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Anal Chem. 2007 Nov 1;79(21):7984-91. Epub 2007 Oct 5.

Top-down proteomics on a chromatographic time scale using linear ion trap fourier transform hybrid mass spectrometers.

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1
Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.

Abstract

Proteomics has grown significantly with the aid of new technologies that consistently are becoming more streamlined. While processing of proteins from a whole cell lysate is typically done in a bottom-up fashion utilizing MS/MS of peptides from enzymatically digested proteins, top-down proteomics is becoming a viable alternative that until recently has been limited largely to offline analysis by tandem mass spectrometry. Here we describe a method for high-resolution tandem mass spectrometery of intact proteins on a chromatographic time scale. In a single liquid chromatography-tandem mass spectrometry (LC-MS/MS) run, we have identified 22 yeast proteins with molecular weights from 14 to 35 kDa. Using anion exchange chromatography to fractionate a whole cell lysate before online LC-MS/MS, we have detected 231 metabolically labeled (14N/15N) protein pairs from Saccharomyces cerevisiae. Thirty-nine additional proteins were identified and characterized from LC-MS/MS of selected anion exchange fractions. Automated localization of multiple acetylations on Histone H4 was also accomplished on an LC time scale from a complex protein mixture. To our knowledge, this is the first demonstration of top-down proteomics (i.e., many identifications) on linear ion trap Fourier transform (LTQ FT) systems using high-resolution MS/MS data obtained on a chromatographic time scale.

PMID:
17915963
PMCID:
PMC2361135
DOI:
10.1021/ac070553t
[Indexed for MEDLINE]
Free PMC Article
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