Measurement of Protein Persulfidation: Improved Tag-Switch Method.

Abstract

Hydrogen sulfide (H2S) is an endogenously produced signaling gasotransmitter, generated bythe enzymes cystathionine γ-lyase, cystathionine β-synthase, and 3-mercaptopyruvate sulfurtransferase. The involvement of H2S in numerous physiological, as well as pathophysiological conditions, was established over the past decade. However, the exact mechanism(s) of regulation of the biological functions by H2S are under active investigations. It is proposed that the oxidative posttranslational modification of protein cysteine residues, known as persulfidation, could be the main mechanism of action of H2S. Protein persulfides show similar reactivity to thiols, which represents one of the main obstacles in the development of a reliable method for detection of this specific protein modification. Subsequently, having a selective method for persulfide detection is of utmost importance in order to fully understand the physiological and pathophysiological role of H2S. Several methods have been proposed for thedetection of proteinpersulfidation, all of which are highlighted in this chapter. Furthermore, we provide a detailed description and protocol for the first selective persulfide labeling method, a tag-switch method, developed in our group.