高等学校化学学报 ›› 2019, Vol. 40 ›› Issue (1): 69-75.doi: 10.7503/cjcu20180456

• 有机化学 • 上一篇    下一篇

银杏种子糖蛋白的SDS-PAGE分离及N-糖链的质谱分析

李成1, 王承健1,2, 晋万军1, 韩健利1, 杨梅芳1, 郜茜1, 黄琳娟1,2, 王仲孚1,2   

  1. 1. 西北大学生命科学学院, 2. 食品科学与工程学院, 西安710069
  • 收稿日期:2018-06-25 出版日期:2019-01-10 发布日期:2018-12-12
  • 作者简介:

    联系人简介: 王仲孚, 男, 博士, 教授, 博士生导师, 主要从事糖生物学与糖工程学方面的研究. E-mail: wangzhf@nwu.edu.cn

  • 基金资助:
    国家自然科学基金(批准号: 31670808, 31600647)、 陕西省教育厅自然科学专项项目(批准号: 16JK1782)和西北大学校内基金(批准号: 15NW18)资助.

Mass Spectrometric Analysis of N-Glycans of Glycoprotein Separated by SDS-PAGE Gel from Ginkgo Seed

LI Cheng1, WANG Chengjian1,2, JIN Wanjun1, HAN Jianli1, YANG Meifang1, GAO Xi1, HUANG Linjuan1,2, WANG Zhongfu1,2,*   

  1. 1. College of Life Sciences, 2. College of Food Sciences and Technology, Northwest University, Xi’an 710069, China
  • Received:2018-06-25 Online:2019-01-10 Published:2018-12-12
  • Contact: WANG Zhongfu
  • Supported by:
    † Supported by the National Natural Scince Foundation of China(Nos.31670808, 31600647), the Natural Science Special Fund of Shaanxi Provincial Education Department, China(No.16JK1782) and the Scientific Research Foundation of Northwest University, China(No.15NW18).

摘要:

利用十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)分离了银杏种子中的糖蛋白组分, 进一步用氨水催化释放N-糖链, 并采用电喷雾离子质谱(ESI-MS)及在线液相色谱-质谱联用(LC-UV-MS/MS2)等技术对胶条上释放的N-糖链进行了定性定量分析. 结果表明, 从银杏种子中分离得到11种糖蛋白, 共检测到11条N-糖链, 包括高甘露糖型(4.88%)和复杂型(95.12%) 2种类型, 其中分子量为21000, 36000和50000的糖蛋白所释放的核心α-1,3-岩藻糖和β-1,2-木糖修饰的N-糖链所占比率较高, 分别为68.23%, 64.37%和75.09%. 本研究对进一步研究银杏糖蛋白功能具有重要意义.

关键词: 银杏糖蛋白, 十二烷基硫酸钠聚丙烯酰胺凝胶电泳, N-糖苷酶F, 氨水催化, 电喷雾离子质谱

Abstract:

The glycoproteins in Ginkgo seeds were separated and isolated by sodium dodecyl sulfate polyacry-lamide gel electrophoresis(SDS-PAGE). The N-glycans released from glycoproteins were qualitatively and quantitatively analyzed by electrospray ionization mass spectrometry(ESI-MS) and on-line liquid chromatography-mass spectrometry(LC-UV-MS/MS2). As a result, eleven glycoproteins containing oligomannose type glycans(4.88%) and complex type glycans(95.12%) were obtained. And most of the N-glycans were modified by the core α-1, 3- and β-1, 2-xylose in the 21000, 36000 and 50000 glycoproteins, which accounted for 68.23%, 64.37% and 75.09%, respectively. This study can highlight the understanding of protein glycosylation in Ginkgo and provide foundation in the research of Ginkgo biloba seed allergens.

Key words: Ginkgo glycoprotein, Sodium dodecyl sulfate polyacrylamide gel electrophoresis, PNGase F, Aqueous ammonia, Electrospray ionization mass spectrometry

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