Table of Contents
International Journal of Proteomics
Volume 2012, Article ID 514847, 9 pages
Research Article

Application of iTRAQ Reagents to Relatively Quantify the Reversible Redox State of Cysteine Residues

1Department of Biochemistry and Molecular Biology, University of Córdoba and IMIBIC, 14071 Córdoba, Spain
2Cardiovascular Proteomics Laboratory, National Center for Cardiovascular Research, 28026 Madrid, Spain
3Department of Biochemistry, University College Cork, Cork, Ireland

Received 12 April 2012; Accepted 30 April 2012

Academic Editor: Qiangwei Xia

Copyright © 2012 Brian McDonagh et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Cysteines are one of the most rarely used amino acids, but when conserved in proteins they often play critical roles in structure, function, or regulation. Reversible cysteine modifications allow for potential redox regulation of proteins. Traditional measurement of the relative absolute quantity of a protein between two samples is not always necessarily proportional to the activity of the protein. We propose application of iTRAQ reagents in combination with a previous thiol selection method to relatively quantify the redox state of cysteines both within and between samples in a single analysis. Our method allows for the identification of the proteins, identification of redox-sensitive cysteines within proteins, and quantification of the redox status of individual cysteine-containing peptides. As a proof of principle, we applied this technique to yeast alcohol dehydrogenase-1 exposed in vitro to H2O2 and also in vivo to the complex proteome of the Gram-negative bacterium Bacillus subtilis.