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Coupling Strong Anion-Exchange Monolithic Capillary with MALDI-TOF MS for Sensitive Detection of Phosphopeptides in Protein Digest
Dong, Mingming2; Wu, Minghuo1; Wang, Fangjun1; Qin, Hongqiang1; Han, Guanghui1; Gong, Jing1; Wu, Ren'an1; Ye, Mingliang1; Liu, Zhen2; Zou, Hanfa1; Ye ML(叶明亮)
Source PublicationANALYTICAL CHEMISTRY
2010-04-01
ISSN10.1021/ac902907w
DOI10.1021/ac902907w
Volume82Issue:7Pages:2907-2915
Indexed BySCI
SubtypeArticle
Department18
Funding Project1809
Contribution Rank2;2
WOS HeadingsScience & Technology ; Physical Sciences
WOS SubjectChemistry, Analytical
WOS Research AreaChemistry
WOS KeywordDESORPTION/IONIZATION MASS-SPECTROMETRY ; PHOSPHOPROTEOME ANALYSIS ; PHOSPHORYLATED PEPTIDES ; SELECTIVE ENRICHMENT ; TITANIUM-DIOXIDE ; INDIVIDUAL PHOSPHOPROTEINS ; CHROMATOGRAPHY ; MATRIX ; DERIVATIZATION
AbstractProtein 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.
Language英语
URL查看原文
WOS IDWOS:000276004000044
Citation statistics
Cited Times:73[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://cas-ir.dicp.ac.cn/handle/321008/103051
Collection中国科学院大连化学物理研究所
Corresponding AuthorYe ML(叶明亮)
Affiliation1.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
GB/T 7714
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.
APA Dong, Mingming.,Wu, Minghuo.,Wang, Fangjun.,Qin, Hongqiang.,Han, Guanghui.,...&叶明亮.(2010).Coupling Strong Anion-Exchange Monolithic Capillary with MALDI-TOF MS for Sensitive Detection of Phosphopeptides in Protein Digest.ANALYTICAL CHEMISTRY,82(7),2907-2915.
MLA Dong, Mingming,et al."Coupling Strong Anion-Exchange Monolithic Capillary with MALDI-TOF MS for Sensitive Detection of Phosphopeptides in Protein Digest".ANALYTICAL CHEMISTRY 82.7(2010):2907-2915.
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