Metal Ion-dependent Adhesion Site Research Articles

Identifying the Putative Metal Ion-dependent Adhesion Site in the β2 (CD18) Subunit Required for αLβ2 and αMβ2 Ligand Interactions

We have previously demonstrated that Asp134 and Ser136 of the beta2 subunit are essential for alphaLbeta2 and alphaMbeta2 ligand recognition. It has been proposed that these residues may be part of a metal ion-dependent adhesion site (MIDAS) within the beta subunit homologous to the alphaM I domain MIDAS structure (Lee, J.-O., Rieu, P., Arnaout, M. A., and Liddington, R. (1995) Cell 80, 631-638). In the present study, we evaluated the role of additional candidate metal ion-coordinating residues in the beta2 subunit in ligand interactions. Cells bearing the recombinant alphaLbeta2 or alphaMbeta2 mutant(s) were tested for the ability to bind to immobilized ligands. Alanine substitution at Asp232 in beta2 produced a complete loss in the capacity of both alphaLbeta2 and alphaMbeta2 to support cell adhesion and suppressed the expression of a divalent cation-dependent conformation recognized by mAb 24. Alanine substitution at Glu235 differentially affected receptor function dependent upon the co-transfected alpha subunit. Cells expressing alphaLbeta2 with a substitution at Glu235 failed to adhere to intercellular adhesion molecule 1 (ICAM-1) but did retain the capacity to bind mAb 24. Moreover, cells expressing alphaMbeta2 with a substitution at Glu235 failed to adhere to fibrinogen or ICAM-1 and did not bind mAb 24. However, these cells did retain the capacity to adhere to iC3b following antibody-induced activation. These results implicate Asp232 and Glu235, along with Asp134 and Ser136, in ligand binding function of alphaLbeta2 and alphaMbeta2. These findings provide evidence in support of the existence of a MIDAS structure in beta2 analogous to that seen in the alphaM I domain.

Crystal structure of the A domain from the a subunit of integrin CR3 (CD11 b/CD18)

We have determined the high resolution crystal structure of the A domain from the a chain of integrin CR3. The domain adopts a classic α/β “Rossmann” fold and contains an unusual Mg 2+ coordination site at its surface. One of the coordinating ligands is the glutamate side chain from another A domain molecule. We suggest that this site represents a general metal ion-dependent adhesion site (MIDAS) for binding protein ligands. We further propose that the subunits of integrins contain a MIDAS motif within a modified A domain. Our crystal structure will allow reliable models to be built for other members of the A domain superfamily and should facilitate development of novel adhesion modulatory drugs.