Human galectin-1 recognition of poly-N-acetyllactosamine and chimeric polysaccharides

Abstract

Human galectin-1 is a dimeric carbohydrate binding protein (Gal-1) (subunit 14.6 kDa) widely expressed by many cells but whose carbohydrate binding specificity is not well understood. Because of conflicting evidence regarding the ability of human Gal-1 to recognize N-acetyllactosamine (LN, Galbeta4GlcNAc) and poly-N-acetyllactosamine sequences (PL, [-3Galbeta4GlcNAcbeta1-]n), we synthesized a number of neoglycoproteins containing galactose, N-acetylgalactosamine, fucose, LN, PL, and chimeric polysaccharides conjugated to bovine serum albumin (BSA). All neoglycoproteins were characterized by MALDI-TOF. Binding was determined in ELISA-type assays with immobilized neoglycoproteins and apparent binding affinities were estimated. For comparison, we also tested the binding of these neoglycoconjugates to Ricinus communis agglutinin I, (RCA-I, a galactose-binding lectin) and Lycopersicon esculentum agglutinin (LEA, or tomato lectin), a PL-binding lectin. Gal-1 bound to immobilized Galbeta4GlcNAcbeta3Galbeta4Glc-BSA with an apparent K(d) of approximately 23 micro M but bound better to BSA conjugates with long PL and chimeric polysaccharide sequences (K(d)'s ranging from 11.9 +/- 2.9 microM to 20.9 +/- 5.1 micro M). By contrast, Gal-1 did not bind glycans lacking a terminal, nonreducing unmodified LN disaccharide and also bound very poorly to lactosyl-BSA (Galbeta4Glc-BSA). By contrast, RCA bound well to all glycans containing terminal, nonreducing Galbeta1-R, including lactosyl-BSA, and bound independently of the modification of the terminal, nonreducing LN or the presence of PL. LEA bound with increasing affinity to unmodified PL in proportion to chain length. Thus Gal-1 binds terminal beta4Gal residues, and its binding affinity is enhanced significantly by the presence of this determinant on long-chain PL or chimeric polysaccharides.

Pre-B cell receptor binding to galectin-1 modifies galectin-1/carbohydrate affinity to modulate specific galectin-1/glycan lattice interactions

“…1c) and the a3 sialyl-N-Acetyllactosamine (a3 SiaLacNAc) belonging to group (ii) (Fig. 1c), for which a relative decrease and increase in GAL1 binding, respectively, are observed in the presence of the l5-UR domain (binding of these two glycans to GAL1 has been demonstrated previously 31,32 ). In our experimental conditions, the dissociation constant (K D ) of GAL1/LNnT interaction obtained is 51±6 mM, whereas when GAL1 is bound to the l5-UR domain, the K D increases to 130±10 mM, that is, a 2.5-fold decrease in binding affinity of LNnT for GAL1 in the presence of l5-UR (Supplementary Table 1).…”

Dimeric Galectin-1 Binds with High Affinity to α2,3-Sialylated and Non-sialylated Terminal N-Acetyllactosamine Units on Surface-bound Extended Glycans

“…By contrast, it was reported that bovine dGal-1 binds with relatively high affinity to immobilized extended (neo)glycolipids in TLC overlay and microwell binding assays, whereas binding to nonextended structures was not detected (42). Similarly, human dGal-1 is able to recognize immobilized neoglycoproteins containing PLs with higher affinity compared with immobilized neoglycoproteins with non-extended oligosaccharides in a solidphase assay (43). The apparent discordance of these results may now be resolved since we have shown that they reflect the differing affinity of Gal-1 for ligands free in solution or immobilized on surfaces.…”

“…In contrast to these studies, recent data have suggested that dGal-1 may not recognize PL structures with higher affinity compared with its recognition of short non-extended structures (44,45). Interestingly, dGal-1 has been suggested to bind primarily to nonreducing terminal LN residues rather than to mid-chain LN residues within a PL chain (42,43,46).…”

“…However, the ability of human or bovine dGal-1 to bind to ␣2,3-sialylated and ␣2,6-sialylated PL structures has not been compared simultaneously under the same conditions. It has also been reported that ␣1,2-fucosylated (but not ␣1,3/4-fucosylated) structures are recognized by dGal-1 (42,43,47).…”

“…Other glycoprotein ligands for dGal-1 have also been identified, and they may also contain PL sequences (36 -39). dGal-1 also binds to glycopeptides containing PL sequences (40,41), glycolipids (42), and neoglycoproteins (43). In contrast to these studies, recent data have suggested that dGal-1 may not recognize PL structures with higher affinity compared with its recognition of short non-extended structures (44,45).…”