色谱

色谱 ›› 2016, Vol. 34 ›› Issue (12): 1234-1239.DOI: 10.3724/SP.J.1123.2016.08038

• 研究论文 • 上一篇    下一篇

基于凝集素的唾液糖蛋白分离、分析及应用

孙凯博, 尚志, 孙妍, 乔智, 刘莎, 樊柳荫, 曹成喜, 肖华   

  1. 微生物代谢国家重点实验室, 上海交通大学生命科学技术学院生物分离与分析实验室, 上海 200240
  • 收稿日期:2016-08-30 出版日期:2016-12-08 发布日期:2013-04-24
  • 通讯作者: 肖华,Tel:(021)34205682,E-mail:huaxiao@sjtu.edu.cn
  • 基金资助:

    国家自然科学基金项目（21275099，21305087，21475086）；“863”项目（2014AA020545）；国家青-千人计划项目（2013）.

Lectin based salivary glycoprotein separation, analysis and its application

SUN Kaibo, SHANG Zhi, SUN Yan, QIAO Zhi, LIU Sha, Fan Liuyin, CAO Chengxi, XIAO Hua   

  1. State Key Laboratory of Microbial Metabolism, Laboratory of Analytical Biochemistry and Bioseparation, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
  • Received:2016-08-30 Online:2016-12-08 Published:2013-04-24
  • Supported by:

    National Natural Science Foundation of China (Nos. 21275099, 21305087, 21475086); "863" Program (No. 2014AA020545); Recruitment Program of Global Youth Experts of China (2013).

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摘要：

唾液中的糖蛋白丰度偏低，给分离、分析带来挑战。该文采用麦胚素（WGA）和橙黄网胞盘菌凝集素（AAL）分别富集糖蛋白，考察了高丰度蛋白质去除和不同酶解方式对糖蛋白分离、分析的影响。结果显示，WGA和AAL提取的唾液糖蛋白经胶内酶解可鉴定到的糖蛋白数量显著多于溶液内酶解的结果，也优于去除高丰度蛋白质后的鉴定结果。选择WGA结合胶内酶解进一步对比分析肺癌患者与健康人唾液糖蛋白的差异，通过免标记定量分析共鉴定到139个蛋白质，其中102个蛋白质存在糖基化位点，包括14个在癌症组和正常组之间存在显著差异（p<0.05）的糖蛋白，表明该策略可用于唾液糖蛋白的有效分离、分析和癌症标志物的发现。

关键词: 标志物, 凝集素, 糖蛋白, 唾液

Abstract:

The abundance of glycoproteins in saliva is relatively low, which brings great challenge to their separation and analysis. In the study, wheat germ agglutinin (WGA) and aleuria aurantia lectin (AAL) were utilized to extract salivary glycoproteins. The effects of high abundant protein removal and two digestion methods on glycoprotein analysis were investigated. Our data demonstrated that the number of identified glycoproteins using WGA and AAL with in-gel digestion was significantly higher than that of in-solution digestion, as well as high abundant protein removal method. The developed strategy was further applied to compare the salivary glycoproteins from lung cancer patients and normal subjects. According to label free quantification results, 139 proteins were found and 102 proteins with the glycosylation sites were predicted. Among them, 14 glycoproteins showed significant difference between the cancer and control ones (p<0.05). Our study demonstrated that the developed strategy could be applied to the efficient separation and analysis of salivary glycoproteins and would contribute to the cancer biomarker discovery.

Key words: biomarker, glycoprotein, lectin, saliva

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