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Information on EC 3.1.2.6 - hydroxyacylglutathione hydrolase and Organism(s) Arabidopsis thaliana and UniProt Accession Q9SID3
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EC Tree
3.1.2.6 hydroxyacylglutathione hydrolaseIUBMB Comments
Also hydrolyses S-acetoacetylglutathione, but more slowly.
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Word Map
- 3.1.2.6
- methylglyoxal
- gsh
- s-d-lactoylglutathione
-
glyoxalases
- d-lactate
-
glycation
-
glutathione-dependent
- dehydroascorbate
- monodehydroascorbate
- hemithioacetal
-
d-lactic
- mdhar
-
dicarbonyls
- trypanothione
- 2-oxoaldehydes
-
4.4.1.5
- lactoylglutathione
- metallo-beta-lactamase
- drug development
- biotechnology
- pharmacology
- medicine
- agriculture
The taxonomic range for the selected organisms is: Arabidopsis thaliana
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
glyoxalase ii, glyoxalase 2, gly ii, glyii, glxii, glx2-2, gloii, hydroxyacylglutathione hydrolase, cgloii, glo ii, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
acetoacetylglutathione hydrolase
Germ cell specific protein
GLO II
GLX2-2
GlxII
glyoxalase II
hydrolase, acetoacetylglutathione
hydrolase, hydroxyacylglutathione
Round spermatid protein RSP29
S-2-hydroxylacylglutathione hydrolase
additional information
acetoacetylglutathione hydrolase
-
-
-
-
Germ cell specific protein
-
-
-
-
GLO II
-
-
-
-
glyoxalase II
-
-
-
-
hydrolase, acetoacetylglutathione
-
-
-
-
hydrolase, hydroxyacylglutathione
-
-
-
-
Round spermatid protein RSP29
-
-
-
-
S-2-hydroxylacylglutathione hydrolase
-
-
-
-
additional information
-
the enzyme belongs to the metallo-beta-lactamase superfamily
additional information
enzyme belongs to the metallo-beta-lactamase superfamily
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
S-(2-hydroxyacyl)glutathione + H2O = glutathione + a 2-hydroxy carboxylate
active site and reaction mechanism
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hydrolysis of thioester
hydrolysis of thioester
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
S-(2-hydroxyacyl)glutathione hydrolase
Also hydrolyses S-acetoacetylglutathione, but more slowly.
CAS REGISTRY NUMBER
COMMENTARY
9025-90-5
-
9025-92-7
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
S-acetoacetylglutathione + H2O
acetoacetate + glutathione
hydrolysis at about half the rate of S-lactoylglutathione
-
-
?
S-acetylglutathione + H2O
acetate + glutathione
hydrolysis about 10fold lower than of S-lactoylglutathione
-
-
?
S-glycolylglutathione + H2O
? + glutathione
hydrolysis at about half the rate of S-lactoylglutathione
-
-
?
S-lactonylglutathione + H2O
? + glutathione
hydrolysis about 10fold lower than of S-lactoylglutathione
-
-
?
S-lactoylglutathione + H2O
D-lactate + glutathione
preferred substrae
-
-
?
S-mandeloylglutathione + H2O
mandelic acid + glutathione
hydrolysis about 10fold lower than of S-lactoylglutathione
-
-
?
S-methyl hydroxy(phenyl)ethanethioate + H2O
methanethiol + hydroxy(phenyl)acetic acid
-
thioester hydrolysis is promoted by an Fe(III)Zn(II) complex
-
-
?
lactoylglutathione + H2O
D-lactate + glutathione
-
S-D-lactoylglutathione
-
-
?
S-D-lactoylglutathione + H2O
D-lactate + glutathione
-
second step in the glyoxalase system
-
-
?
S-D-lactoylglutathione + H2O
D-lactate + glutathione
-
second step in the glyoxalase system, detoxification of methylglyoxal
-
-
?
S-D-lactoylglutathione + H2O
glutathione + D-lactate
-
-
?
S-D-lactoylglutathione + H2O
glutathione + D-lactate
-
-
?
S-D-lactoylglutathione + H2O
glutathione + D-lactate
-
-
?
additional information
?
-
no substrate: S-formylglutathione
-
-
?
additional information
?
-
the enzyme participates in the detoxification of cytotoxic and mutagenic 2-oxoaldehydes
-
?
additional information
?
-
-
the glyoxalase system is an ubiquitous pathway for the detoxification of highly reactive ketoaldehydes
-
-
?
additional information
?
-
-
the glyoxalase system is an ubiquitous pathway for the detoxification of highly reactive ketoaldehydes
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
S-D-lactoylglutathione + H2O
D-lactate + glutathione
-
second step in the glyoxalase system
-
-
?
S-D-lactoylglutathione + H2O
D-lactate + glutathione
-
second step in the glyoxalase system, detoxification of methylglyoxal
-
-
?
additional information
?
-
the enzyme participates in the detoxification of cytotoxic and mutagenic 2-oxoaldehydes
-
?
additional information
?
-
-
the glyoxalase system is an ubiquitous pathway for the detoxification of highly reactive ketoaldehydes
-
-
?
additional information
?
