b.
Integrin binding to physiologic ligands requires divalent cations and an inside-out-driven switch of the integrin to a high-affinity state. Divalent cations at the metal ion-dependent adhesion site (MIDAS) face of the alpha subunit-derived A domain provide a direct bridge between ligands and the integrin, and it has been proposed that activation dependency is caused by reorientation of the surrounding residues relative to the metal ion, forming an optimal binding interface. To gain more insight into the functional significance of the protein movements on the MIDAS face, we raised and characterized a murine mAb 107 directed against the MIDAS face of the A domain from integrin CD11b. We find that mAb 107 behaves as a ligand mimic. It binds in a divalent-cation-dependent manner to solvent-exposed residues on the MIDAS face of CD11b, blocks interaction of 11bA or the holoreceptor with ligands, and inhibits spreading and phagocytosis by human neutrophils. However, in contrast to physiologic ligands, mAb 107 preferentially binds to the inactive low-affinity form of the integrin, suggesting that its antagonistic effects are exerted in part by stabilizing the receptor in the low-affinity state. These data support a functional relevance of the protein movements on the MIDAS face and suggest that stabilizing the A domain in the low-affinity state may have therapeutic benefit.
Li
Rui
R
Leukocyte Biology and Inflammation Program, Structural Biology Program, Renal Unit, Massachusetts General Hospital, Charlestown, MA 02129, USA.
Haruta
Ikuko
I
Rieu
Philippe
P
Sugimori
Takashi
T
Xiong
Jian-Ping
JP
Arnaout
M Amin
MA
D000818:Animals; D019138:Antibodies, Blocking / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D000915:Antibody Affinity; D001666:Binding Sites, Antibody / Q000276:immunology*; D001667:Binding, Competitive / Q000276:immunology; D018821:CD18 Antigens / Q000378:metabolism / Q000502:physiology; D016466:CHO Cells; D019556:COS Cells; D002413:Cations, Divalent / Q000378:metabolism; D006224:Cricetinae; D006801:Humans; D016023:Integrins / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D008024:Ligands; D016177:Macrophage-1 Antigen / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D008670:Metals / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D011487:Protein Conformation
D019138:Antibodies, Blocking; D000911:Antibodies, Monoclonal; D018821:CD18 Antigens; D002413:Cations, Divalent; D016023:Integrins; D008024:Ligands; D016177:Macrophage-1 Antigen; D008670:Metals