Shotgun Annotation of Histone Modifications: A New Approach for Streamlined Characterization of Proteins by Top Down Mass Spectrometry

James J. Pesavento, Yong Bin Kim, Gregory K. Taylor, Neil L. Kelleher*

*Corresponding author for this work

Research output: Contribution to journalArticle

132 Scopus citations

Abstract

Eukaryotic histones serve as prototypical examples of posttranslational complexity with diverse modifications (PTMs) on many different residues that comprise a "Histone Code". To help crack this code more efficiently, we demonstrate a new strategy for protein characterization wherein complete PTM descriptions are obtained by database retrieval instead of manual interpretation of information-rich data from high-resolution tandem mass spectrometry (MS/MS). A database of nearly 50000 modified histone H4 sequences was created and queried with 91 fragment ions from electron capture dissociation of a histone form +112 Da (versus unmodified mass) selectively accumulated in a quadrupole Fourier transform hybrid mass spectrometer. The correct form atop the retrieval list indicated dimethylation at Lys20, acetylation at the N terminus, and acetylation at Lys16 (resolved from trimethylation, Δm = 0.036 Da). A statistical evaluation reveals the critical role of mass accuracy and that PTM "isomers" are retrieved as next-best matches. The applicability of shotgun annotation to forms of H4 with up to six PTMs is demonstrated, with extensibility to other histones (e.g., H2A, H2B, H3) and other protein classes projected.

Original languageEnglish (US)
Pages (from-to)3386-3387
Number of pages2
JournalJournal of the American Chemical Society
Volume126
Issue number11
DOIs
StatePublished - Mar 22 2004

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

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