一种利用DSA-FACE分析寡糖链的方法  

An Optimized Method Based on DSA-FACE Analyzing Oligosaccharides from Glycoproteins

作　　者：刘波[1] 宋淼[1,2] 巩新[1] 唱韶红[1] 王凌雪[1,3] 杨依丽[1] 汪丽娜[1,2] 马清钧[1] 吴军[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071 [2]沈阳药科大学,辽宁沈阳110016 [3]西南大学,重庆400000

出　　处：《生物技术通讯》2008年第6期885-888,共4页Letters in Biotechnology

基　　金：国家高技术研究发展计划(2007AA02Z103)

摘　　要：目的:优化糖链结构分析方法,满足高通量、高灵敏度和快速分析糖基结构的要求。方法:基于DNA测序仪的荧光糖电泳(DSA-FACE),将糖蛋白经过糖苷酶酶切获得糖链,并与荧光标记物8-氨基芘基-1,3,6-三磺酸(APTS)进行衍生化反应,标记样品经过Sephadex-G10、HPLC纯化后,用DNA3100测序仪电泳分离,用Genescan3.7软件进行数据分析。结果:利用DSA-FACE技术分析了标准品Man5GlcNAc2和Gal2GlcNAc2Man3GlcNac2,并得到糖蛋白牛核糖核酸酶B的5种N-糖基结构(Man5～9GlcNAc2)。结论:DSA-FACE是分析糖基结构的有效技术手段,能够实现对糖基结构的高通量、高灵敏度和快速分析。Objective： To observe the structures of N-glycan on glycoproteins in a high-throughput, uhrasensitive and fast way, an optimized method for the oligosaccharides sequencing based on DNA sequencer assisted fluorophore-assisted carbohydrate electrophoresis（DSA-FACE） was established. Methods： Firstly, the N-glycans from glycoproteins were derivati- zatied with 8-aminopyrene-l,3,6-trisulfonate（APTS）. Secondly, the sample was separated and purified using Sephadex-G10, HPLC. Samples electrophoresis was done by DNA sequencer 3100, and the data were analyzed by Genescan 3.7 software. Results： Standard glycans MansGleNAc2, Gal2GlcNAc2Man3GlcNAc2 were analyzed by DSA-FACE. Subsequently, the N-gly- can from glycoprotein bovine ribonuclease B was also analyzed by DSA-FACE, and the five N-glycans （Mans_gGlcNAc2） were determined by comparison to the standard glycans. Conclusion： DSA-FACE is a good method on sample quantity supplied and detection sensitivity, and it should bring high-performance, high-throughput and ultrasensitive glycosylation analysis.

关 键 词：基于DNA测序仪的荧光糖电泳 8-氨基芘基-1 3 6-三磺酸 糖链分析 糖蛋白 

分 类 号：Q503[生物学—生物化学]