β-D-Manp-(1->4)-β-D-GlcpNAc-(1->4)-D-GlcpNAc

• 分子结构分类: 有机化合物 - 有机氧化合物
• 英文名称: β-D-Manp-(1->4)-β-D-GlcpNAc-(1->4)-D-GlcpNAc
• 英文别名: Manβ(1-4)GlcNAcβ(1-4)GlcNAc；Gn2-Man；O-β-D-mannopyranosyl-(1[*]4)-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-(1[*]4)-2-acetamido-2-deoxy-D-glucopyranose；Man(b1-4)GlcNAc(b1-4)GlcNAc；N-[(3R,4R,5S,6R)-5-[(2S,3R,4R,5S,6R)-3-acetamido-4-hydroxy-6-(hydroxymethyl)-5-[(2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyoxan-2-yl]oxy-2,4-dihydroxy-6-(hydroxymethyl)oxan-3-yl]acetamide
• 化学式: C₂₂H₃₈N₂O₁₆
• 分子量: 586.548
• InChiKey: OCPQLCXSCTTXMX-WODXAAMMSA-N

计算性质

• 辛醇/水分配系数(LogP)：
  -6.9
• 重原子数：
  40
• 可旋转键数：
  9
• 环数：
  3.0
• sp3杂化的碳原子比例：
  0.91
• 拓扑面积：
  286
• 氢给体数：
  11
• 氢受体数：
  16

反应信息

• 作为反应物：
  描述：

  β-D-Manp-(1->4)-β-D-GlcpNAc-(1->4)-D-GlcpNAc 在 Helix pomatia β-mannosidase 作用下， 以 aq. buffer 为溶剂， 反应 16.0h， 生成 N,N'-Diacetylchitobiose
  参考文献：
  名称：

  Quantitative study of yeast Alg1 beta-1, 4 mannosyltransferase activity, a key enzyme involved in protein N-glycosylation

  摘要：

  Background: Asparagine (N)-linked glycosylation begins with a stepwise synthesis of the dolichol-linked oligosaccharide (DLO) precursor, Glc3Man9GlcNAc2-PP-Dol, which is catalyzed by a series of endoplasmic reticulum membrane-associated glycosyltransferases. Yeast ALG1 (asparagine-linked glycosylation 1) encodes a beta-1, 4 mannosyltransferase that adds the first mannose onto GlcNAc2-PP-Dol to produce a core trisaccharide Man1GlcNAc2-PP-Dol.ALG1 is essential for yeast viability, and in humans mutations in the ALG1 cause congenital disorders of glycosylation known as ALG1-CDG. Alg1 is difficult to purify because of its low expression level and as a consequence, has not been well studied biochemically. Here we report a new method to purify recombinant Alg1 in high yield, and a mass spectral approach for accurately measuring its beta-1, 4 mannosyltransferase activity.Methods: N-terminally truncated yeast His-tagged Alg1 protein was expressed in Escherichia coli and purified by HisTrap HP affinity chromatography. In combination with LC-MS technology, we established a novel assay to accurately measure Alg1 enzyme activity. In this assay, a chemically synthesized dolichol-linked oligosaccharide analogue, phytanyl-pyrophosphoryl-alpha-N,N'-diacetylchitobioside (PPGn2), was used as the acceptor for the beta-1, 4 mannosyl transfer reaction.Results: Using purified Alg1, its biochemical characteristics were investigated, including the apparent K-m and V-max values for acceptor, optimal conditions of activity, and the specificity of its nucleotide sugar donor. Furthermore, the effect of ALG1-CDG mutations on enzyme activity was also measured.General significance: This work provides an efficient method for production of Alg1 and a new MS-based quantitative assay of its activity. (C) 2016 Elsevier B.V. All rights reserved.

  DOI：

  10.1016/j.bbagen.2016.09.023
• 作为产物：
  描述：

  guanosine 5'-diphospho-D-mannose 在 盐酸 、 乙二胺四乙酸 、 dithiothreitol 、 recombinant His6-tagged S. cerevisiae Alg1 Δtransmembrane-spanning domain 、 magnesium chloride 、 NP-40 作用下， 以 甲醇 为溶剂， 反应 1.5h， 生成 β-D-Manp-(1->4)-β-D-GlcpNAc-(1->4)-D-GlcpNAc
  参考文献：
  名称：

  Quantitative study of yeast Alg1 beta-1, 4 mannosyltransferase activity, a key enzyme involved in protein N-glycosylation

  摘要：

  Background: Asparagine (N)-linked glycosylation begins with a stepwise synthesis of the dolichol-linked oligosaccharide (DLO) precursor, Glc3Man9GlcNAc2-PP-Dol, which is catalyzed by a series of endoplasmic reticulum membrane-associated glycosyltransferases. Yeast ALG1 (asparagine-linked glycosylation 1) encodes a beta-1, 4 mannosyltransferase that adds the first mannose onto GlcNAc2-PP-Dol to produce a core trisaccharide Man1GlcNAc2-PP-Dol.ALG1 is essential for yeast viability, and in humans mutations in the ALG1 cause congenital disorders of glycosylation known as ALG1-CDG. Alg1 is difficult to purify because of its low expression level and as a consequence, has not been well studied biochemically. Here we report a new method to purify recombinant Alg1 in high yield, and a mass spectral approach for accurately measuring its beta-1, 4 mannosyltransferase activity.Methods: N-terminally truncated yeast His-tagged Alg1 protein was expressed in Escherichia coli and purified by HisTrap HP affinity chromatography. In combination with LC-MS technology, we established a novel assay to accurately measure Alg1 enzyme activity. In this assay, a chemically synthesized dolichol-linked oligosaccharide analogue, phytanyl-pyrophosphoryl-alpha-N,N'-diacetylchitobioside (PPGn2), was used as the acceptor for the beta-1, 4 mannosyl transfer reaction.Results: Using purified Alg1, its biochemical characteristics were investigated, including the apparent K-m and V-max values for acceptor, optimal conditions of activity, and the specificity of its nucleotide sugar donor. Furthermore, the effect of ALG1-CDG mutations on enzyme activity was also measured.General significance: This work provides an efficient method for production of Alg1 and a new MS-based quantitative assay of its activity. (C) 2016 Elsevier B.V. All rights reserved.

