Poly-N-acetyllactosamine Extension inN-Glycans and Core 2- and Core 4-branchedO-Glycans Is Differentially Controlled by i-Extension Enzyme and Different Members of the β1,4-Galactosyltransferase Gene Family

Abstract

Poly-N-acetyllactosamines are attached to N-glycans, O-glycans, and glycolipids and serve as underlying glycans that provide functional oligosaccharides such as sialyl Lewis X . Poly-N-acetyllactosaminyl repeats are synthesized by the alternate addition of ␤1,3-linked GlcNAc and ␤1,4-linked Gal by i-extension enzyme (iGnT) and a member of the ␤1,4-galactosyltransferase (␤4Gal-T) gene family. In the present study, we first found that poly-N-acetyllactosamines in N-glycans are most efficiently synthesized by ␤4Gal-TI and iGnT. We also found that iGnT acts less efficiently on acceptors containing increasing numbers of N-acetyllactosamine repeats, in contrast to ␤4Gal-TI, which exhibits no significant change. In O-glycan biosynthesis, N-acetyllactosamine extension of core 4 branches was found to be synthesized most efficiently by iGnT and ␤4Gal-TI, in contrast to core 2 branch synthesis, which requires iGnT and ␤4Gal-TIV. Poly-N-acetyllactosamine extension of core 4 branches is, however, less efficient than that of N-glycans or core 2 branches. Such inefficiency is apparently due to competition between a donor substrate and acceptor in both galactosylation and N-acetylglucosaminylation, since a core 4-branched acceptor contains both Gal and GlcNAc terminals. These results, taken together, indicate that poly-N-acetyllactosamine synthesis in N-glycans and core 2-and core 4-branched Oglycans is achieved by iGnT and distinct members of the ␤4Gal-T gene family. The results also exemplify intricate interactions between acceptors and specific glycosyltransferases, which play important roles in how poly-Nacetyllactosamines are synthesized in different acceptor molecules.Poly-N-acetyllactosamines are unique glycans having N-acetyllactosamine repeats (Gal␤134GlcNAc␤133) n in one side chain (1). Poly-N-acetyllactosamines are attached to Nglycans (2-4), O-glycans (5-7), and glycolipids (8 -10). Poly-Nacetyllactosamines are often modified to express differentiation antigens and functional oligosaccharides. One of those oligosaccharides is sialyl Le X , 1 NeuNAc␣233Gal␤134(Fuc␣133)-GlcNAc3 R discovered in human granulocytes and monocytes (11,12). Sialyl Le X and its sulfated forms, such as 6-sulfo sialyl Le X , NeuNAc␣233Gal␤134[Fuc␣133(sulfo36)]GlcNAc3 R in mucin-type glycoproteins, have been shown to be ligands for E-, P-, and L-selectin (13-15).Since these O-glycans are present as clusters in mucin-type glycoproteins, mucin-type glycoproteins can present multiple ligands to a selectin. In mucin-type glycoproteins of blood cells, sialyl Le X can be found in core 2-branched oligosaccharides (5, 6, 16). Similarly, 6-sulfo sialyl Le X in L-selectin ligands found in high endothelial venules are synthesized in core 2-branched oligosaccharides such as NeuNAc␣233Gal␤134[Fuc␣133-(sulfo36)]GlcNAc␤136(Gal␤133)GalNAc␣13serine/threonine (17-19).The enzyme responsible for core 2 branching is called core 2 ␤1,6-N-acetylglucosaminyltransferase (C2GnT), and its cDNA has been cloned (20). When C2GnT was inactivated by gene targeting, leukocytes...

β-1,4-Galactosyltransferase III Enhances Invasive Phenotypes Via β1-Integrin and Predicts Poor Prognosis in Neuroblastoma

“…Poly-N-acetyllactosamine structures have been found on b1 subunit of a3b1-integrin (22, 23), a receptor for laminin, fibronectin, and collagen. In vitro enzyme activity assays indicate that B4GALT3 catalyzes the formation of poly-Nacetyllactosamine (13). Consistent with these results, we found that B4GALT3 changes the amount of poly-N-acetyllactosamine on b1-integrin as revealed by altered RCA I and LEL binding.…”

“…B4GALT3 is widely expressed in human tissues and the fetal brain expresses much higher levels of B4GALT3 than does adult brain (11,12). Previous in vitro study showed that B4GALT3 prefers adding galactose to the beginning of a poly-N-acetyllactosamine chain (13). However, roles of B4GALT3 in NB and its effects on NB tumor cells are still unclear.…”

Dendritic Cell Maturation Results in Pronounced Changes in Glycan Expression Affecting Recognition by Siglecs and Galectins

“…The CRDs of galectin-4 bind to the 3-O-sulfated glycolipids SM4, SM3, and SB1a, but also to some glycoproteins (44). iDC express relatively high levels of the transcripts for the enzymes ␤4GalT-1, -4, and iGnT, previously described to be involved in the expression of poly-N-acetyllactosamine chains (45), which are clearly detected by MS analyses of the N-glycans. This observation could explain the high binding of galectin-3 and -8 to iDC.…”

Keratan Sulfate Biosynthesis

“…Comparison of substrate specificity of recombinant forms of these enzymes has shown the β4Gal-T1 enzyme to be most efficient at synthesis of linear polylactosamine chains in N -linked oligosaccharides; however, the transferase β4Gal-T4 appears to be essential in generating the initial branches found in core-2 O-linked polylactosamine (42). The structural identity between N -linked polylactosamine and KSI chains suggests that the same enzymes are likely to be involved in their elongation, adding to the body of evidence pointing to β4Gal-T1 as the galactosyltransferase involved in keratan sulfate elongation.…”