Differential ligand binding specificities of recombinant CD11b/CD18 integrin I-domain.

Abstract

The (Y subunits of leukocyte CDll/CDlS integrins contain an -200-amino acid "inserted" domain (I-domain) that may be important for multivalent adhesive recognitions. A recombinant form of the I-domain of CDllbl CDl8 was generated and analyzed directly for interaction with complementary integrin ligands. CDllb I-domain bound the activation-dependent monoclonal antibody 7E3, and the functionally blocking anti-CDllb monoclonal antibodies 0-9, 60.1, and LM2/1, but not 0-1 or M1/70. Fibrinogen or soluble intercellular adhesion molecule-1 associated with CDllb I-domain in a concentration-dependent manner. Binding of laSI-fibrinogen to recombinant CDllb I-domain was saturable, governed by a Kd of "0.22 2 0.06 p ~, and fully inhibited by molar excess of unlabeled fibrinogen, or by the P1 peptide (KY)GWTVFQKRLDGSV (IC, -2 . 5 6 pd, duplicating the fibrinogen y chain sequence Gly'Bo-Va1202. In contrast, 'zsI-factor X binding to CDllb I-domain was only partially inhibited ( 5 0 4 0 % ) by a molar excess of unlabeled factor X, and entirely unaffected by functionally blocking anti-CDllb monoclonal antibodies or by factor X-derived synthetic peptidyl antagonists. W e conclude that the I-domain of CDllb participates in qualitative mechanisms of receptor activation and contains the binding site(s) for the CDllb/CDlS ligands fibrinogen and intercellular adhesion molecule-1, while it is only minimally implicated in the recognition of factor X. Leukocyte pz integrins comprise three heterodimeric transmembrane receptors CDlldCD18 (LFA-l), CDllbKD18 (Macl ) , a n d C D l l d C D 1 8 ( ~1 5 0 , 951, generated by the assembly of the common p subunit CD18 with the three respective a chains C D l l a (a,), C D l l b (aM), and C D l l c (a,) (1-3). Broadly distributed on all leukocytes of lymphoid and myelomonocytic lineage,