Bioorthogonal Chemical Reporters for Analyzing Protein Sulfenylation in Cells

  1. Thu H. Truong1,
  2. Kate S. Carroll2

Published Online: 1 JUN 2012

DOI: 10.1002/9780470559277.ch110219

Current Protocols in Chemical Biology

Current Protocols in Chemical Biology

How to Cite

Truong, T. H. and Carroll, K. S. 2012. Bioorthogonal Chemical Reporters for Analyzing Protein Sulfenylation in Cells. Current Protocols in Chemical Biology. 4:101–122.

Author Information

  1. 1

    Department of Chemistry, University of Michigan, Ann Arbor, Michigan

  2. 2

    Department of Chemistry, The Scripps Research Institute, Jupiter, Florida

Publication History

  1. Published Online: 1 JUN 2012
  2. Published Print: JUN 2012


Protein sulfenylation (RSOH), the redox-based modification of cysteine thiol side chains by hydrogen peroxide (H2O2), is an important mechanism in signal transduction. Likewise, dysregulated protein sulfenylation contributes to a range of human pathologies, including cancer. Efforts to elucidate the diverse roles of protein sulfenylation in physiology and disease have been hampered by the lack of techniques to probe these modifications in native environments. To address this problem, selective chemical reporters have been developed for the detection and identification of sulfenylated proteins directly in cells. In the approach described here, a cyclic β-diketone warhead is functionalized with an azide or alkyne chemical handle. An orthogonally functionalized biotin or fluorescent reporter is then appended to the probe post-homogenization via click chemistry for downstream analysis. These bi-functional probes are exquisitely selective for protein sulfenyl modifications, non-toxic, and do not perturb intracellular redox balance. These reagents have been utilized to investigate sulfenylation in vitro and to identify intracellular protein targets of H2O2 during cell signaling. These methods provide a facile way to detect protein sulfenic acids and to study the biological role of cysteine oxidation with regard to physiological and pathological events. Curr. Protoc. Chem. Biol. 4:101-122 © 2012 by John Wiley & Sons, Inc.


  • thiol modification;
  • protein sulfenylation;
  • redox signaling;
  • click chemistry;
  • in-gel fluorescence;
  • western blotting