antioxidants Review Biosynthesis, Quantification and Genetic Diseases of the Smallest Signaling Thiol Metabolite: Hydrogen Sulfide Joanna Myszkowska 1, Ilia Derevenkov 2 , Sergei V. Makarov 2, Ute Spiekerkoetter 3 and Luciana Hannibal 1,* 1 Laboratory of Clinical Biochemistry and Metabolism, Department of General Pediatrics, Adolescent Medicine and Neonatology, Medical Center, Faculty of Medicine, University of Freiburg, 79106 Freiburg, Germany; [email protected] 2 Department of Food Chemistry, Ivanovo State University of Chemistry and Technology, 153000 Ivanovo, Russia; [email protected] (I.D.); [email protected] (S.V.M.) 3 Department of General Pediatrics, Adolescent Medicine and Neonatology, Medical Center, Faculty of Medicine, University of Freiburg, 79106 Freiburg, Germany; [email protected] * Correspondence: [email protected] Abstract: Hydrogen sulfide (H2S) is a gasotransmitter and the smallest signaling thiol metabolite with important roles in human health. The turnover of H2S in humans is mainly governed by enzymes of sulfur amino acid metabolism and also by the microbiome. As is the case with other small signaling molecules, disease-promoting effects of H2S largely depend on its concentration and compartmentalization. Genetic defects that impair the biogenesis and catabolism of H2S have been described; however, a gap in knowledge remains concerning physiological steady-state concentra- tions of H2S and their direct clinical implications. The small size and considerable reactivity of H2S Citation: Myszkowska, J.; renders its quantification in biological samples an experimental challenge. A compilation of methods Derevenkov, I.; Makarov, S.V.; currently employed to quantify H2S in biological specimens is provided in this review. Substan- Spiekerkoetter, U.; Hannibal, L. tial discrepancy exists in the concentrations of H2S determined by different techniques. Available Biosynthesis, Quantification and methodologies permit end-point measurement of H2S concentration, yet no definitive protocol exists Genetic Diseases of the Smallest for the continuous, real-time measurement of H S produced by its enzymatic sources. We present a Signaling Thiol Metabolite: 2 Hydrogen Sulfide. Antioxidants 2021, summary of available animal models, monogenic diseases that impair H2S metabolism in humans 10, 1065. https://doi.org/10.3390/ including structure-function relationships of pathogenic mutations, and discuss possible approaches antiox10071065 to overcome current limitations of study. Academic Editors: João Vicente and Keywords: hydrogen sulfide; transsulfuration; sulfur metabolite; genetic disease; cystathionine Alessandro Giuffré beta-synthase; metabolism Received: 19 May 2021 Accepted: 24 June 2021 Published: 1 July 2021 1. Introduction 1.1. Biological Chemistry of H2S Publisher’s Note: MDPI stays neutral Hydrogen sulfide (H2S) is the smallest biological thiol. In the biosphere, H2S is both with regard to jurisdictional claims in an environmental toxin and a signaling metabolite with increasingly investigated roles published maps and institutional affil- iations. in human physiology. An excellent review about the timeline of H2S investigation from its discovery by Swedish-German chemist Carl Wilhelm Scheele until present is available from Csaba Szabo [1]. This review focuses on the biosynthesis and quantification of H2S and on possible disturbances caused by genetic defects of metabolism that impair its turnover in humans. Thiols are nucleophiles of generally good reducing capacity that can Copyright: © 2021 by the authors. engage in one- and two-electron transfer reactions [2]. Reaction mechanisms involving Licensee MDPI, Basel, Switzerland. the transfer of one electron produce the corresponding reactive thiyl radical species. Two- This article is an open access article electron oxidation of thiols yields disulfide species. The reactivity of H S is comparable to distributed under the terms and 2 that of other low molecular weight thiols, including its undergoing reversible as well as conditions of the Creative Commons Attribution (CC BY) license (https:// irreversible modifications. Reactions of H2S with other reactive species, including oxygen- creativecommons.org/licenses/by/ and nitrogen-centered radicals have been comprehensively reviewed elsewhere [3–5]. 