The Arg-Gly-Asp binding domain of the vitronectin receptor. Photoaffinity cross-linking implicates amino acid residues 61-203 of the beta subunit.

Abstract

We have employed photoaffinity cross-linking to examine RGD recognition by the human placental vitronectin receptor. The peptide GRGDSPK was coupled to a thiol-cleavable radioiodinatable aryl azide (sulfosuccinimidyl 2-(p-azido-salicylamido)-1,3'-dithiopropionate. When 125I-sulfosuccinimidyl 2-(p-azido-salicylamido)-1,3'-dithiopropionate-GRGDSPK was cross-linked to the vitronectin receptor in solution, 80% of the label was associated with the beta subunit. Cross-linking to both subunits of the receptor was highly specific and dependent upon the presence of divalent cations. Ca2+ and Mg2+ promoted RGD recognition by the receptor; however, the effects of each divalent cation were kinetically distinct. We have also identified and determined the amino acid sequence of chymotryptic and V8 protease-generated peptides of the beta subunit that were radiolabeled as a result of cross-linking. The results of these studies demonstrate that amino acid residues 61-203 are proximal to the RGD binding domain of the vitronectin receptor.