INTEGRIN LFA-1 ALPHA-SUBUNIT CONTAINS AN ICAM-1 BINDING-SITE IN DOMAIN-V AND DOMAIN-VI

In order to identify a binding site for ligand intercellular adhesion molecule-1 (ICAM-1) on the beta 2 integrin lymphocyte function-associa ted antigen-1 (LFA-1), protein fragments of LFA-1 were made by in vitr o translation of a series of constructs which featured domain-sized de letions starting from the N-terminus of the alpha subunit of LFA-1. Mo noclonal antibodies and ICAM-1 were tested for their ability to bind t o these protein fragments. Results show that the putative divalent cat ion binding domains V and VI contain an ICAM-1 binding site. A series of consecutive peptides covering these domains indicated two discontin uous areas as specific contact sites: residues 458-467 in domain V and residues 497-516 in domain VI. A three-dimensional model of these dom ains of LFA-1 was constructed based on the sequence similarity to know n EF hands. The two regions critical for the interaction of LFA-1 with ICAM-1 lie adjacent to each other, the first next to the nonfunctiona l EF hand in domain V and the second coinciding with the potential div alent cation binding loop in domain VI. The binding of ICAM-1 with the domain V and VI region in solution was not sensitive to divalent cati on chelation. In short, a critical motif for ICAM-1 binding to the alp ha subunit of LFA-1 is shared between two regions of domains V and VI.