Bisecting GlcNAc Is a General Suppressor of Terminal Modification of N-glycan*[S]

Abstract

Glycoproteins are decorated with complex glycans for protein functions. However, regulation mechanisms of complex glycan biosynthesis are largely unclear. Here we found that bisecting GlcNAc, a branching sugar residue in N-glycan, suppresses the biosynthesis of various types of terminal epitopes in N-glycans, including fucose, sialic acid and human natural killer-1. Expression of these epitopes in N-glycan was elevated in mice lacking the biosynthetic enzyme of bisecting GlcNAc, GnT-III, and was conversely suppressed by GnT-III overexpression in cells. Many glycosyltransferases for N-glycan terminals were revealed to prefer a nonbisected N-glycan as a substrate to its bisected counterpart, whereas no up-regulation of their mRNAs was found. This indicates that the elevated expression of the terminal N-glycan epitopes in GnT-III-deficient mice is attributed to the substrate specificity of the biosynthetic enzymes. Molecular dynamics simulations further confirmed that nonbisected glycans were preferentially accepted by those glycosyltransferases. These findings unveil a new regulation mechanism of protein N-glycosylation.

Discovery of Pancreatic Ductal Adenocarcinoma-Related Aberrant Glycosylations: A Multilateral Approach of Lectin Microarray-Based Tissue Glycomic Profiling With Public Transcriptomic Datasets

“…The alterations in gene expression and glycome levels are also consistent with a previous report showing the increased degree of MGAT3 and MGAT5 activities in the tissues of pancreatic carcinoma patients (45). Notably, a more recent study has demonstrated that the introduction of bisecting GlcNAc suppresses terminal modifications of N-glycans such as fucosylation and sialylation in genetically modified mice (46). The present tissue glycomic profiling results are consistent with this previous report in that the increased level of bisecting GlcNAc was accompanied by decreased fucosylation.…”

3D Structure and Function of Glycosyltransferases Involved in N-glycan Maturation

“…The second feature of bisecting GlcNAc is its inhibitory effects on other glycosyltransferases. The enzymes responsible for producing other N-glycan branches (e.g., GnT-IV, GnT-V, and FUT8) as well as the enzymes acting on N-glycan terminals (fucosyltransferases, sialyltransferases and biosynthetic enzymes for human natural killer-1 epitope) are inhibited partially or completely by the presence of a bisecting GlcNAc in N-glycan [54][55][56]. This is probably because the presence of bisecting GlcNAc alters glycan conformation and restricts preferable conformers of N-glycan.…”

“…Molecular dynamic simulation, NMR analysis, and X-ray crystallography suggest that bisected N-glycans tend to prefer back-fold conformations in which the α1-6 branch flips back to the reducing end [56][57][58][59][60]. Our MS glycan analysis revealed increases in various terminal epitopes of N-glycans in Mgat3-knockout brain, which is concomitant with complete loss of bisecting GlcNAc, suggesting that one of the physiological functions of bisecting GlcNAc is to suppress formation of mature and complex N-glycans [56]. Similarly, Dr. Gu's group reported that the level of bisecting GlcNAc is negatively correlated with the level of sialylation in various cell lines [61].…”

“…When a bisecting GlcNAc residue is added on β-mannose, this residue lies close to OH6 of the α1-6 branched mannose (Man-2) and sterically clashes with Man-2. In addition, bisected glycan prefers to take extend-b and back-fold conformations, rather than the extend-a conformation that is observed in the complex structure [56][57][58][59][60]. Structural superposition of these two conformations shows that they cannot fit into the surface groove of GnT-V without severe steric clashes.…”

Abberant Immunoglobulin G Glycosylation in Rheumatoid Arthritis by LTQ-ESI-MS

“…Recently, bisecting GlcNAc has been recognized as a suppressor of terminal modifications of N-glycan, which may further influence biological functions of glycans on glycoproteins [31]. However, the effects of bisecting GlcNAc on functional properties of IgG are not well understood.…”