View source for Glutathione synthetase

{{Short description|Enzyme}}
{{infobox protein
|Name=Glutathione synthetase
|caption=Structure of glutathione synthetase in [[yeast]]. Generated from 1M0W.<ref name="Gogos_2002">{{Cite journal |vauthors=Gogos A, Shapiro L |date=Dec 2002 |title=Large conformational changes in the catalytic cycle of glutathione synthase |journal=Structure |volume=10 |issue=12 |pages=1669–76 |doi=10.1016/S0969-2126(02)00906-1 |pmid=12467574 |doi-access=free}}</ref>
|image=Glutathione_synthetase_1M0W.png
|width=
|HGNCid=4624
|Symbol=GSS
|AltSymbols=
|EntrezGene=2937
|OMIM=601002
|RefSeq=NM_000178
|UniProt=P48637
|PDB=
|ECnumber=6.3.2.3
|Chromosome=20
|Arm=q
|Band=11.2
|LocusSupplementaryData=
}}
{{Infobox protein family
| Symbol = GSH_synthase
| Name = Eukaryotic glutathione synthase
| image = PDB 2hgs EBI.jpg
| width =
| caption = Human glutathione synthetase
| Pfam = PF03199
| Pfam_clan = CL0483
| InterPro = IPR004887
| SMART =
| PROSITE =
| MEROPS =
| SCOP = 2hgs
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}
{{infobox enzyme
| Name       = glutathione synthase
| EC_number  = 6.3.2.3
| CAS_number = 9023-62-5
| GO_code    = 0004363
| image      = 2hgs.jpg
| width      = 270
| caption    = glutathione synthetase dimer, Human
}}
{{Infobox protein family
| Symbol = GSH_synth_ATP
| Name = Eukaryotic glutathione synthase, ATP binding domain
| image = PDB 2hgs EBI.jpg
| width =
| caption = Human glutathione synthetase
| Pfam = PF03917
| Pfam_clan =
| InterPro = IPR005615
| SMART =
| PROSITE =
| MEROPS =
| SCOP = 1m0t
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}
{{Infobox protein family
| Symbol = GSH-S_N
| Name = Prokaryotic glutathione synthetase, N-terminal domain
| image = PDB 1gsh EBI.jpg
| width =
| caption = Structure of escherichia coli glutathione synthetase at ph 7.5
| Pfam = PF02951
| Pfam_clan =
| InterPro = IPR004215
| SMART =
| PROSITE =
| MEROPS =
| SCOP = 1glv
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}
{{Infobox protein family
| Symbol = GSH-S_ATP
| Name = Prokaryotic glutathione synthetase, ATP-grasp domain
| image = PDB 1gsh EBI.jpg
| width =
| caption = Structure of escherichia coli glutathione synthetase at ph 7.5
| Pfam = PF02955
| Pfam_clan = CL0179
| InterPro = IPR004218
| SMART =
| PROSITE =
| MEROPS =
| SCOP = 1glv
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}

'''Glutathione synthetase''' ('''GSS''') ({{EnzExplorer|6.3.2.3}}) is the second enzyme in the [[glutathione]] (GSH) biosynthesis pathway. It catalyses the condensation of [[gamma-glutamylcysteine]] and [[glycine]], to form glutathione.<ref>{{Cite journal |vauthors=Njålsson R, Norgren S |year=2005 |title=Physiological and pathological aspects of GSH metabolism. |journal=Acta Paediatr |volume=94 |issue=2 |pages=132–7 |doi=10.1111/j.1651-2227.2005.tb01878.x |pmid=15981742}}</ref> Glutathione synthetase is also a potent antioxidant. It is found in many species including bacteria, yeast, mammals, and plants.<ref name="Li_2003">{{Cite journal |vauthors=Li H, Xu H, Graham DE, White RH |date=Aug 2003 |title=Glutathione synthetase homologs encode alpha-L-glutamate ligases for methanogenic coenzyme F420 and tetrahydrosarcinapterin biosyntheses |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=100 |issue=17 |pages=9785–90 |bibcode=2003PNAS..100.9785L |doi=10.1073/pnas.1733391100 |pmc=187843 |pmid=12909715 |doi-access=free}}</ref>

