Molecular-level description of proteins from Saccharomyces cerevisiae using quadrupole FT hybrid mass spectrometry for top down proteomics

被引:59
作者
Meng, FY [1 ]
Du, Y [1 ]
Miller, LM [1 ]
Patrie, SM [1 ]
Robinson, DE [1 ]
Kelleher, NL [1 ]
机构
[1] Univ Illinois, Dept Chem, Urbana, IL 61801 USA
关键词
D O I
10.1021/ac0354903
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
For improved detection of diverse posttranslational modifications (PTMs), direct fragmentation of protein ions by top down mass spectrometry holds promise but has yet to 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, 18 duplicate genes, and 44 proteolytic fragments. The platform for this study combined continuous-elution gel electrophoresis, reversed-phase liquid chromatography, automated nanospray coupled with a quadrupole-FT hybrid mass spectrometer, and a new search engine for querying a custom database. The proteins identified required no manual validation, ranged from 5 to 39 kDa, had codon biases from 0.93 to 0.083, and were primarily associated with glycolysis and protein synthesis. Illustrations of gene-specific identifications, PTM detection and subsequent PTM localization (using either electron capture dissociation or known PTM data stored in a database) show how larger scale proteome projects incorporating top down may proceed in the future using commercial Q-FT instruments.
引用
收藏
页码:2852 / 2858
页数:7
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