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Galectins are a family of lectins which share similar carbohydrate recognition domains (CRDs) and affinity for small ␤-galactosides, but which show significant differences in binding specificity for more complex glycoconjugates. We report here the x-ray crystal structure of the human galectin-3 CRD, in complex with lactose and N-acetyllactosamine, at 2.1-Å resolution. This structure represents the first example of a CRD determined from a galectin which does not show the canonical 2-fold symmetric dimer organization. Comparison with the published structures of galectins-1 and -2 provides an explanation for the differences in carbohydrate-binding specificity shown by galectin-3, and for the fact that it fails to form dimers by analogous CRD-CRD interactions.Galectin-3 is a member of the galectin family of lectins defined by a conserved ϳ14-kDa carbohydrate recognition domain (CRD) 1 showing affinity for ␤-galactosides (1, 2). Abundantly expressed in a few cell types, such as macrophages and polarized epithelial cells in adults (2, 3) and others during embryogenesis (4), it tends to be localized in the cytoplasm and the nucleus. Although functions for galectin-3 have been proposed in each of these subcellular locations (5-7), it is also secreted by a nonclassical pathway (8, 9) and is found on the cell surface and in the extracellular matrix. There it binds and cross-links selected carbohydrate-containing ligands (10,11) and is thought to modulate cell adhesion (12-14) and cell signaling (15, 16). Many groups are currently studying the roles and uses of galectin-3 in cancer, inflammation, hostpathogen interaction, and nerve injury, among others (17, 18).Galectin-3 is unique among the known galectins in that, in addition to the canonical CRD (located at the C terminus), it contains an unrelated, non-carbohydrate-binding N-terminal domain of between 120 (in human) and 166 (in dog) amino acids (3,19). In contrast, galectins-1 and -2 are homodimers composed of the CRD alone, while galectins-4, -6, -8, and -9 possess an N-and C-terminal CRD linked in tandem by a short polypeptide segment (18). The galectin-3 CRD shows sequence identity ranging from 30 -40% with galectins-4 through -10, to 20 -25% with galectins-1 and -2. It has an affinity for lactose (Lac, K d ϭ 1 mM), and N-acetyllactosamine (LacNAc, K d ϭ 0.2 mM) similar to that of other galectins, but has a distinct profile for larger oligosaccharides (20, 21), including polyNAc-lactosaminoglycan, a polymer of ␤(1,3)-linked LacNAc units found on many extracellular matrix and cell surface molecules.Intact galectin-3, but not its CRD alone, shows avidity for multivalent glycoconjugates (10, 11), modulates cell adhesion (14), and induces intracellular signals (15). Thus it is thought that the N-terminal domain of galectin-3 promotes the formation of dimers or higher order oligomers, thereby permitting multivalent interactions essential for its biological activities.We report here the x-ray crystal structure of the CRD of human galectin-3 in complex with Lac and LacNA...

Abstract. A soluble lactose-binding lectin with subunit Mr of 14,500 is believed to function by interacting with extracellular glycoconjugates, because it has been detected extracellularly by immunohistochemistry. This localization has been questioned, however, since the lectin lacks a secretion signal sequence, which challenges the contention that it is secreted. We have demonstrated externalization of this lectin from C2 mouse muscle cells by both immunoprecipitation of metabolically labeled protein and immunohistochemical localization. We further show that externalization of the lectin is a developmentally regulated process that accompanies myoblast differentiation and that the lectin codistributes with laminin in myotube extracellular matrix. Immunohistochemical localization during intermediate stages of externalization suggests that the lectin becomes concentrated in evaginations of plasma membrane, which pinch off to form labile lectin-rich extracellular vesicles. This suggests a possible mechanism for lectin export from the cytosol to the extracellular matrix.

The soluble mammalian lactose-binding lectins L-14-I and L-29 are both secreted and bind to oligosaccharides on laminin, a large extracellular matrix glycoprotein containing polylactosamine chains. Because of the potential functional significance of these lectin-laminin interactions, we compared quantitative aspects of L-14-I and L-29 binding to immobilized laminin using recombinant lectins labeled with 125I. We report that the concentration-dependent binding of L-29 exhibits positive cooperativity whereas binding of L-14-I does not. Cooperative binding of L-29 can also occur on glycoconjugate substrates other than laminin and is not dependent on cystine bond formation or aggregation in solution. L-29 contains repetitive sequences within the N-terminal domain not present in L-14-I. This domain is not required for binding activity, but is required for positive cooperativity. Though the precise mechanism of interaction of L-29 with laminin remains to be determined, it apparently results in assembly of a lectin aggregate on the substrate surface, which could have important functional consequences.

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