4.0/). Antioxidants 2021, 10, 1065. https://doi.org/10.3390/antiox10071065 https://www.mdpi.com/journal/antioxidants Antioxidants 2021, 10, x FOR PEER REVIEW 2 of 44 Antioxidants 2021, 10, 1065 2 of 41 Hydrogen sulfide is a weak acid with pKa~7.0 (Figure 1). Recent Raman studies of sodium sulfideHydrogen dissolved sulfide in hyper is a weak-concentrated acid with NaOH pKa~7.0 (aq) (Figure and CsOH1). Recent (aq) cast Raman serious studies of doubt onsodium the widely sulfide assumed dissolved existence in hyper-concentrated of S2- (aq) [6]. Under NaOH physiologi (aq) andcal CsOH pH, H (aq)2S exists cast serious 2− predominantlydoubt on as the a widelyhydrosulfide assumed anion existence (HS—). of It S has(aq) been [6 ].estimated Under physiological that approximately pH, H2S exists − 70–80%predominantly of hydrogen sulfide as a hydrosulfide exists as HS anion— in (HSplasma). It[7,8] has. beenIn the estimated cytosol (pH that 6. approximately9–7.0), − 70–80% of hydrogen sulfide exists as HS in plasma— [7,8]. In the cytosol (pH 6.9–7.0), approximately 50% of hydrogen sulfide exists as HS− . In specific organelles, the proportionapproximately of the different 50% of ionized hydrogen forms sulfide of hydrogen exists as HSsulfide. In may specific vary. organelles, For example, the proportion in of the different ionized forms of hydrogen sulfide may vary. For example, in lysosomes lysosomes (pH 4.7–5.0), H2S would be the predominant species, whereas in the more (pH 4.7–5.0), H2S would be the predominant species, whereas in the more— alkaline microen- alkaline microenvironment of the mitochondrial matrix (pH− > 7.8), HS has been estimatedvironment to represent of the 92% mitochondrial of all available matrix sulfide. (pH > 7.8), HS has been estimated to represent 92% of all available sulfide. Figure 1. Ionization properties of H2S. Hydrogen sulfide exists as an equilibrium of its gaseous and Figure 1. Ionization properties of H2S. Hydrogen sulfide exists− as an equilibrium of its gaseous and aqueous forms. Deprotonation of H S to form— HS has a pKa of 6.9–7.1. aqueous forms. Deprotonation of H2S to form2 HS has a pKa of 6.9–7.1. − In this review, the term H2S refers to the sum of H2S/HS . The signaling properties In this review, the term H2S refers to the sum of H2S/HS—. The signaling properties of H2S have often been compared to those of the gasotransmitters nitric oxide (NO) and of H2S have often been compared to those of the gasotransmitters nitric oxide (NO) and carbon monoxide (CO) [9]. Among the similarities, all three signaling molecules are gases, carbon monoxide (CO) [9]. Among the similarities, all three signaling molecules are gases, are synthesized by enzymes and possess in vivo half-lives of seconds to minutes. The are synthesized by enzymes and possess in vivo half-lives of seconds to minutes. The major difference that prevails until present time is the absence of a conserved canonical major difference that prevails until present time is the absence of a conserved canonical receptor molecule for H2S, which is in contrast to the case of NO and CO, both of which receptor molecule for H2S, which is in contrast to the case of NO and CO, both of which exert their functions via the sGC/cGMP signaling axis [9]. An excellent review on H2S exert theirsignaling functions in the via context the sGC/cGMP of other known signaling gasotransmitters axis [9]. An isexcellent available review elsewhere on H [102S –12]. signaling in the context of other known gasotransmitters is available elsewhere [10–12]. 1.2. Concentration, Signaling and Cellular Response to H2S 1.2. Concentration, Signaling and Cellular Response to H2S As a redox signaling metabolite, the effects of H2S in physiology are determined Asby a redox three majorsignaling factors, metaboli namely,te, the concentration, effects of H2 compartmentalizationS in physiology are determined and reactivity by [13]. three majorFirstly ,factors, the concentrations namely, concentration, of H2S in cells compartmentalization and extracellular compartments and reactivity are maintained [13]. in Firstly, athe narrow concentrations physiological of H range2S in cells by modulation and extracellular of its biogenesiscompartments (transsulfuration are maintained pathway in a narrowand microbiome physiological production) range by and modulation catabolism of (mitochondrial its biogenesis oxidation) (transsulfuration (Table1). The pathwayoccurrence and microbiome of mutations production) in genes and encoding catabolism enzymes (mitochondrial responsible oxidation) for the biosynthesis (Table and 1). Thecatabolism occurrence of Hof2S canmutations increase in and genes reduce encoding the concentration enzymes of Hresponsible2S outside optimalfor the homeo- biosynthesisstatic concentration.and catabolism The of exact H2S contributioncan increase of and the reduce microbiome the concentration to overall H2S of concentration H2S outsideis optimal a matter homeostatic of active investigation concentration. [14– 16The]. Itexact is noteworthy contribution that of the the reported microbiome concentrations to overall ofH2 HS 2concentrationS in biological is compartments a matter of active vary