High-throughput global peptide proteomic analysis by combining stable isotope amino acid labeling and data-dependent multiplexed-MS/MS

Scott J. Berger, Sang Won Lee, Gordon A. Anderson, Liljana Paša-Tolić, Nikola Tolić, Yufeng Shen, Rui Zhao, Richard D. Smith

Research output: Contribution to journalArticlepeer-review

63 Citations (Scopus)

Abstract

In this work, we describe the application of a stable isotope amino acid (lysine) labeling in conjunction with data-dependent multiplexed tandem mass spectrometry (MS/MS) to facilitate the characterization and identification of peptides from proteomic (global protein) digests. Lysine auxotrophic yeast was grown in the presence of 13C-labeled or unlabeled lysine and combined after harvesting in equal proportions. Endoproteinase LysC digestion of the cytosolic fraction produced a global proteomic sample, consisting of heavy/light labeled peptide pairs. Then data-dependent multiplexed-MS/MS was applied to simultaneously select and dissociate only labeled peptide ion pairs. The approach allows differentiation between N-terminal (e.g., b-type ions) and C-terminal fragment ions (e.g., y-type ions) in resulting tandem mass spectra, as well as the capability of differentiation between nearisobaric glutamine and lysine residues. We also describe the utility of peptide composition and fragment information to support peptide identifications and examine the potential application of lysine labeling for differential quantitative protein analysis.

Original languageEnglish
Pages (from-to)4994-5000
Number of pages7
JournalAnalytical chemistry
Volume74
Issue number19
DOIs
Publication statusPublished - 2002 Oct 1

ASJC Scopus subject areas

  • Analytical Chemistry

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