The tripeptide γ-
l-glutamyl-
l-cysteinyl-glycine, or glutathione (GSH), is the most abundant eukaryotic intracellular thiol, found in a concentration range of 1–10 mM. It is mainly localized in the cytosol (up to 90%), but it is also present in the mitochondria (10%), the endoplasmic reticulum, and the nucleus in a small percentage
[1]. The antioxidant activity of GSH is probably the best known and popular function of this molecule; in fact, thanks to the reactivity and the reducing ability of its cysteinyl residue’s sulfhydryl (-SH) group, it can quickly react with and neutralize potentially dangerous molecules, both of endogenous and exogenous origin, as reactive oxygen and nitrogen species (ROS and RNS, respectively), xenobiotics, and metals
[2].
However, today it is well known that the essential role of GSH overcomes its scavenger activity and it plays a crucial role in cell fate, regulating processes such as proliferation, differentiation, and apoptosis, and several other cellular functions, ranging from metabolism to immune and inflammatory response
[3][4][5][6]. In fact, the concentration of GSH and its oxidized form, glutathione disulfide (GSSG), mainly determines the redox state of the cell. Together with other thiol redox couples, such as Trx-SH/Trx-SS, GSH/GSSG acts as a sensor of intracellular redox changes and couples these modifications to redox-dependent biochemical processes
[7]. What are the mechanisms by which GSH/GSSG plays a role in signaling? Keeping in mind that the GSH/GSSG couple is the main determinant of H
2O
2 level, which is a well-known intracellular messenger in signal transduction (see reviews
[8][9][10][11][12][13]), it can play a direct role in signaling through glutathionylation, that is, the formation of mixed disulfides between its cysteine and the cysteine of a protein
[14]. As for the processes regulated by phosphorylation, the redox-mediated signal impacts proteins through a specific amino acid residue (i.e., cysteine). Several modifications of the thiol group can occur for structural needs, such as inter/intrachain disulfide –SS–, or different degrees of oxidative stress, such as the sulfenic –SOH, sulfinic –SO
2H, sulfonic –SO
3H, and mixed disulfide with cysteine or glutathione –SSR
[15]. A prerequisite for cysteine residue modification is its accessibility by the solvent and a peculiar reactivity imposed by near amino acids. Indeed, the pK value of the cysteine thiol group is around 8.0 and, therefore, always protonated at physiological pH. The so-called “reactive cysteine”, instead, is localized in a basic milieu that stabilizes the thiolated form –S
− and represents a very reactive group. The formation of glutathionylated proteins (GSSPs) has been known for many years. Initially considered an effect of oxidative stress and damage
[16][17], it then became clear that it occurred also under physiological conditions and, being a reversible process, could have a role in redox signaling
[18][19][20]. GSSPs can be formed by different biochemical reactions: through the formation of sulfenic acids (reaction below (1)) or the thiol–disulfide exchange that occurs between GSSG and protein thiols (2). The reaction can be reversibly catalyzed by different enzymes belonging to the family of thiol–disulfide oxidoreductases or redoxins, including glutaredoxin (Grx)
[21], where reversibility and catalysis are important requirements for the regulatory role of the process
[22][23].