IDENTIFICATION OF PUTATIVE LIGAND-BINDING SITES WITHIN I-DOMAIN OF INTEGRIN ALPHA-2-BETA-1 (VLA-2, CD49B CD29)/

Integrin alpha2beta1 is a cell surface adhesion receptor for collagen and echovirus 1. Here we localized the epitopes for anti-alpha2 monocl onal antibodies using interspecies (human/bovine) alpha2 chimeras with different lengths of human alpha2 sequence on the amino-terminal side and site-directed mutagenesis. The antibodies that block the collagen and/or echovirus 1 binding to human alpha2beta1 (6F1, RMAC11, 12F1, a nd AA10) recognizes a small region (residues 173-259) within the I dom ain. Asp-160 and Arg-242 are critical for binding of the two other fun ction-inhibiting antibodies, P1H5 and 5E8, respectively. Notably, muta tions of Asp-151 and Asp-254 block the binding of alpha2beta1 to colla gen. These data suggest that the I domain (residues 140-359) is critic ally involved in the ligand/receptor interactions, and collagen and ec hovirus 1 binding sites are adjacent or overlapping within the I domai n. The sequence of the residues 173-259 of alpha2 overlap with the pep tide sequences (M11 and M20) that derive from von Willebrand factor A1 and A3 domains (homologous to the alpha2 I domain) and block von Will ebrand factor/collagen interaction, suggesting that the epitope region of alpha2 (residues 173-259) may really be involved in ligand recogni tion.