Glutathione (GSH) is an antioxidant in plants, animals, fungi, and some bacteria and archaea. Glutathione is capable of preventing damage to important cellular components caused by reactive oxygen species such as free radicals, peroxides, lipid peroxides, and heavy metals. It is a tripeptide with a gamma peptide linkage between the carboxyl group of the glutamate side chain and the amine group of cysteine, and the carboxyl group of cysteine is attached by normal peptide linkage to a glycine.
Thiol groups are reducing agents, existing at a concentration around 5mM in animal cells. Glutathione reduces disulfide bonds formed within cytoplasmic proteins to cysteines by serving as an electron donor. In the process, glutathione is converted to its oxidized form, glutathione disulfide (GSSG), also called L-(–)-glutathione (GSH and GSSG structure in the above scheme).
Once oxidized, glutathione can be reduced back by glutathione reductase, using NADPH as an electron donor. The ratio of reduced glutathione to oxidized glutathione within cells is often used as a measure of cellular oxidative stress.
Glutathione is not an essential nutrient for humans, since it can be synthesized in the body from the amino acids L-cysteine, L-glutamic acid, and glycine; it does not have to be present as a supplement in the diet. The sulfhydryl group (SH) of cysteine serves as a proton donor and is responsible for its biological activity. Cysteine is the rate-limiting factor in cellular glutathione biosynthesis, since this amino acid is relatively rare in foods.
Glutathione exists in both reduced (GSH) and oxidized (GSSG) states. In the reduced state, the thiol group of cysteine is able to donate a reducing equivalent (H++ e−) to other molecules, such as reactive oxygen species to neutralize them, or to protein cysteines to maintain their reduced forms. With donating an electron, glutathione itself becomes reactive and readily reacts with another reactive glutathione to form glutathione disulfide (GSSG). Such a reaction is probable due to the relatively high concentration of glutathione in cells (up to 7 mM in the liver).
Generally, interactions between GSH and other molecules with higher relative electrophilicity deplete GSH levels within the cell. An exception to this case involves the sensitivity of GSH to the electrophilic compound's relative concentration. In high concentrations, the organic molecule diethyl maleate fully depleted GSH levels in cells. However, in low concentrations, a minor decrease in cellular GSH levels was followed by a two-fold increase.
GSH can be regenerated from GSSG by the enzyme glutathione reductase (GSR): NADPH reduces FAD present in GSR to produce a transient FADH-anion. This anion then quickly breaks a disulfide bond (Cys58 – Cys63) and leads to Cys63's nucleophilically attacking the nearest sulfide unit in the GSSG molecule (promoted by His467), which creates a mixed disulfide bond (GS-Cys58) and a GS-anion. His467 of GSR then protonates the GS-anion to form the first GSH. Next, Cys63 nucleophilically attacks the sulfide of Cys58, releasing a GS-anion, which, in turn, picks up a solvent proton and is released from the enzyme, thereby creating the second GSH. So, for every GSSG and NADPH, two reduced GSH molecules are gained, which can again act as antioxidants scavenging reactive oxygen species in the cell.
In healthy cells and tissue, more than 90% of the total glutathione pool is in the reduced form (GSH) and less than 10% exists in the disulfide form (GSSG). An increased GSSG-to-GSH ratio is considered indicative of oxidative stress.
Glutathione participates in thiol protection and redox regulation of cellular thiol proteins under oxidative stress by protein S-glutathionylation, a redox-regulated post-translational thiol modification.
Glutathione has multiple functions:
It maintains levels of reduced glutaredoxin and glutathione peroxidase.
It is one of the major endogenous antioxidants produced by the cells, participating directly in the neutralization of free radicals and reactive oxygen compounds, as well as maintaining exogenous antioxidants such as vitamins C and E in their reduced (active) forms.
Regulation of the nitric oxide cycle is critical for life, but can be problematic if unregulated. Glutathione enhances the function of citrulline as part of the nitric oxide cycle.
It is used in metabolic and biochemical reactions such as DNA synthesis and repair, protein synthesis, prostaglandin synthesis, amino acid transport, and enzyme activation. Thus, every system in the body can be affected by the state of the glutathione system, especially the immune system, the nervous system, the gastrointestinal system, and the lungs.
It has a vital function in iron metabolism. Yeast cells depleted of GSH or containing toxic levels of GSH show an intense iron starvation-like response and impairment of the activity of extramitochondrial ISC enzymes thus inhibiting oxidative endoplasmic reticulum folding, followed by death.
It has roles in progression of the cell cycle, including cell death. GSH levels regulate redox changes to nuclear proteins necessary for the initiation of cell differentiation. Differences in GSH levels also determine the expressed mode of cell death, being either apoptosis or cell necrosis. Manageably low levels result in the systematic breakage of the cell whereas excessively low levels result in rapid cell death.
Function in animals
GSH is known as a substrate in conjugation reactions, which is catalyzed by glutathione S-transferase enzymes in cytosol, microsomes, and mitochondria. However, GSH is also capable of participating in nonenzymatic conjugation with some chemicals.
