A practical guide to working with H2S at the interface of chemistry and biology

Abstract

Hydrogen sulfide (H₂S) is the most recently accepted endogenously produced gasotransmitter and is now implicated in a variety of physiological functions. In this tutorial review, our goal is to provide researchers new to the field of H₂S chemical biology with practical considerations, pitfalls, and best practices to enable smooth entry into investigations focused on biological H₂S. We present practical handling and safety considerations for working with this reactive biomolecule, and cover basic roles of H₂S biogenesis and action. Experimental methods for modulating H₂S levels, including enzymatic knockout, RNA silencing, enzymatic inhibition, and use of small molecule H₂S donors are highlighted. Complementing H₂S modulation techniques, we also highlight current strategies for H₂S detection and quantification.

Figure 1

Enzymatic H₂S production pathways. CSE: cystathionine γ-lyase; CBS: cystathionine β-synthase; CAT: cysteine aminotransferase; DAO: D-aminoacid oxidase; 3-MST: 3-mercaptopyruvate sulfurtransferase. Grey boxes indicate common substrates for H₂S-producing enzymes.

Figure 2

Selected common CSE and CBS inhibitors with associated IC₅₀ values.

Scheme 1

(a) Cold cyanolysis detection/quantification of sulfane sulfur. (b) General strategy for tag-switch labelling of protein persulfides.

Scheme 2

Common motifs for small-molecule H₂S donors. (a) Organic polysulfides, (b) Cysteine-activated H₂S donors, (c) Cysteine-activated H₂S donors with N-SH motifs, (d) Hydrolysis-based donors, and (e) Anethole 1,2-dithiole-3-thione (ADT) derived donors.

Scheme 3

(a) Methylene blue (MB) and (b) monobromobimane (mBB) methods for H₂S quantification.

Scheme 4

Common motifs in reaction-based probes for H₂S, including reactions based on (a) reduction, (b) nucleophilic attack, and (c) metal precipitation.