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学科主题分析化学
Integration of click maltose hydrophilic interaction chromatography, SCX preolumn and PNGase F immobilized enzymatic reactor for N-linked glycosylation sites profiling
Qu YY(曲焱焱); Yuan HM(袁辉明); Zhang LH(张丽华); Liang Z(梁振); Zhang YK(张玉奎)
会议文集26th International Symposium on Microscale Bioseparations
会议名称26th International Symposium on Microscale Bioseparations
会议日期2011-5-1
2011
会议地点圣地亚哥
页码75-0
出版者待补充
出版地待补充
合作性质墙报
部门归属1810
主办者CASSS
英文摘要N-glycosylation is one of the most common and complex post-translational modifications of proteins, and plays an important role in many cellular interactions and pathogenesis of diseases. Global mapping of N-linked glycosylation sites is a prerequisite for fully understanding the biological functions of N-linked glycoproteins. Therefore, the development of a fast and specific strategy to achieve high-throughput and high-sensitivity glycoproteome analysis becomes imperative. The traditional N-linked glycosylation sites profiling is usually off-line performed, with drawbacks such as long analysis time, resultant sample loss and manual manipulation. In our recent work, an integrated sample pretreatment system, composed of a click maltose hydrophilic interaction chromatography (HILIC) column, a strong cation exchange (SCX) precolumn and a PNGase F immobilized enzymatic reactor (IMER), was established for the simultaneous glycopeptide enrichment, sample buffer exchange, and on-line deglycosylation. Compared with the conventional off-line method, the analysis time, including glycopeptide enrichment and deglycosylation was shortened to 1 h, and the detection limit as low as 5 fmol was achieved. Moreover, such a system was successfully applied for analyzing the digest of soluble fraction extracted from rat brain. A total of 120 unique glycoprotein groups and 196 N-linked glycosylation sites were identified, with the injected digests amount as 6 µg. All these results demonstrated that the integrated system is of great promise to achieve fast and high sensitive N-linked glycosylation sites profiling, which could be further online coupled with nano HPLC-ESI/MS/MS to achieve high-throughput glycoproteome analysis.; N-glycosylation is one of the most common and complex post-translational modifications of proteins, and plays an important role in many cellular interactions and pathogenesis of diseases. Global mapping of N-linked glycosylation sites is a prerequisite for fully understanding the biological functions of N-linked glycoproteins. Therefore, the development of a fast and specific strategy to achieve high-throughput and high-sensitivity glycoproteome analysis becomes imperative. The traditional N-linked glycosylation sites profiling is usually off-line performed, with drawbacks such as long analysis time, resultant sample loss and manual manipulation. In our recent work, an integrated sample pretreatment system, composed of a click maltose hydrophilic interaction chromatography (HILIC) column, a strong cation exchange (SCX) precolumn and a PNGase F immobilized enzymatic reactor (IMER), was established for the simultaneous glycopeptide enrichment, sample buffer exchange, and on-line deglycosylation. Compared with the conventional off-line method, the analysis time, including glycopeptide enrichment and deglycosylation was shortened to 1 h, and the detection limit as low as 5 fmol was achieved. Moreover, such a system was successfully applied for analyzing the digest of soluble fraction extracted from rat brain. A total of 120 unique glycoprotein groups and 196 N-linked glycosylation sites were identified, with the injected digests amount as 6 µg. All these results demonstrated that the integrated system is of great promise to achieve fast and high sensitive N-linked glycosylation sites profiling, which could be further online coupled with nano HPLC-ESI/MS/MS to achieve high-throughput glycoproteome analysis.
语种英语
文献类型会议论文
条目标识符http://cas-ir.dicp.ac.cn/handle/321008/116025
专题中国科学院大连化学物理研究所
通讯作者Zhang LH(张丽华)
推荐引用方式
GB/T 7714
Qu YY,Yuan HM,Zhang LH,et al. Integration of click maltose hydrophilic interaction chromatography, SCX preolumn and PNGase F immobilized enzymatic reactor for N-linked glycosylation sites profiling[C]. 待补充:待补充,2011:75-0.
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