In humans, defects in GSS are inherited in an [[autosomal recessive]] way and are the cause of severe [[metabolic acidosis]], [[5-oxoprolinuria]], increased rate of [[haemolysis]], and defective function of the [[central nervous system]].<ref name="Njålsson_2005" /> Deficiencies in GSS can cause a spectrum of deleterious symptoms in plants and human beings alike.<ref name="O'Neill 2005">{{Cite web |last=O'Neill |first=Marla |name-list-style=vanc |title=Glutathione Synthetase Deficiency |url=http://www.omim.org/entry/266130 |website=Online Mendelian Inheritance in Man |access-date=2016-03-02 |archive-date=2016-02-10 |archive-url=https://web.archive.org/web/20160210014326/http://omim.org/entry/266130 |url-status=live }}</ref>

In [[eukaryotes]], this is a [[protein dimer|homodimeric]] enzyme. The substrate-binding domain has a three-layer [[alpha helix|alpha]]/[[beta sheet|beta]]/alpha [[secondary structure|structure]].<ref name="Polekhina_1999" /> This enzyme utilizes and stabilizes an acylphosphate intermediate to later perform a favorable [[nucleophilic substitution|nucleophilic attack]] of [[glycine]].{{cn|date=November 2024}}

== Structure ==

Human and yeast glutathione synthetases are [[protein dimer|homodimers]], meaning they are composed of two identical [[protein subunit|subunits]] of itself [[non-covalent interactions|non-covalently]] bound to each other. On the other hand, ''E. coli'' glutathione synthetase is a [[tetramer|homotetramer]].<ref name="Li_2003" /> Nevertheless, they are part of the [[ATP-grasp]] [[protein superfamily|superfamily]], which consists of 21 enzymes that contain an ATP-grasp fold.<ref name="Banerjee_2007">{{Cite book |last=Banerjee |first=Ruma |title=Redox Biochemistry |date=2007 |publisher=Wiley |isbn=978-0-471-78624-5 |location=Hoboken, N.J. |page=16 |chapter=Antioxidant Molecules and Redox Factors |chapter-url=https://books.google.com/books?id=ynFUOEgjXYgC&q=glutathione+synthetase&pg=PA16 |name-list-style=vanc}}</ref> Each subunit interacts with each other through [[alpha helix]] and [[beta sheet]] [[hydrogen bond]]ing interactions and contains two domains. One domain facilitates the ATP-grasp mechanism<ref name="Fawaz_2011">{{Cite journal |vauthors=Fawaz MV, Topper ME, Firestine SM |date=Dec 2011 |title=The ATP-grasp enzymes |journal=Bioorganic Chemistry |volume=39 |issue=5–6 |pages=185–91 |doi=10.1016/j.bioorg.2011.08.004 |pmc=3243065 |pmid=21920581}}</ref> and the other is the catalytic [[active site]] for [[gamma-glutamylcysteine|γ-glutamylcysteine]]. The [[ATP-grasp fold]] is conserved within the ATP-grasp superfamily and is characterized by two alpha helices and beta sheets that hold onto the ATP molecule between them.<ref name="Fyfe_2010">{{Cite journal |vauthors=Fyfe PK, Alphey MS, Hunter WN |date=Apr 2010 |title=Structure of Trypanosoma brucei glutathione synthetase: domain and loop alterations in the catalytic cycle of a highly conserved enzyme |journal=Molecular and Biochemical Parasitology |volume=170 |issue=2 |pages=93–9 |doi=10.1016/j.molbiopara.2009.12.011 |pmc=2845819 |pmid=20045436}}</ref> The domain containing the active site exhibits interesting properties of specificity. In contrast to γ-glutamylcysteine synthetase, glutathione synthetase accepts a large variety of glutamyl-modified analogs of γ-glutamylcysteine, but is much more specific for cysteine-modified analogs of γ-glutamylcysteine.<ref name="Galperin_1997">{{Cite journal |vauthors=Galperin MY, Koonin EV |year=1997 |title=A diverse superfamily of enzymes with ATP-dependent carboxylate-amine/thiol ligase activity |journal=Protein Science |volume=6 |issue=12 |pages=2639–43 |doi=10.1002/pro.5560061218 |pmc=2143612 |pmid=9416615}}</ref> Crystalline structures have shown glutathione synthetase bound to GSH, ADP, two [[magnesium]] ions, and a sulfate ion.<ref name="Meister_1978">{{Cite book |last=Meister |first=A. |title=Functions of Glutathione in Liver and Kidney |date=1978 |publisher=Springer Berlin Heidelberg |isbn=978-3-642-67132-6 |editor-last=Wendel |editor-first=Albrecht |location=Berlin, Heidelberg |page=49 |chapter=Current Status of the γ-Glutamyl Cycle |editor-last2=Sies |editor-first2=Helmut |chapter-url=https://books.google.com/books?id=epf6CAAAQBAJ&q=specificity+of+glutathione+synthetase&pg=PA49 |name-list-style=vanc}}</ref> Two magnesium ions function to stabilize the acylphosphate intermediate, facilitate binding of ATP, and activate removal of phosphate group from ATP. Sulfate ion serves as a replacement for inorganic phosphate once the acylphosphate intermediate is formed inside the active site.<ref name="Polekhina_1999">{{Cite journal |vauthors=Polekhina G, Board PG, Gali RR, Rossjohn J, Parker MW |date=Jun 1999 |title=Molecular basis of glutathione synthetase deficiency and a rare gene permutation event |journal=The EMBO Journal |volume=18 |issue=12 |pages=3204–13 |doi=10.1093/emboj/18.12.3204 |pmc=1171401 |pmid=10369661}}</ref>

