The intrinsically disordered N-terminal domain of galectin-3 dynamically mediates multisite self-association of the protein through fuzzy interactions

Abstract

Galectins are a family of lectins that bind -galactosides through their conserved carbohydrate recognition domain (CRD) and can induce aggregation with glycoproteins or glycolipids on the cell surface and thereby regulate cell activation, migration, adhesion, and signaling. Galectin-3 has an intrinsically disordered N-terminal domain and a canonical CRD. Unlike the other 14 known galectins in mammalian cells, which have dimeric or tandem-repeated CRDs enabling multivalency for various functions, galectin-3 is monomeric, and its functional multivalency therefore is somewhat of a mystery. Here, we used NMR spectroscopy, mutagenesis, small-angle X-ray scattering, and computational modeling to study the self-association–related multivalency of galectin-3 at the residue-specific level. We show that the disordered N-terminal domain (residues 20 –100) interacts with itself and with a part of the CRD not involved in carbohydrate recognition (-strands 7–9; residues 200 –220), forming a fuzzy complex via inter- and intramolecular interactions, mainly through hydrophobicity. These fuzzy interactions are characteristic of intrinsically disordered proteins to achieve liquid–liquid phase separation, and we demonstrated that galectin-3 can also undergo liquid–liquid phase separation. We propose that galectin-3 may achieve multivalency through this multisite self-association mechanism facilitated by fuzzy interactions.