De novo analysis of protein N-terminal sequence utilizing MALDI signal enhancing derivatization with Br signature

Jong Seo Kim, Jin Su Song, Yongju Kim, Seung Bum Park, Hie Joon Kim

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

De novo analysis of protein N-terminal sequence is important for identification of N-terminal proteolytic processing such as N-terminal methionine or signal peptide removal, or for the genome annotation of uncharacterized proteins. We introduce a de novo sequencing method of protein N terminus utilizing matrix-assisted laser esorption/ionization (MALDI) signal enhancing picolinamidination with bromine isotopic tag incorporated to the N terminus. The doublet signature of bromine in the tandem mass (MS/MS) spectrum distinguished N-terminal ion series from C-terminal ion series, facilitating de novo N-terminal sequencing of protein. The dual advantage of MALDI signal enhancement by the basic picolinamidine and b-ion selection aided by Br signature is demonstrated using a variety of peptides. The N-terminal sequences of myoglobin and hemoglobin as model proteins were determined by incorporating the Br tag to the N terminus of the proteins and obtaining a series of b-ions with Br signature by MS/MS analysis after chymotryptic digestion of the tagged proteins. The N-terminal peptide was selected for MS/MS analysis from the chymotryptic digest based on the Br signature in the mass spectrum. Identification of phosphorylation site as well as N- terminal sequencing of a phosphopeptide was straightforward.

Original languageEnglish
Pages (from-to)1911-1919
Number of pages9
JournalAnalytical and Bioanalytical Chemistry
Volume402
Issue number5
DOIs
Publication statusPublished - 2012 Feb 1
Externally publishedYes

Keywords

  • Amidination
  • Bromine signature
  • De novo sequencing
  • MALDI signal enhancement
  • N-terminal

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry

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