N-Acetylglucosaminyltransferase (GnT) Assays Using Fluorescent Oligosaccharide Acceptor Substrates: GnT-III, IV, V, and IX (GnT-Vb)

Takamatsu, Shinji; Korekane, Hiroaki; Ohtsubo, Kazuaki; Oguri, Suguru; Park, Jong Yi; Matsumoto, Akio; Taniguchi, Naoyuki

doi:10.1007/978-1-62703-465-4_21

Cited by 14 publications

(14 citation statements)

References 20 publications

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“…GnGnbi-PA was prepared as described previously from egg yolk sialylglycopeptide (Fushimi Pharmaceutical) by releasing N-glycans with PNGase F, labelling with 2-aminopyridine, and treating with sialidase and galactosidase (42). To prepare GGnGGnbi-PA (type-2 galactosylated form of GnGnbi-PA), GnGnbi-PA was incubated with purified proteinA-B4GALT1 in 125 mM MES pH 6.2, 10 mM MnCl2, 10 mM UDP-Gal at 37˚C overnight.…”

Section: Preparation Of Pa-labelled Sugarsmentioning

confidence: 99%

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

Nakano

Mishra

Tokoro

et al. 2019

Molecular & Cellular Proteomics

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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.

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Section: Preparation Of Pa-labelled Sugarsmentioning

confidence: 99%

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

Nakano

Mishra

Tokoro

et al. 2019

Molecular & Cellular Proteomics

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“…Enzymatic Reaction and HPLC Analysis. The GnT-V enzymatic activity analyses in T cells from patients with UC and controls were performed as previously described by Takamatsu et al (45).…”

Section: Isolation Of Cd3 + T Cells From Fresh Colonic Biopsies and Bmentioning

confidence: 99%

Metabolic control of T cell immune response through glycans in inflammatory bowel disease

Dias

Correia

Pereira

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

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Mucosal T lymphocytes from patients with ulcerative colitis (UC) were previously shown to display a deficiency in branched N-glycosylation associated with disease severity. However, whether this glycosylation pathway shapes the course of the T cell response constituting a targeted-specific mechanism in UC remains largely unknown. In this study, we demonstrated that metabolic supplementation of ex vivo mucosal T cells from patients with active UC with -acetylglucosamine (GlcNAc) resulted in enhancement of branched N-glycosylation in the T cell receptor (TCR), leading to suppression of T cell growth, inhibition of the T helper 1 (Th1)/Th17 immune response, and controlled T cell activity. We further demonstrated that mouse models displaying a deficiency in the branched N-glycosylation pathway (, ) exhibited increased susceptibility to severe forms of colitis and early-onset disease. Importantly, the treatment of these mice with GlcNAc reduced disease severity and suppressed disease progression due to a controlled T cell-mediated immune response at the intestinal mucosa. In conclusion, our human ex vivo and preclinical results demonstrate the targeted-specific immunomodulatory properties of this simple glycan, proposing a therapeutic approach for patients with UC.

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“…The enzyme acts on biantennary complex-type N-glycan and forms the 3-antennary core structure (Figures 3 and 4). The third branch extends by addition of LacNAc repeats, poly-LacNAcs that bear galectin-binding epitopes (Takamatsu et al, 2010(Takamatsu et al, , 2013Taniguchi and Korekane, 2011). Secreted galectins are bound to β-galactoside sugar on poly-LacNAcs and cross-link glycoconjugates on the cell surface and in the ECM by producing a lattice formation.…”

Section: N-acetylglucosaminyltransferases (Glcnacts Gnts)mentioning

confidence: 99%

“…GnT-IV has 2 isoenzymes, GnT-IVa and GnT-IVb, and they both contribute to the galectin-mediated glycoprotein clustering on the cell surface (Takamatsu et al, 2013). In humans, GnT-IV isoenzymes have the same specificities but different affinities for sugar donors and acceptors.…”

Section: N-acetylglucosaminyltransferases (Glcnacts Gnts)mentioning

confidence: 99%

Glycosylation changes leading to the increase in size on the common core of N-glycans, required enzymes, and related cancer-associated proteins

Karaçalı¹,

İzzetoğlu²,

Deveci³

2014

Turk J Biol

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Glycan parts of glycoconjugates on the surfaces of cells regulate many kinds of interactions between the cells and their immediate environments. Alterations in glycosylation on the cancer-associated glycoproteins are responsible for changes in their molecular interactions and biological functions. Glycosylation changes occur in the core and/or at the nonreducing end of the oligosaccharide chains of N-glycans. In this review, we focus on the branching of the common core structure of N-glycans, the responsible enzyme, and the extensions of some of the branches causing size increases on the surface of tumor cells. Abnormal branching, elongation of the branches, and increasing size of the common core of N-glycans are the typical features of these changes and are related with malignant transformations. Seven N-acetylglucosaminyltransferases (GnTs) (GnT-I, GnT-II, GnT-III, GnT-IV, GnT-V, GnT-VI, and GnT-IX) and α1,6-fucosyltransferase (FUT8) initiate the new branches on the core. GnT-IV, GnT-V, and GnT-IX initiate the branches available for poly-LacNAc extensions, which are responsible for tumor progression and metastasis. GnT-III prevents the catalytic activity of GnT-II, GnT-IV, GnT-V, and FUT8 to form branching and elongation of the branches. The contributions of GnT-III and the other enzymes to the cancer progression are in conflict with each other. While GnT-III prevents cancer, the others increase metastasis. The function of FUT8 is related to signal transduction and its activity is higher in tumor tissue than in healthy tissue. The impact of glycosylation changes on some of the cancer-associated proteins (growth factor receptors, adhesion and signal molecules, CD147, TIMP-1, and matriptase) is also summarized.

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N-Acetylglucosaminyltransferase (GnT) Assays Using Fluorescent Oligosaccharide Acceptor Substrates: GnT-III, IV, V, and IX (GnT-Vb)

Cited by 14 publications

References 20 publications

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

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

Metabolic control of T cell immune response through glycans in inflammatory bowel disease

Glycosylation changes leading to the increase in size on the common core of N-glycans, required enzymes, and related cancer-associated proteins

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