In the case of N-acetyl-p-benzoquinone imine (NAPQI), the reactive cytochrome P450-reactive metabolite formed by paracetamol (acetaminophen), which becomes toxic when GSH is depleted by an overdose of acetaminophen, glutathione is an essential antidote to overdose. Glutathione conjugates to NAPQI and helps to detoxify it. In this capacity, it protects cellular protein thiol groups, which would otherwise become covalently modified; when all GSH has been spent, NAPQI begins to react with the cellular proteins, killing the cells in the process. The preferred treatment for an overdose of this painkiller is the administration (usually in atomized form) of N-acetyl-L-cysteine (often as a preparation called Mucomyst), which is processed by cells to L-cysteine and used in the de novo synthesis of GSH.
Glutathione (GSH) participates in leukotriene synthesis and is a cofactor for the enzyme glutathione peroxidase. It is also important as a hydrophilic molecule that is added to lipophilic toxins and waste in the liver during biotransformation before they can become part of the bile. Glutathione is also needed for the detoxification of methylglyoxal, a toxin produced as a byproduct of metabolism.
This detoxification reaction is carried out by the glyoxalase system. Glyoxalase I catalyzes the conversion of methylglyoxal and reduced glutathione to S-D-lactoyl-glutathione. Glyoxalase II catalyzes the hydrolysis of S-D-lactoyl-glutathione to glutathione and D-lactic acid.
Glutathione, along with oxidized glutathione (GSSG) and S-nitrosoglutathione (GSNO), have been found to bind to the glutamate recognition site of the NMDA and AMPA receptors (via their γ-glutamyl moieties), and may be endogenous neuromodulators. At millimolar concentrations, they may also modulate the redox state of the NMDA receptor complex. Glutathione has been found to bind to and activate ionotropic receptors that are different from any other excitatory amino acid receptor, and which may constitute glutathione receptors, potentially making it a neurotransmitter. Glutathione is also able to activate the purinergic P2X7 receptor from Müller glia, inducing acute calcium transient signals and GABA release from both retinal neurons and glial cells.
Function in plants
In plants, glutathione is crucial for biotic and abiotic stress management. It is a pivotal component of the glutathione-ascorbate cycle, a system that reduces poisonous hydrogen peroxide. It is the precursor of phytochelatins, glutathione oligomers that chelate heavy metals such as cadmium. Glutathione is required for efficient defence against plant pathogens such as Pseudomonas syringae and Phytophthora brassicae. Adenylyl-sulfate reductase, an enzyme of the sulfur assimilationpathway, uses glutathione as an electron donor. Other enzymes using glutathione as a substrate are glutaredoxins. These small oxidoreductases are involved in flower development, salicylic acid, and plant defence signalling. In a recent report it is shown that seeds of Cassia occidentalis plants which contains multiple anthraquinones are capable of forming conjugates with glutathione. It was also found that Rhein have the most cytotoxic response with maximum oxidization of glutathione followed by emodin and aloe-emodin.
The content of glutathione in must, the first raw form of wine, determines the browning, or caramelizing effect, during the production of white wine by trapping the caffeoyltartaric acid quinones generated by enzymic oxidation as grape reaction product. Its concentration in wine can be determined by UPLC-MRM mass spectrometry.
Glutathione plays an important role in preventing oxidative damage to the skin. In addition to its many recognized biological functions, glutathione has also been associated with skin lightening ability. The role of glutathione as a skin whitener was discovered as a side effect of large doses of glutathione. Glutathione utilizes different mechanisms to exert its action as a skin whitening agent at various levels of melanogenesis. It inhibits melanin synthesis by means of stopping the neurotransmitter precursor L-DOPA's ability to interact with tyrosinase in the process of melanin production. Glutathione inhibits the actual production as well as agglutination of melanin by interrupting the function of L-DOPA. Another study found that glutathione inhibits melanin formation by direct inactivation of the enzyme tyrosinase by binding and chelating copper within the enzyme's active site. Glutathione's antioxidant property allows it to inhibit melanin synthesis by quenching of free radicals and peroxides that contribute to tyrosinase activation and melanin formation. Its antioxidant property also protects the skin from UV radiation and other environmental as well as internal stressors that generate free radicals that cause skin damage and hyperpigmentation. In most mammals, melanin formation consists of eumelanin (brown-black pigment) and pheomelanin ( yellow-red pigment) as either mixtures or co-polymers. Increase in glutathione level may induce the pigment cell to produce pheomelanin instead of eumelanin pigments. A research by Te-Sheng Chang found lowest levels of reduced glutathione to be associated with eumelanin type pigmentation, whereas the highest ones were associated with the pheomelanin. As a result, it is reasonable to assume that depletion of glutathione would result in eumelanin formation. Prota observed that decreased glutathione concentration led to the conversion of L-Dopaquinone to Dopachrome, increasing the formation of brown-black pigment (eumelanin).