Characteristics of cation binding to the I domains of LFA-1 and MAC-1 -: The LFA-1 I domain contains a Ca2+-binding site

The crystal structures of the I domains of integrins MAC-1 (alpha(M)beta(2); CD11lb/CD18) and LFA-1 (alpha(L)beta(2); CD11a/CD18) show that a single conserved cation-binding site is present in each protein. Purified recombinant I domains have intrinsic ligand binding activity, and in several systems this interaction has been demonstrated to be cation-dependent, It has been proposed that the I domain cation-binding site represents a general metal ion-dependent adhesion motif utilized for binding protein ligands, Here we show that the purified recombinant I domain of LFA-I (alpha(L)I) binds cations, but with significantly different characteristics compared with the I domain of MAC-1 (alpha(M)I) Both alpha(L)I and alpha(M)I bind Mn-54(2+) in a conformation-dependent manner, and in general, cations with charge and size characteristics similar to Mn2+ most effectively inhibit Mn-54(2+) binding. Surprisingly, however, physiological levels of Ca2+ (1-2 mM) inhibited Mn-54(2+) binding to purified alpha(L)I, but not to alpha(M)I. Using Ca-45(2+) and Mn-54(2+) in direct binding studies, the dissociation constants (K-D) for the interactions between these cations and alpha(L)I were estimated to be 5-6 x 10(-5) and 1-2 x 10(-5) M, respectively. Together with the available structural information, the data suggest differential affinities for Mn2+ and Ca2+ binding to the single conserved site within alpha(L)I. Antagonism of LFA-1, but not MAC-1, -mediated cell adhesion by Ca2+ may be related to the Ca2+ binding activity of the LFA-1 I domain.