Special issue on “Free Radical and Redox Biochemistry of Thiols”

Thiols are compounds that contain a carbon-bonded sulfhydryl (–SH) functional group, i.e. they are sulfur analogues of alcohols (R–OH). Biologically relevant thiols, or biothiols, include low molecular mass (LMM) compounds as well as protein cysteine residues. The formers greatly vary among different organisms, from the more ubiquitous tripeptide glutathione (γ-L-glutamyl L-cysteinylglycine, GSH) found in mM concentrations in most eukaryotic and many prokaryotic cells, towards compounds that are more restrictedly synthesized by particular species [1,2]. They participate in key metabolic and regulatory routes that in many cases involve the intermediacy of mixed-disulfide bond formation. In turn, protein cysteine residues serve diverse functions, such as structural roles through disulfide bond formation and metal binding, and are targets of post-translational modifications frequently involved in the regulation of protein activity or sub-cellular localization and participate in enzyme catalysis [3]. Cysteine residues are usually underrepresented in proteins from various organisms, and its abundance shows a positive correlation with the complexity of the species [4].

Both LMM and protein thiols are powerful nucleophiles, mostly when deprotonated to thiolates (RS⁻) and can be oxidized by one- and two-electron mechanisms, or react with different electrophiles. When those reactions occur at cysteine residues, they promote protein post-translational modifications that may affect protein structure/function as well as cellular distribution. As a consequence, many important functions including DNA and protein synthesis, cell differentiation, apoptotic cell death and antioxidant defenses, among others, can be regulated through thiol oxidations and other modifications.

This special issue is aimed to review the mechanisms by which oxidation of thiols (and their reactions with electrophiles) participates not only in cellular antioxidant defenses but also in redox signaling and regulation of biological processes. The chapter by Trujillo et al. [5] summarizes current knowledge on the species that are responsible for the one- or two-electron oxidation of thiols to thiyl radicals and sulfenic acids, respectively, and the mechanisms of these reactions. Available kinetic data are presented and discussed so as to predict the contribution of the different thiol-containing compounds to the overall reactivity of one- or two-electrons oxidants in selected biological compartments. Moreover, some thiol-containing proteins are highly reactive towards specific oxidants, and this is considered to provide the selectivity that redox regulation requires. Current understanding of the molecular determinants of such selectivity, which is particularly striking in the case of hydroperoxide-mediated oxidation in thiol-dependent peroxidases, is analyzed. Thiyl radicals and sulfenic acids are unstable species, and their subsequent reactions and main fates in vivo are also presented [5].

A more profound discussion on thiyl radical fates under biological conditions, taking into account published kinetic data, can be found in the review presented by Schoneich [6]. Those include thiyl radical repair by ascorbate and glutathione as well as reaction with oxygen, together with hydrogen abstraction from C–H bonds in peptides and proteins to form carbon-centered radicals that could result in protein damage [6]. Mechanisms leading to such protein damage include generation of peroxyl radicals at the ^αC position followed by additional hydrogen and electron transfer processes and fragmentation reactions as well as thiyl radical elimination promoting the electrophilic dehydroalanine formation, which can further react with nucleophilic protein residues [7,8].

Sulfenic acids are also usually unstable species, particularly when formed in LMM compounds. However, sulfenic acids in proteins can be relatively stabilized [9]. The review by Devarie-Baez et al. [10] focuses on the mechanisms of protein sulfenylation as well as the factors that govern the reactivity and stability of this reversible protein modification. The review presents examples in which oxidative cysteine residue modifications, and particularly sulfenic acid formation, have been detected and either (a) take part in enzymatic catalytic mechanisms or (b) are proposed to alter protein function, with emphasis on their role in cell signaling, metabolism and epigenetics. Currently available analytical methods that allow the detection and characterization of these species are discussed.

In another review in this issue, Flohé [11] presents an exhaustive review of the biology of the main thiol- (or selenol-)dependent peroxidases, namely peroxiredoxins and glutathione peroxidases, emphasizing their functions in metabolic regulation. The kinetic data presented (although still scarce) points towards these enzymes playing a preferential role as hydroperoxide sensors. Subsequent interactions of oxidized thiol-dependent peroxidases with particular protein thiols, leading to their indirect oxidation, are in the basis of the redox-regulation of specific cellular processes. Examples of indirect redox regulation of transcription factors/signaling proteins, initially described in yeast, are emerging in different organisms [12,13].

Thiol–disulfide exchange reactions play fundamental roles in oxidative protein folding, antioxidant systems as well as in redox signaling and regulation of protein function. They are reversible processes that occur through SN2 nucleophilic substitution mechanisms. Different oxido-reductases, such as thioredoxins, glutaredoxins and protein disulfide isomerase, catalyze thiol–disulfide exchange reactions by factors of up to 10⁶. The factors conferring specificity in enzymatic thiol/disulfide exchange reactions are reviewed by Netto et al. in this issue [14]. Protein–protein interaction during thiol–disulfide exchange reactions provides specificity to different pathways. Selected examples are particularly related to the regulation of gene expression which could provide evolutionary insights.

The direction of electron flow through cellular redox systems in general and thiol/disulfide couples in particular depends on thermodynamics. However, the rate at which electron transfer occurs is dictated by kinetics, and thiol/disulfide ratios vary according to cellular situations. In most cells, two redox systems, namely the thioredoxin and glutaredoxin systems, are in charge of kinetically transduced changes in thiol/disulfide pools to molecular targets that could impact in cellular functions. The fact that they are not in rapid equilibrium indicate that it is not possible, or correct, to refer to a “global thiol/disulfide redox status” in cells or compartments [15]. The review written by Comini [16] summarizes the main methods and tools currently available to quantify redox status of different thiol/disulfide systems in cells and cellular compartments. The main applications and potential drawbacks associated with their use are critically discussed.

Finally, the review presented by LoPachin and Gavin [17] focuses on the reactions of cysteine thiolates with electrophiles and the pathophysiological consequences that can arise from the resulting modifications of the sulfur centre. The authors utilize the principles of the Hard and Soft, Acids and Bases (HSAB) theory of Pearson together with computational and proteomic evidences to indicate that soft electrophiles have a common mechanism of reactivity by selectively forming Michael-type adducts with soft, highly reactive cysteine thiolate nucleophiles. The toxicological consequences of environmental exposure to, or endogenous generation of, these types of electrophiles, particularly unsaturated aldehydes, are thoroughly discussed. Moreover, the review presents the molecular basis for the development of rational therapeutic strategies for pathological conditions related to electrophile-mediated cysteine modifications.

Overall, this special issue of Free Radical Research brings together substantial amount of current kinetic, mechanistic, structural and functional data on the free radical, redox and electrophilic chemical biology of thiols. We firmly believe that the fine works presented herein contribute to a more profound understanding of the several roles and actions that thiol redox biochemistry plays in cell regulation, physiology and pathology.