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题名: Coupling Strong Anion-Exchange Monolithic Capillary with MALDI-TOF MS for Sensitive Detection of Phosphopeptides in Protein Digest
作者: Dong, Mingming2;  Wu, Minghuo1;  Wang, Fangjun1;  Qin, Hongqiang1;  Han, Guanghui1;  Gong, Jing1;  Wu, Ren'an1;  Ye, Mingliang1;  Liu, Zhen2;  Zou, Hanfa1
通讯作者: 叶明亮
刊名: ANALYTICAL CHEMISTRY
发表日期: 2010-04-01
DOI: 10.1021/ac902907w
卷: 82, 期:7, 页:2907-2915
收录类别: SCI
文章类型: Article
部门归属: 18
项目归属: 1809
产权排名: 2;2
WOS标题词: Science & Technology ;  Physical Sciences
类目[WOS]: Chemistry, Analytical
研究领域[WOS]: Chemistry
英文摘要: Protein phosphorylation is one of the most biologically relevant and ubiquitous post-translational modifications. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a powerful tool for the analysis of protein phosphorylation by detection of phosphopeptides in phosphoprotein digest. Enrichment of phosphopeptides by immobilized metal ion affinity chromatography (IMAC) or metal oxide affinity chromatography (MOAC) followed with MALDI analysis is the common approach. However, the pH for loading and elution of phosphopeptides is incompatible with protein digestion as well as the preparation of the MALDI matrix solution. Therefore, some pretreatment steps, such as pH adjustment and desalting, are required, which make the approach tedious and insensitive. In this study, a strong anion-exchange (SAX) capillary monolith was prepared to enrich phosphopeptides from protein digest for MALDI-TOF MS analysis. It was found that phosphopeptides could be specifically retained on the SAX column at high pH around 8 and could be eluted by 5% formic acid. Thus, the protein digests without any pretreatment could be loaded onto the SAX column under basic pH condition; after removing nonphosphopeptides by washing, the bound phosphopeptides could be eluted directly onto a MALDI target and analyzed by MALDI-TOF MS. This approach significantly simplified the analytical procedures and reduced the sample loss. Because of the excellent MALDI MS compatible procedure and the microscale SAX column, a detection limit as low as 50 amol for the analysis of phosphopeptides from beta-casein digest was achieved. To circumvent the inconvenience of the sample loading, a new simple sample introducing method based on capillary action was proposed, which further reduced the detection limit to 10 amol.
关键词[WOS]: DESORPTION/IONIZATION MASS-SPECTROMETRY ;  PHOSPHOPROTEOME ANALYSIS ;  PHOSPHORYLATED PEPTIDES ;  SELECTIVE ENRICHMENT ;  TITANIUM-DIOXIDE ;  INDIVIDUAL PHOSPHOPROTEINS ;  CHROMATOGRAPHY ;  MATRIX ;  DERIVATIZATION
语种: 英语
原文出处: 查看原文
WOS记录号: WOS:000276004000044
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/103051
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, Natl Chromatog R&A Ctr, Key Lab Separat Sci Analyt Chem, Dalian 116023, Peoples R China
2.Nanjing Univ, Sch Chem & Chem Engn, Key Lab Analyt Chem Life Sci, Nanjing 210093, Peoples R China

Recommended Citation:
Dong, Mingming,Wu, Minghuo,Wang, Fangjun,et al. Coupling Strong Anion-Exchange Monolithic Capillary with MALDI-TOF MS for Sensitive Detection of Phosphopeptides in Protein Digest[J]. ANALYTICAL CHEMISTRY,2010,82(7):2907-2915.
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