Molecular-Level Description of Proteins from Saccharomyces cerevisiae Using Quadrupole FT Hybrid Mass Spectrometry for Top Down Proteomics

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• 作者：Fanyu Meng ; Yi Du ; Leah M. Miller ; Steven M. Patrie ; Dana E. Robinson ; and Neil L. Kelleher
• 刊名：Analytical Chemistry
• 出版年：2004
• 出版时间：May 15, 2004
• 年：2004
• 卷：76
• 期：10
• 页码：2852 - 2858
• 全文大小：178K
• 年卷期：v.76,no.10(May 15, 2004)
• ISSN：1520-6882

文摘

For improved detection of diverse posttranslational modifications (PTMs), direct fragmentation of protein ions bytop down mass spectrometry holds promise but has yetto be achieved on a large scale. Using lysate from Saccharomyces cerevisiae, 117 gene products were identified with 100% sequence coverage revealing 26 acetylations, 1 N-terminal dimethylation, 1 phosphorylation, 18duplicate genes, and 44 proteolytic fragments. The platform for this study combined continuous-elution gelelectrophoresis, reversed-phase liquid chromatography,automated nanospray coupled with a quadrupole-FThybrid mass spectrometer, and a new search engine forquerying a custom database. The proteins identifiedrequired no manual validation, ranged from 5 to 39 kDa,had codon biases from 0.93 to 0.083, and were primarilyassociated with glycolysis and protein synthesis. Illustrations of gene-specific identifications, PTM detection andsubsequent PTM localization (using either electron capture dissociation or known PTM data stored in a database)show how larger scale proteome projects incorporating topdown may proceed in the future using commercial Q-FTinstruments.