Role of the Carboxyl-Terminal Lectin Domain in Self-Association of Galectin-3

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Abstract
Galectin-3 is a member of a large family of β-galactoside-binding animal lectins and is composed of a carboxyl-terminal lectin domain connected to an amino-terminal nonlectin part. Previous experimental results suggest that, when bound to multivalent glycoconjugates, galectin-3 self-associates through intermolecular interactions involving the amino-terminal domain. In this study, we obtained evidence suggesting that the protein self-associates in the absence of its saccharide ligands, in a manner that is dependent on the carboxyl-terminal domain. This mode of self-association is inhibitable by the lectin's saccharide ligands. Specifically, recombinant human galectin-3 was found to bind to galectin-3C (the carboxyl-terminal domain fragment) conjugated to Sepharose 4B and the binding was inhibitable by lactose. In addition, biotinylated galectin-3 bound to galectin-3 immobilized on plastic surfaces and the binding could also be inhibited by various saccharide ligands of the lectin. A mutant with a tryptophan to leucine replacement in the carboxyl-terminal domain, which exhibited diminished carbohydrate-binding activity, did not bind to galectin-3C−Sepharose 4B. Furthermore, galectin-3C formed covalent homodimers when it was treated with a chemical cross-linker and the dimer formation was completely inhibited by lactose. Therefore, galectin-3 can self-associate through intermolecular interactions involving both the amino- and the carboxyl-terminal domains and the relative contribution of each depends on whether the lectin is bound to its saccharide ligands.