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学科主题: 物理化学
题名: High throughput tryptic digestion via poly (acrylamide-co-methylenebisacrylamide) monolith based immobilized enzyme reactor
作者: Wu, Shuaibin1, 2;  Sun, Liangliang1, 2;  Ma, Junfeng1, 2;  Yang, Kaiguang1;  Liang, Zhen1;  Zhang, Lihua1;  Zhang, Yukui1
通讯作者: 张丽华
关键词: Immobilized enzyme reactor ;  Poly (acrylamide-co-methylenebisacrylamide) monolith ;  Protein digestion ;  Protein identification ;  Proteomics
刊名: TALANTA
发表日期: 2011-02-15
DOI: 10.1016/j.talanta.2010.12.011
卷: 83, 期:5, 页:1748-1753
收录类别: SCI
文章类型: Article
部门归属: 18
项目归属: 1810
产权排名: 1,1
WOS标题词: Science & Technology ;  Physical Sciences
类目[WOS]: Chemistry, Analytical
摘要: High throughput tryptic digestion via poly (acrylamide-co-methylenebisacrylamide) monolith based immobilized enzyme reactor
研究领域[WOS]: Chemistry
英文摘要: A poly (acrylamide-co-methylenebisacrylamide) (poly (AAm-co-MBA)) monolith was prepared by thermal polymerization in the 100 or 250 mu m i.d. capillary. The monolithic support was activated by ethylenediamine followed by glutaraldehyde. Trypsin was then introduced to form an immobilized enzyme reactor (IMER). The prepared IMER showed a reliable mechanical stability and permeability (permeability constant K = 2.65 x 10(-13) m(2)). With BSA as the model protein, efficient digestion was completed within 20 s, yielding the sequence coverage of 57%, better than that obtained from the traditional in-solution digestion (42%), which took about 12 h. Moreover. BSA down to femtomole was efficiently digested by the IMER and positively identified by matrix assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). To test the applicability of IMER for complex sample profiling, proteins extracted from Escherichia coli were digested by the IMER and further analyzed by nanoreversed phase liquid chromatography-electrospray ionization-mass spectrometry (nanoRPLC-ESI-MS/MS). In comparison to in-solution digestion, despite slightly fewer proteins were positively identified at a false discovery rate (FDR) of similar to 1% (333 vs 411), the digestion time used was largely shortened (20 s vs 24 h), implying superior digestion performance for the high throughput analysis of complex samples. (C) 2011 Elsevier B.V. All rights reserved.
关键词[WOS]: ENABLING PROTEIN DIGESTION ;  MASS-SPECTROMETRY ;  CAPILLARY ELECTROCHROMATOGRAPHY ;  PEPTIDE SEPARATION ;  TRYPSIN REACTOR ;  IDENTIFICATION ;  PROTEOMICS ;  SYSTEM ;  ONLINE
语种: 英语
WOS记录号: WOS:000287110100065
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/115315
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

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作者单位: 1.Chinese Acad Sci, Dalian Inst Chem Phys, Key Lab Separat Sci Analyt Chem, Natl Chromatog R&A Ctr, Dalian 116023, Peoples R China
2.Chinese Acad Sci, Grad Sch, Beijing 100049, Peoples R China

Recommended Citation:
Wu, Shuaibin,Sun, Liangliang,Ma, Junfeng,et al. High throughput tryptic digestion via poly (acrylamide-co-methylenebisacrylamide) monolith based immobilized enzyme reactor[J]. TALANTA,2011,83(5):1748-1753.
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