健康人肝组织麦胚凝集素亲和型糖蛋白表达谱分析  

Wheatgermagglutinin affinity glycoprotein expression profile of normal human liver tissue

作　　者：卢雯静[1] 刘银坤[1] 孙强玲[1] 周海军[1] 周新文[2] 郭坤[1] 陆豪杰[2] 杨芃原[2] 

机构地区：[1]复旦大学附属中山医院肝癌研究所,上海200032 [2]复旦大学化学系蛋白质组学研究中心,上海200433

出　　处：《复旦学报（医学版）》2007年第1期29-36,共8页Fudan University Journal of Medical Sciences

基　　金：国家重点基础研究发展计划(973项目)(001CB510205;2004CB520802);国家高技术研究发展计划(863项目)(2002BA711A11)

摘　　要：目的分析健康人肝组织麦胚凝集素(wheatgermagglutinin,WGA)亲和型糖蛋白表达谱。方法从30例健康人肝组织混合样本的总蛋白中用WGA凝集素亲和层析分离纯化糖蛋白,再利用双向电泳结合SYPRO Ruby荧光染色进一步分离WGA亲和型糖蛋白。目的蛋白质点经质谱鉴定后对其进行生物信息学分析及功能分类。结果初步建立WGA亲和型糖蛋白的双向电泳图谱,图谱均点数为(650±50)个,质谱鉴定并去冗余共获116个蛋白。经生物信息学分析104个蛋白具有不同程度的N糖基化位点,最后按Gene Ontology的分类原则进行功能分类。结论凝集素亲和层析结合双向电泳荧光染色、质谱鉴定是一种高通量的检测方法,WGA亲和型糖蛋白表达谱的初步构建为后续研究奠定了基础。Purpose To establish wheatgermagglutinin （WGA） affinity glycoprotein database of normal human liver tissue. Methods A proteome analysis coupled to lectin affinity chromatography was carried out with the following stages： protein sample preparation of normal human liver tissue, WGA lectin affinity chromatography, two dimensional electrophoresis（2-DE） separation, SYPRO Ruby fluorescent stain and MALDI-TOF-MS/MS identification, as well as data-dependent bioinformatics. Resuits 2-DE gel represented （650 ＋ 50） WGA affinity glycoprotein. Having identified 116 proteins by MALDI-TOF-MS/MS identification, we found 104 proteins had predicted N-glycosylated sites. After that, we sub-classified these proteins into several functional clusters which be useful in exploring these proteins＇ role in derail, according to the three principal categories provided by Gene Ontology. Conclusions Lectin affinity chromatography combined with 2-DE, fluorescent stain and MS is a high throughput technique for screening glycoprotein. The establishment of WGA affinity glycoprotein database will contribute to the subsequent research.

关 键 词：凝集素亲和层析 双向电泳 SYPRO Ruby荧光染色 质谱 生物信息学 

分 类 号：TQ936.223[化学工程]