Integrin/ligand interaction is a therapeutic target for many diseases. We previously reported that residues critical for ligand binding are clustered in N-terminal repeat 3 (in the predicted 2-3 loop) of α4, α5 and αIIb. Here we have localized residues critical for ligand binding in the α3 subunit of integrin α3β1 with distinct ligand specificity (laminin-5). We identified an α3 epitope common to several function-blocking anti-α3 antibodies at the boundary between repeats 1 and 2 (residues 75-80). We found that swapping the predicted 4-1 loop (residues 153-165) at the boundary between repeats 2 and 3 with the corresponding α4 sequence and mutating Thr- 162 and Gly-163 residues in this predicted loop block laminin-5 binding. Thr- 162 and Gly-163 and the antibody epitope are separated in the primary structure; however, they are close to each other in the proposed β-propeller model. Mutating residues recently reported to block (Tyr-186 and Trp-188) or enhance (Asp-122) laminin-5 binding to α3β1 [Krukonis, E.S., Dersch, P., Eble, J.A., and Isberg, R.R. (1998) J. Biol. Chem. 273, 31837-31843] did not affect laminin-5 binding under the assay conditions used. Thr-162 and Gly-163 are not critical for adhesion to invasin, indicating that laminin-5 and invasin may use different recognition mechanisms, and that mutation of Thr- 162 and Gly-163 does not drastically affect the integrity of α3β1. These results suggest that residues critical for ligand binding may be similarly (but not identically) located in repeat 3 of the α subunit regardless of ligand specificity.
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