[[File:Key residues that interact with ADP within the active site.png|thumb|center|upright=1.4|Key residues that interact with ATP near the active site. Magnesium ions are shown in black. Generated from 2HGS.<ref name="Polekhina_1999" />]]

As of late 2007, 7 [[tertiary structure|structures]] have been solved for this class of enzymes, with [[Protein Data Bank|PDB]] accession codes {{PDB link|1GLV}}, {{PDB link|1GSA}}, {{PDB link|1GSH}}, {{PDB link|1M0T}}, {{PDB link|1M0W}}, {{PDB link|2GLT}}, and {{PDB link|2HGS}}.

== Mechanism ==
Glutathione synthase [[catalysis|catalyzes]] the [[chemical reaction]]{{cn|date=November 2024}}

:ATP + gamma-L-glutamyl-L-cysteine + glycine <math>\rightleftharpoons</math> ADP + phosphate + glutathione

The 3 [[substrate (biochemistry)|substrates]] of this enzyme are [[adenosine triphosphate|ATP]], [[gamma-L-glutamyl-L-cysteine]], and [[glycine]], whereas its 3 [[product (chemistry)|products]] are [[adenosine diphosphate|ADP]], [[phosphate]], and [[glutathione]].

This enzyme belongs to the family of [[ligase]]s, specifically those forming carbon-nitrogen bonds as acid-D-amino-acid ligases (peptide synthases).  The [[List of enzymes|systematic name]] of this enzyme class is '''gamma-L-glutamyl-L-cysteine:glycine ligase (ADP-forming)'''. Other names in common use include '''glutathione synthetase''', and '''GSH synthetase'''.  This enzyme participates in [[glutamate metabolism]] and [[glutathione metabolism]].  At least one compound, [[Phosphinate]] is known to [[enzyme inhibitor|inhibit this enzyme]].{{cn|date=November 2024}}

