Interaction profile of galectin-5 with free saccharides and mammalian glycoproteins:: probing its fine specificity and the effect of naturally clustered ligand presentation

Cell-surface glycans are functional docking sites for tissue lectins such as the members of the galectin family. This interaction triggers a wide variety of responses; hence, there is a keen interest in defining its structural features. Toward this aim, we have used enzyme-linked lectinosorbent (ELLSA) and inhibition assays with the prototype rat galectin-5 and panels of free saccharides and glycoconjugates. Among 45 natural glycans tested for lectin binding, galectin-5 reacted best with glycoproteins (gps) presenting a high density of Gal beta 1-3/4GlcNAc (I/II) and multiantennary N-glycans with II termini. Their reactivities, on a nanogram basis, were up to 4.3 x 10(2), 3.2 x 10(2), 2.5 x 10(2), and 1.7 x 10(4) times higher than monomeric Gal beta 1-3/4GlcNAc (I/II), triantennary-II (Tri-II), and Gal, respectively. Galectin-5 also bound well to several blood group type B Gal alpha 1-3Gal)- and A (GalNAc alpha 1-3Gal)-containing gps. It reacted weakly or not at all with tumor-associated Tn (GalNAc alpha 1-Ser/Thr) and sialylated gps. Among the mono-, di-, and oligosaccharides and mammalian glycoconjugates tested, blood group B-active II (Gal alpha 1-3Gal beta 1-4GlcNAc), B-active II beta 1-3L (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc), and Tri-II were the best. It is concluded that (1) Gal beta 1-3/4GlcNAc and other Gal beta 1-related oligosaccharides with alpha 1-3 extensions are essential for binding, their polyvalent form in cellular glycoconjugates being a key recognition force for galectin-5; (2) the combining site of galectin-5 appears to be of a shallow-groove type sufficiently large to accommodate a substituted beta-galactoside, especially with alpha-anomeric extension at the non-reducing end (e.g., human blood group B-activeII and B-active II beta 1-3L); (3) the preference within beta-anomeric positioning is Gal beta 1-4 >= Gal beta 1-3 > Gal beta 1-6; and (4) hydrophobic interactions in the vicinity of the core galactose unit can enhance binding. These results are important for the systematic comparison of ligand selection in this family of adhesion/growth-regulatory effectors with potential for medical applications.