REGULATION OF INTEGRIN ALPHA-5-BETA-1-FIBRONECTIN INTERACTIONS BY DIVALENT-CATIONS - EVIDENCE FOR DISTINCT CLASSES OF BINDING-SITES FOR MN2+, MG2+, AND CA2+

Integrin-ligand interactions are known to be dependent on divalent cat ions, although the precise role of cations in ligand binding is still unclear. Using the interaction between alpha 5 beta 1 and fibronectin as a model system, we have performed a comprehensive analysis of the e ffects of Mn2+, Mg2+, and Ca2+ on Ligand binding. Each cation had dist inct effects on the ligand-binding capacity of alpha 5 beta 1:Mn2+ pro moted high levels of Ligand binding, Mg2+ promoted low levels of bindi ng, and Ca2+ failed to support binding Studies of the effects of diffe rent combinations of cations on Ligand binding indicated that the cati on-binding sites within alpha 5 beta 1 are not all identical, or of br oad specificity, but instead each site shows a distinct preference for one or more cations, Ca2+ strongly inhibited Mn2+-supported Ligand bi nding, but this inhibition was noncompetitive, suggesting that Ca2+ re cognizes different cation-binding sites to Mn2+. In contrast, Ca2+ act ed as a direct competitive inhibitor of Mg2+-supported Ligand binding, implying that Ca2+ can displace Mg2+ from the integrin. However, low concentrations of Ca2+ greatly increased the apparent affinity of Mg2 for its binding site, suggesting the existence of a distinct high aff inity Ca2+-binding site. Taken together, our results imply that the li gand-binding capacity of alpha 5 beta 1 can be regulated in a complex manner through separate classes of binding sites for Mn2+, Mg2+, and C a2+.