ADAMs (a disintegrin and metalloprotease) are a family of proteins that possess functional adhesive and proteolytic domains. ADAM 28 (MDC-L) is expressed by human lymphocytes and contains a disintegrin-like domain that serves as a ligand for the leukocyte integrin, α4β1. To elucidate which residues comprise the α4βl binding site in the ADAM 28 disintegrin domain, a charge-to-alanine mutagenesis strategy was utilized. Each alanine substitution mutant was evaluated and compared to the native sequence for its ability to support cell adhesion of the T-lymphoma cell line, Jurkat. This approach identified ADAM 28 residues Lys[sup 437], Lys[sup 442], Lys[sup 455], Lys[sup 459], Lys[sup 460], Lys[sup 469], and Glu[sup 476] as being essential for α4βl-dependent cell adhesion. The epitope for a function-blocking monoclonal antibody, Dis 1-1, was localized to the N-terminal end of the ADAM 28 disintegrin domain using these same charge-toalanine mutants. Three distinct molecular models based upon the known structures of snake venom disintegrins suggested that residues contributing to α4β1 recognition are aligned on one face of the domain. This study demonstrates that residues located outside of the disintegrin loop participate in integrin recognition of mammalian disintegrins. [ABSTRACT FROM AUTHOR]