The biosynthetic mechanisms for [[ligase|synthetases]] use energy from [[nucleoside triphosphates]], whereas [[synthases]] do not.<ref name="Hara_1996">{{Cite journal |vauthors=Hara T, Kato H, Katsube Y, Oda J |date=Sep 1996 |title=A pseudo-michaelis quaternary complex in the reverse reaction of a ligase: structure of Escherichia coli B glutathione synthetase complexed with ADP, glutathione, and sulfate at 2.0 A resolution |journal=Biochemistry |volume=35 |issue=37 |pages=11967–74 |doi=10.1021/bi9605245 |pmid=8810901}}</ref> Glutathione synthetase stays true to this rule, in that it uses the energy generated by ATP. Initially, the [[carboxylate]] group on γ-glutamylcysteine is converted into an [[acyl]] phosphate by the transfer of an inorganic phosphate group of ATP to generate an acyl phosphate intermediate. Then the amino group of glycine participates in a nucleophilic attack, displacing the phosphate group and forming GSH.<ref name="urlsynthase and ligase">{{Cite web |date=1984 |title=Synthases and Ligases |url=http://www.chem.qmul.ac.uk/iubmb/newsletter/misc/synthase.html |url-status=dead |archive-url=https://web.archive.org/web/20121015094308/http://www.chem.qmul.ac.uk/iubmb/newsletter/misc/synthase.html |archive-date=2012-10-15 |access-date=2016-03-02 |website=IUPAC-IUB Joint Commission on Biochemical Nomenclature (JCBN), and Nomenclature Commission of IUB (NC-IUB), Newsletter}}</ref> After the final GSH product is made, it can be used by [[glutathione peroxidase]] to neutralize [[reactive oxygen species]] (ROS) such as H<sub>2</sub>O<sub>2</sub> or [[Glutathione S-transferase]]s in the detoxification of [[xenobiotics]].<ref name="Banerjee_2007" />
[[File:Overall synthesis rxn of GSH.jpg|thumb|center|upright=2.4|Reaction mechanism for GSH biosynthesis.<ref name="Herrera_2007">{{Cite journal |vauthors=Herrera K, Cahoon RE, Kumaran S, Jez J |date=Jun 2007 |title=Reaction mechanism of glutathione synthetase from Arabidopsis thaliana: site-directed mutagenesis of active site residues |journal=The Journal of Biological Chemistry |volume=282 |issue=23 |pages=17157–65 |doi=10.1074/jbc.M700804200 |pmid=17452339 |doi-access=free}}</ref> [[Glutamate]] and [[cysteine]] side chains are shown in red and green, respectively.]]

== Function ==

Glutathione synthetase is important for a variety of biological functions in multiple organisms. In ''[[Arabidopsis thaliana]]'', low levels of glutathione synthetase have resulted in increased vulnerability to stressors such as [[toxic heavy metal|heavy metals]], toxic organic chemicals, and [[oxidative stress]].<ref name="Moyer_2010">{{Cite journal |vauthors=Moyer AM, Sun Z, Batzler AJ, Li L, Schaid DJ, Yang P, Weinshilboum RM |date=Mar 2010 |title=Glutathione pathway genetic polymorphisms and lung cancer survival after platinum-based chemotherapy |journal=Cancer Epidemiology, Biomarkers & Prevention |volume=19 |issue=3 |pages=811–21 |doi=10.1158/1055-9965.EPI-09-0871 |pmc=2837367 |pmid=20200426}}</ref> The presence of a [[thiol]] functional group allows its product GSH to serve both as an effective [[oxidizing agent|oxidizing]] and [[reducing agent]] in numerous biological scenarios. Thiols can easily accept a pair of electrons and become [[redox|oxidized]] to [[disulfides]], and the disulfides can be readily [[redox|reduced]] to regenerate thiols. Additionally, the thiol side chain of cysteines serve as potent [[nucleophiles]] and react with oxidants and electrophilic species that would otherwise cause damage to the cell.<ref name="Xiang_2001">{{Cite journal |vauthors=Xiang C, Werner BL, Christensen EM, Oliver DJ |date=Jun 2001 |title=The biological functions of glutathione revisited in arabidopsis transgenic plants with altered glutathione levels |journal=Plant Physiology |volume=126 |issue=2 |pages=564–74 |doi=10.1104/pp.126.2.564 |pmc=111149 |pmid=11402187}}</ref> Interactions with certain metals also stabilize thiolate intermediates.<ref name="Conte_2013">{{Cite book |last=Conte |first=Mauro Lo |title=Oxidative Stress and Redox Regulation |last2=Carroll |first2=Kate S. |date=14 February 2013 |isbn=978-94-007-5786-8 |pages=1–42 |chapter=The Chemistry of Thiol Oxidation and Detection |doi=10.1007/978-94-007-5787-5_1 |access-date=2 March 2016 |chapter-url=http://www.scripps.edu/carroll/papers/loconte_bookchapter2012.pdf |archive-url=https://web.archive.org/web/20151123160434/http://www.scripps.edu/carroll/papers/loconte_bookchapter2012.pdf |archive-date=23 November 2015 |url-status=dead |name-list-style=vanc}}</ref>

In humans, glutathione synthetase functions in a similar manner. Its product GSH participates in cellular pathways involved in homeostasis and cellular maintenance. For instance, [[glutathione peroxidases]] catalyze the oxidation of GSH to [[glutathione disulfide]] (GSSG) by reducing free radicals and reactive oxygen species such as hydrogen peroxide.<ref name="Suzuki_2002">{{Cite journal |vauthors=Suzuki N, Higuchi T, Nagano T |date=Aug 2002 |title=Multiple active intermediates in oxidation reaction catalyzed by synthetic heme-thiolate complex relevant to cytochrome p450 |journal=Journal of the American Chemical Society |volume=124 |issue=32 |pages=9622–8 |doi=10.1021/ja0115013 |pmid=12167058}}</ref> [[Glutathione S-transferase]] uses GSH to clean up various metabolites, xenobiotics, and electrophiles to [[mercapturates]] for excretion.<ref name="Fang_2002">{{Cite journal |vauthors=Fang YZ, Yang S, Wu G |date=Oct 2002 |title=Free radicals, antioxidants, and nutrition |journal=Nutrition |volume=18 |issue=10 |pages=872–9 |doi=10.1016/S0899-9007(02)00916-4 |pmid=12361782}}</ref> Because of its antioxidant role, GSS mostly produce GSH inside the cytoplasm of liver cells and imported to mitochondria where detoxification occurs.<ref name="Ribas 2014">{{Cite journal |vauthors=Ribas V, García-Ruiz C, Fernández-Checa JC |date=Jul 2014 |title=Glutathione and mitochondria |journal=Frontiers in Pharmacology |volume=5 |pages=151 |doi=10.3389/fphar.2014.00151 |pmc=4079069 |pmid=25024695 |doi-access=free}}</ref> GSH is also essential for the activation of the immune system to generate robust defense mechanisms against invading pathogens.<ref name="Fang_2002" /> GSH is capable of preventing infection from the influenza virus.<ref name="Townsend_2003">{{Cite journal |vauthors=Townsend DM, Tew KD, Tapiero H |year=2003 |title=The importance of glutathione in human disease |journal=Biomedicine & Pharmacotherapy |volume=57 |issue=3–4 |pages=145–55 |doi=10.1016/S0753-3322(03)00043-X |pmc=6522248 |pmid=12818476}}</ref><ref name="Cai_2003">{{Cite journal |vauthors=Cai J, Chen Y, Seth S, Furukawa S, Compans RW, Jones DP |date=Apr 2003 |title=Inhibition of influenza infection by glutathione |journal=Free Radical Biology & Medicine |volume=34 |issue=7 |pages=928–36 |doi=10.1016/S0891-5849(03)00023-6 |pmid=12654482}}</ref>

[[File:Gsh mitochondria.jpg|thumb|center|upright=1.4|[[Glutathione]] (GSH) is synthesized in the cytoplasm of liver cells and imported to the [[mitochondria]] where it functions as a [[Cofactor (biochemistry)|cofactor]] for a number of [[antioxidant]] and detoxifying enzymes.<ref name="Ribas 2014" />]]