  DOI：

  10.1016/j.bbagen.2016.09.023

文献信息

• Purification of the β-N-acetylhexosaminidase from Aspergillus oryzae and the β-mannosidases from Helix pomatia and A. oryzae and their application to the enzymic synthesis of the core trisaccharide of the N-linked glycoproteins
  作者：Michaela Scigelova、Suddham Singh、David H. G. Crout

  DOI：10.1039/a900773c

  日期：——

  The β-N-acetylhexosaminidase from Aspergillus oryzae and the β-mannosidases from Helix pomatia and A. oryzae were purified and used to synthesise the core trisaccharide of N-linked glycoproteins, Manp-(1→4)-β-D-GlcpNAc-(1→4)-D-GlcpNAc.

  从黑曲霉（Aspergillus oryzae）中分离纯化出δ-N-乙酰己糖胺酶，并从Helix pomatia和黑曲霉（A. oryzae）中分离纯化出δ-甘露糖苷酶，用于合成N-连接糖蛋白的核心三糖--Manp-(1â4)-δ-D-GlcpNAc-(1â4)-D-GlcpNAc。
• The synthesis of O-β-d-mannopyranosyl-(1→4)-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-(1→4)-2-acetamido-2-deoxy-D-glucopyranose. Part I
  作者：Christopher D. Warren、Claudine Augé、Murray L. Laver、Shigeo Suzuki、Diane Power、Roger W. Jeanloz

  DOI：10.1016/s0008-6215(00)85521-4

  日期：1980.6

  allyl O-(2-O-acetyl-3,4,6-tri-O-benzyl-β- D -glucopyranosyl)-(1→4)-O-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β- D -glucopyranosyl)-(1→4)-2-acetamido-3,6-di-O-(2-butenyl)-2-deoxy-β- D -glucopyranoside. O-Deacetylation, oxidation with acetic anhydride-dimethyl sulfoxide, and stereoselective reduction with sodium borohydride gave mainly allyl O-(3,4,6-tri-O-benzyl-β- D -mannopyranosyl)-(1→4)-O-(2-acetamido-3

  摘要制备了2-乙酰氨基-3,6-二-O-（2-丁烯基）-2-脱氧-β-D-吡喃葡萄糖苷烯丙基，并与2-甲基-（4-O-乙酰基-3,6-二-O-苄基-1,2-二脱氧-α-D-吡喃葡萄糖）-[2,1-d] -2-恶唑啉。所得的受保护的二糖烯丙基2-乙酰氨基-4-O-（2-乙酰氨基-4-O-乙酰基-3,6-二-O-苄基-2-脱氧-β-D-吡喃葡萄糖基）-3,6-将二-O-（2-丁烯基）-2-脱氧-β-D-吡喃葡萄糖苷进行O-脱乙酰化，并将产物与2-O-乙酰基-3,4,6-tri-O-苄基-α-D-偶联。在三氟甲磺酸银和1，1,3,3-四甲基脲的存在下，将吡喃葡萄糖基溴化，得到三糖烯丙基O-（2-O-乙酰基-3,4,6-tri-O-苄基-β-D-吡喃葡萄糖基）-（1→4）-O-（2-乙酰氨基-3,6-二-O-苄基-2-脱氧-β-D-吡喃葡萄糖基）-（1→4）-2-乙酰氨基-3,6 -二-O-（2-
• Glycosidase-catalysed synthesis of oligosaccharides: a two-step synthesis of the core trisaccharide of N-linked glycoproteins using the β-N-acetylhexosaminidase and the β-mannosidase from Aspergillus oryzae
  作者：Suddham Singh、Michaela Scigelova、David H. G. Crout

  DOI：10.1039/cc9960000993

  日期：——

  Using a partially purified beta-mannosidase from Aspergillus oryzae, a beta-mannosyl unit is transferred from p-nitrophenyl beta-D-mannopyranoside 5 to di-N-acetylchitobiose 4 to give the core trisaccharide beta-D-Manp-(1-->4)-beta-D-GlcpNAc-(1-->4)-D-GlcpNAc 6 of the N-linked glycoproteins.
• Efficient Enzymatic Synthesis of the Core Trisaccharide ofN-Glycans with a Recombinantβ-Mannosyltransferase
  作者：Gregory M. Watt、Leigh Revers、Matthew C. Webberley、Iain B. H. Wilson、Sabine L. Flitsch

  DOI：10.1002/anie.199723541

  日期：1997.11.14
• The synthesis of O-β-D-mannopyranosyl-(1→4)-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-(1→4)-2-acetamido-2-deoxy-D-glucopyranose. part II
  作者：Claudine Augé、Christopher D. Warren、Roger W. Jeanloz、Makoto Kiso、Laurens Anderson

  DOI：10.1016/s0008-6215(00)85522-6

  日期：1980.6