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

The alpha subunits of leukocyte CD11/CD18 integrins contain an approximately 200-amino acid "inserted" domain (I-domain) that may be important for multivalent adhesive recognitions. A recombinant form of the I-domain of CD11b/CD18 was generated and analyzed directly for interaction with complementary integrin ligands. CD11b I-domain bound the activation-dependent monoclonal antibody 7E3, and the functionally blocking anti-CD11b monoclonal antibodies OKM9, 60.1, and LM2/1, but not OKM1 or M1/70. Fibrinogen or soluble intercellular adhesion molecule-1 associated with CD11b I-domain in a concentration-dependent manner. Binding of 125I-fibrinogen to recombinant CD11b I-domain was saturable, governed by a Kd of approximately 0.22 +/- 0.06 microM, and fully inhibited by molar excess of unlabeled fibrinogen, or by the P1 peptide (KY)GWTVFQKRLDGSV (IC50 approximately 2.5-5 microM), duplicating the fibrinogen gamma chain sequence Gly190-Val202. In contrast, 125I-factor X binding to CD11b I-domain was only partially inhibited (50-60%) by a molar excess of unlabeled factor X, and entirely unaffected by functionally blocking anti-CD11b monoclonal antibodies or by factor X-derived synthetic peptidyl antagonists. We conclude that the I-domain of CD11b participates in qualitative mechanisms of receptor activation and contains the binding site(s) for the CD11b/CD18 ligands fibrinogen and intercellular adhesion molecule-1, while it is only minimally implicated in the recognition of factor X.