Kinetic measurements of binding of galectin 3 to a laminin substratum.

Galectin 3, a β-galactoside binding protein, contains a C-terminal carbohydrate recognition domain (CRD) and an N-terminal segment including multiple repeats of a proline/tyrosine/glycine-rich motif. Previous work has shown that galectin 3 but not the isolated CRD binds to laminin, a multivalent ligand, with positive cooperativity indicating the formation of multiple interactions although the lectin in solution is monomeric. Using surface plasmon resonance, we find that hamster galectin 3 at sub-µmolar concentrations or its isolated CRD at all concentrations binds to a laminin substratum with similar association (kass; 10 – 30 000 M−1 S−1) and dissociation (kdiss; 0.2 – 0.3 S 1 −1 ) rates and weak affinity (Ka; 1 - 3 X 105 M−1). At higher concentrations of galectin 3 the off rate decreases ten fold leading to increased affinity. Ligation of an N-terminal epitope of galectin 3 with a monoclonal Fab fragment increases association and dissociation rates ten fold. A recombinant protein obtained by deletion of the first 93 N-terminal residues binds to laminin with positive cooperativity and a slowly dissociating fraction (Kdiss; 0.002 S−1) accummulates on the substratum. The data suggest that homophilic interactions between CRD as well as N terminal domains are implicated in galectin 3 aggregation on the substratum leading to positive binding cooperativity.