MULTIPLE LOOP STRUCTURES CRITICAL FOR LIGAND-BINDING OF THE INTEGRIN ALPHA-4 SUBUNIT IN THE UPPER FACE OF THE BETA-PROPELLER MODE-1

A non-I-domain integrin, alpha 4 beta 1, recognizes vascular cell adhe sion molecule 1 (VCAM-1) and the IIICS portion of fibronectin. To loca lize regions of alpha 4 critical for ligand binding, we swapped severa l predicted loops within or near the putative ligand-binding site of a lpha 4 (which spans repeats 25 of the seven N-terminal repeats) with t he corresponding regions of alpha 5. Swapping residues 112-131 in repe at 2, or residues 237-247 in repeat 4, completely blocked adhesion to immobilized VCAM-1 and connecting segment 1 (CS-1) peptide. However, s wapping residues 4062 in repeat 1, residues 151-164 in repeat 3, or re sidues 282-288 (which contain a putative cation binding motif) in repe at 5 did not affect or only slightly reduced adhesion to these ligands . The binding of several function-blocking antibodies is blocked by sw apping residues 112-131, 151-164, and 186-191 (which contain previousl y identified residues critical for ligand binding, Tyr-187 and Gly-190 ). These results are consistent with the recently published beta-prope ller folding model of the integrin alpha 4 subunit [Springer, T. A. (1 997) Proc. Natl. Acad. Sci. USA 94, 65-72], in which seven four-strand ed beta-sheets are arranged in a torus around a pseudosymmetric axis. The regions of alpha 4 critical for ligand binding are adjacent to eac h other and are located in the upper face, the predicted ligand-bindin g site, of the beta-propeller model, although they are not adjacent in the primary structure.