Practical Application of Fluorescence Labeling to Component Sugar Analysis of Glycoproteins
Practical Application of Fluorescence Labeling to Component Sugar Analysis of Glycoproteins
批准号:
62880019
负责人:
HASE Sumihiro
金额:
$8.77万
依托单位:
依托单位国家:
日本
项目类别:
Grant-in-Aid for Developmental Scientific Research
财政年份:
1987
资助国家:
日本
项目状态:
已结题
起止时间:
1987 至 1988
中文摘要
报道了一种用pyridylamination法测定糖蛋白组分糖的方法(H. Takemoto, et al. 1985)。该方法包括糖蛋白的酸水解,氨基的n -乙酰化,用NaBH_3CN, 2-氨基吡啶和HCl对单糖的还原端进行吡啶层化,用凝胶色谱(HPLC)纯化糖的吡啶氨基(PA-)衍生物,用反相HPLC分析PA-糖。为研制全自动糖分析仪,改进了吡啶层合工艺条件。1)挥发性试剂,如二甲胺。用硼烷络合物和2-氨基吡啶-乙酸代替NaBH_3CN和2-氨基吡啶-盐酸。2)选择反应条件获得pa -糖的最大产率(中性糖70%,n -乙酰氨基糖87%)。3)开发了减压氮气流,使试剂有效蒸发。利用新的反应条件和蒸发过程,可以省去凝胶色谱法(HPLC)纯化pa -糖。采用自动进样器和数据处理机对合成的pa糖进行高效液相色谱分析。糖蛋白酸水解产物的糖用量为1 ~ 10 nmol,得到了合理的结果。
英文摘要
A method for determination of component sugars of glycoproteins by pyridylamination was reported (H. Takemoto, et al. 1985). The method includes acid hydrolysis of glycoprotein, N-acetylation of amino groups, pyridylamination of the reducing ends of the monosaccharides with NaBH_3CN, 2-aminopyridine, and HCl, purification of pyridylamino (PA-) derivatives of sugars by gel chromatography (HPLC), and the analysis of PA-sugars by reversed-phase HPLC. The pyridylamination conditions were improved for the purpose of the making of an automatic sugar analyzer. 1) Volatile reagents such as dimethylamine. borane complex, and 2-aminopyridine-acetic acid were used intead of NaBH_3CN and 2-aminopyridine-HCl. 2) The reaction conditions were chosen to obtain maximum yield of PA-sugars (neutral sugars 70% and N-acetylaminosugars 87%). 3) Nitrogen flow under reduced pressure was developed to evaporate the reagents effectively. Using the new reaction conditions and the evaporation procedre, purification of PA-sugars by gel chromatography (HPLC) could be omitted. An automatic pyridylamination apparatus using the new conditions was constructed and PA-sugars thus obtained were analyzed by HPLC equipped with an automatic injector and a data processor. Reasonable results were obtained by using 1-10 nmol of sugars of acid hydrolysates of glycoproteins.
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S. Hase: "Analysis of Tissue Glycoprotein Sugar Chains by Two-Dimensional High-Performance Liquid Chromatographic Mapping" J. Chromatogr.434. 51-60 (1988)
S. Hase:“通过二维高效液相色谱图分析组织糖蛋白糖链”J. Chromatogr.434。
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S.Hase: Anal.Biochem.67. 321-326 (1987)
S.Hase:Anal.Biochem.67。
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S. Hase: "A New Trisaccharide Sugar Chain Linked to a Serine Residue in Bovine Blood Coagulation Factors VII and IX" J. Biochem.104. 867-868 (1988)
S. Hase:“与牛凝血因子 VII 和 IX 中的丝氨酸残基相连的新三糖糖链”J. Biochem.104。
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S.Hase: J.Biochem.104. 867-868 (1988)
S.Hase:J.Biochem.104。
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S.Yamamoto: J.Biochem.105. inpress (1989)
S.Yamamoto:J.Biochem.105。
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共 16 条
Analysis of Oligosaccharide Structures from the Reducing End Terminal by Combining Partial Acid Hydrolysis and a Two-Dimensional Sugar Map
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批准号:10558100
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项目类别:Grant-in-Aid for Scientific Research (B)
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资助金额:$5.06万
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财政年份:1998
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负责人:HASE Sumihiro
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依托单位:
Analysis of activation of neutral alpha-mannosidase activated with Co(II)
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批准号:05453211
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项目类别:Grant-in-Aid for General Scientific Research (B)
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资助金额:$4.74万
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财政年份:1993
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负责人:HASE Sumihiro
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依托单位:
Search for xylosylglucose-type sugar chains by pyridylamination
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批准号:03453164
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项目类别:Grant-in-Aid for General Scientific Research (B)
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资助金额:$4.42万
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财政年份:1991
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负责人:HASE Sumihiro
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依托单位:
Sensitive analysis of O-linked sugar chains by fluorescence labeling
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批准号:01470147
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项目类别:Grant-in-Aid for General Scientific Research (B)
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资助金额:$4.35万
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财政年份:1989
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负责人:HASE Sumihiro
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依托单位:
Structure analysis of glycoproteins by two dimensional HPLC mapping with high sensitivity
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批准号:62580120
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项目类别:Grant-in-Aid for General Scientific Research (C)
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资助金额:$0.96万
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财政年份:1987
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负责人:HASE Sumihiro
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依托单位: