N-glycans released from glycoproteins using a commercial kit and comprehensively analyzed with a hypothetical database  

作　　者：Xue Sun Lei Tao Lin Yi Yilan Ouyang Naiyu Xu Duxin Li Robert J.Linhardt Zhenqing Zhang 

机构地区：[1]Jiangsu Key Laboratory of Translational Research and Therapy for Neuro-Psycho-Diseases and College of Pharmaceutical Sciences,Soochow University [2]Center for Biotechnology and Interdisciplinary Studies,Rensselaer Polytechnic Institute

出　　处：《Journal of Pharmaceutical Analysis》2017年第2期87-94,共8页药物分析学报（英文版）

基　　金：the National Natural Science Foundation of China (81473179 and 81673388);Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD, YX13200111);the funding for Jiangsu Key Laboratory of Translational Research and Therapy for Neuro-PsychoDiseases (BM2013003)

摘　　要：The glycosylation of proteins is responsible for their structural and functional roles in many cellular activities.This work describes a strategy that combines an efficient release, labeling and liquid chromatography–mass spectral analysis with the use of a comprehensive database to analyze N-glycans. The analytical method described relies on a recently commercialized kit in which quick deglycosylation is followed by rapid labeling and cleanup of labeled glycans. This greatly improves the separation, mass spectrometry（MS） analysis and fluorescence detection of N-glycans. A hypothetical database, constructed using Glyc Resoft, provides all compositional possibilities of N-glycans based on the common sugar residues found in N-glycans. In the initial version this database contains ＆gt; 8,700 N-glycans, and is compatible with MS instrument software and expandable. N-glycans from four different well-studied glycoproteins were analyzed by this strategy. The results provided much more accurate and comprehensive data than that had been previously reported. This strategy was then used to analyze the N-glycans present on the membrane glycoproteins of gastric carcinoma cells with different degrees of differentiation. Accurate and comprehensive N-glycan data from those cells was obtained efficiently and their differences were compared corresponding to their differentiation states. Thus, the novel strategy developed greatly improves accuracy, efficiency and comprehensiveness of N-glycan analysis.The glycosylation of proteins is responsible for their structural and functional roles in many cellular activities.This work describes a strategy that combines an efficient release, labeling and liquid chromatography–mass spectral analysis with the use of a comprehensive database to analyze N-glycans. The analytical method described relies on a recently commercialized kit in which quick deglycosylation is followed by rapid labeling and cleanup of labeled glycans. This greatly improves the separation, mass spectrometry（MS） analysis and fluorescence detection of N-glycans. A hypothetical database, constructed using Glyc Resoft, provides all compositional possibilities of N-glycans based on the common sugar residues found in N-glycans. In the initial version this database contains ＆gt; 8,700 N-glycans, and is compatible with MS instrument software and expandable. N-glycans from four different well-studied glycoproteins were analyzed by this strategy. The results provided much more accurate and comprehensive data than that had been previously reported. This strategy was then used to analyze the N-glycans present on the membrane glycoproteins of gastric carcinoma cells with different degrees of differentiation. Accurate and comprehensive N-glycan data from those cells was obtained efficiently and their differences were compared corresponding to their differentiation states. Thus, the novel strategy developed greatly improves accuracy, efficiency and comprehensiveness of N-glycan analysis.

关 键 词：N-glycan Hypothetical database Glycoproteins Gastric carcinoma cells Glycomics 

分 类 号：R3411[医药卫生—基础医学]