== Clinical significance ==

Patients with mutations in the ''GSS'' gene develop [[glutathione synthetase deficiency|glutathione synthetase (GSS) deficiency]], an autosomal recessive disorder.<ref name="Ristoff_2001">{{Cite journal |vauthors=Ristoff E, Mayatepek E, Larsson A |date=Jul 2001 |title=Long-term clinical outcome in patients with glutathione synthetase deficiency |journal=The Journal of Pediatrics |volume=139 |issue=1 |pages=79–84 |doi=10.1067/mpd.2001.114480 |pmid=11445798}}</ref> Patients develop a wide range of symptoms depending on the severity of the mutations. Mildly affected patients experience a compensated haemolytic anaemia because mutations affect stability of the enzyme. Moderately and severely affected individuals have enzymes with dysfunctional catalytic sites, rendering it unable to participate in detoxification reactions. Physiological symptoms include [[metabolic acidosis]], neurological defects, and increased susceptibility to pathogenic infections.<ref name="Njålsson_2005">{{Cite journal |vauthors=Njålsson R |date=Sep 2005 |title=Glutathione synthetase deficiency |journal=Cellular and Molecular Life Sciences |volume=62 |issue=17 |pages=1938–45 |doi=10.1007/s00018-005-5163-7 |pmc=11139123 |pmid=15990954 |s2cid=59244}}</ref>

Treatment of individuals with [[glutathione synthetase deficiency]] generally involve therapeutic treatments to address mild to severe symptoms and conditions. In order to treat [[metabolic acidosis]], severely affected patients are given large amounts of [[bicarbonate]] and [[antioxidants]] such as [[vitamin E]] and [[vitamin C]].<ref name="Kraut_2010">{{Cite journal |vauthors=Kraut JA, Madias NE |date=May 2010 |title=Metabolic acidosis: pathophysiology, diagnosis and management |journal=Nature Reviews. Nephrology |volume=6 |issue=5 |pages=274–85 |doi=10.1038/nrneph.2010.33 |pmid=20308999 |s2cid=205512465}}</ref> In mild cases, [[ascorbate]] and [[acetylcysteine|''N''-acetylcysteine]] have been shown to increase [[glutathione]] levels and increase [[erythrocyte]] production.<ref name="Jain_1994">{{Cite journal |vauthors=Jain A, Buist NR, Kennaway NG, Powell BR, Auld PA, Mårtensson J |date=Feb 1994 |title=Effect of ascorbate or N-acetylcysteine treatment in a patient with hereditary glutathione synthetase deficiency |journal=The Journal of Pediatrics |volume=124 |issue=2 |pages=229–33 |doi=10.1016/S0022-3476(94)70309-4 |pmid=8301428}}</ref> It is important to note that because glutathione synthetase deficiency is so rare, it is poorly understood. The disease also appears on a spectrum, so it is even more difficult to generalize among the few cases that occur.<ref name="Ristoff 2007">{{Cite journal |vauthors=Ristoff E, Larsson A |year=2007 |title=Inborn errors in the metabolism of glutathione |journal=Orphanet Journal of Rare Diseases |volume=2 |pages=16 |doi=10.1186/1750-1172-2-16 |pmc=1852094 |pmid=17397529 |doi-access=free}}</ref>

== See also ==
* [[Glutathione synthetase deficiency]]
* [[Glutathione]]
* [[Liver]]

== References ==
{{Reflist|33em}}
* {{Cite journal |vauthors=Law MY, Halliwell B |date=1986 |title=Purification and properties of glutathione synthetase from (Spinacia oleracea) leaves |journal=Plant Sci. |volume=43 |issue=3 |pages=185–191 |doi=10.1016/0168-9452(86)90016-6}}
* {{Cite journal |vauthors=Macnicol PK |date=1987 |title=Homoglutathione and glutathione synthetases of legume seedlings - partial-purification and substrate-specificity |journal=Plant Sci. |volume=53 |issue=3 |pages=229–235 |bibcode=1987PlnSc..53..229M |doi=10.1016/0168-9452(87)90159-2}}

== External links ==
* {{MeshName|Glutathione+Synthetase}}

{{Amino acid metabolism enzymes}}
{{Ligases CO CS and CN}}
{{Enzymes}}
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[[Category:EC 6.3.2]]
[[Category:Enzymes of known structure]]