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

Galectin 3, a beta-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 cooperativety 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-micromolar concentrations or its isolated CRD at all concentrations binds to a laminin substratum with similar association (k(ass); 10-30,000 M(-1) S(-1)) and dissociation (k(diss); 0.2-0.3 S1(-1)) rates and weak affinity (Ka; 1-3 x 10(5) 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 (K(diss); 0.002 S(-1)) accumulates 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.