Quantitative Proteome Analysis by Solid-Phase Isotope Tagging and Mass Spectrometry

Nat Biotechnol. 2002 May;20(5):512-5. doi: 10.1038/nbt0502-512.

Abstract

The adaptation of sequences of chemical reactions to a solid-phase format has been essential to the automation, reproducibility, and efficiency of a number of biotechnological processes including peptide and oligonucleotide synthesis and sequencing. Here we describe a method for the site-specific, stable isotopic labeling of cysteinyl peptides in complex peptide mixtures through a solid-phase capture and release process, and the concomitant isolation of the labeled peptides. The recovered peptides were analyzed by microcapillary liquid chromatography and tandem mass spectrometry (microLC-MS/MS) to determine their sequences and relative quantities. The method was used to detect galactose-induced changes in protein abundance in the yeast Saccharomyces cerevisiae. A side-by-side comparison with the isotope-coded affinity tag (ICAT) method demonstrated that the solid-phase method for stable isotope tagging of peptides is comparatively simpler, more efficient, and more sensitive.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Chromatography, Liquid
  • Cysteine / chemistry
  • Gas Chromatography-Mass Spectrometry
  • Isotopes*
  • Mass Spectrometry / methods*
  • Models, Chemical
  • Peptides / chemistry
  • Proteins / analysis*
  • Proteins / chemistry*
  • Saccharomyces cerevisiae / metabolism
  • Time Factors

Substances

  • Isotopes
  • Peptides
  • Proteins
  • Cysteine