-
-
the glyoxalase system is an ubiquitous pathway for the detoxification of highly reactive ketoaldehydes
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Iron
1.04 mol per mol of enzyme, contains a predominant Fe(III)Zn(II) center and an anti-ferromagnetically coupled Fe(III)Fe(II) center
Fe2+
Fe3+
Iron
-
0.55 mol per mol of protein in minimal medium, located in the dinuclear site, metalloenzyme, enzyme binds a mixture of zinc, iron, and manganese when produced in cells grown in rich medium
Mn2+
Zn2+
additional information
Fe2+
0.8 mol per mol of protein, wild-type enzyme, 0.4 mol per mol of protein, R248W mutant enzyme
Fe2+
cytosolic isozyme, enzyme contains a zinc/iron binuclear metal center that is essential for substrate binding and catalysis, content of wild-type and mutant enzymes, overview
Fe3+
-
Arabidopsis thaliana mitochondrial and cytosolic gly II can accomodate a number of different metal centers, enzyme contains an iron-zinc binuclear metal center that os essential for activity
Fe3+
-
thioester hydrolysis is promoted by an Fe(III)Zn(II) complex. Thioester does not initially interact with the Fe(III) center, changes occur at this site over the course of the reaction. Formation of a Fe(III)-S-methyl ester moiety, which likely results from redox activity involving an iron(III) thiolate species. Thioester hydrolysis may involve initial coordination of the deprotonated alpha-hydroxy thioester to the zinc center followed by nucleophilic attack by a terminal Fe(III)-OH moiety and thiolate leaving group stabilization by the Fe(III) center
Mn2+
-
0.05 mol per mol of protein in minimal medium, metalloenzyme, enzyme binds a mixture of zinc, iron, and manganese when produced in cells grown in rich medium
Mn2+
-
no binding to Arabidopsis thaliana mitochondrial enzyme, but binding to cytosolic gly II
Zn2+
-
enzyme binds 2.1 mol of Zn(II) per monomer. The binding of Zn(II) is essential for enzyme viability and activity
Zn2+
0.4 mol per mol of protein, wild-type enzyme, 2.1 mol per mol of protein, R248W mutant enzyme
Zn2+
-
0.5 mol per mol of protein in minimal medium, located in the dinuclear site, metalloenzyme, enzyme binds a mixture of zinc, iron, and manganese when produced in cells grown in rich medium
Zn2+
cytosolic isozyme, enzyme contains a zinc/iron binuclear metal center that is essential for substrate binding and catalysis, content of wild-type and mutant enzymes, overview
Zn2+
-
glxII is a binuclear metalloenzyme with Zn2+ as the probable active site metal ion
Zn2+
-
thioester hydrolysis is promoted by an Fe(III)Zn(II) complex. Thioester hydrolysis may involve initial coordination of the deprotonated alpha-hydroxy thioester to the zinc center followed by nucleophilic attack by a terminal Fe(III)-OH moiety and thiolate leaving group stabilization by the Fe(III) center
additional information
enzyme possesses a binuclear active site, metal ion binding structure and kinetics, overview
additional information
-
enzyme possesses a binuclear active site, metal ion binding structure and kinetics, overview
additional information
-
metal binding structure, and metal content under different media conditions, overview, enzyme shows positive cooperative metal binding
additional information
-
overexpresssion of enzyme in minmal media containing either Zn, Fe, or Mn results in enzyme binding an average of 1 equivalent of metal ion, and presence of antiferomagnetically and ferromagnetically coupled dinuclear metal centers. Enzyme does not exist as a mononuclear metal ion containing enzyme, and shows positive cooperativity in metal binding
additional information
-
the cytosolic and mitochondrial glxII from Arabidopsis thaliana contain varying ratios of Zn2+, Fe2+ and Mn2+ and exhibit broad metal activation
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Fe2+
addition to the medium slightly reduces the activity in vivo
N-(9-fluorenylmethoxycarbonyl)-S-phenylmethoxycarbonylglutathione
-
N-phenylmethoxycarbonyl-S-(9-fluorenylmethoxycarbonyl)glutathione
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
-
the Fe(III)Zn(II) complex [(2-{[bis(2-pyridylmethyl)amino]methyl}-6-[{[2-hydroxyphenyl)methyl]-(2-pyridylmethyl)amino}methyl]-4-methylphenolFe(III)Zn(II)(micro-O2C2H3)2]ClO4 promotes thioester hydrolysis
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.12
S-D-lactoylglutathione
recombinant enzyme grown in medium supplemented with Mn2+, pH 7.2, 25°C
0.14
S-D-lactoylglutathione
-
recombinant cytosolic isozyme, medium with Zn added, pH 7.2, 25°C
0.151
S-D-lactoylglutathione
recombinant mutant L9A, pH 7.2, 25°C
0.21
S-D-lactoylglutathione
-
recombinant cytosolic isozyme, minimal medium, pH 7.2, 25°C
0.22
S-D-lactoylglutathione
recombinant enzyme grown in medium not supplemented with metal ions or supplemented with Fe2+, pH 7.2, 25°C
0.256
S-D-lactoylglutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.268
S-D-lactoylglutathione
recombinant mutant W57D, pH 7.2, 25°C
0.288
S-D-lactoylglutathione
recombinant mutant K65A, pH 7.2, 25°C
0.318
S-D-lactoylglutathione
recombinant mutant W57N, pH 7.2, 25°C
0.6
S-D-lactoylglutathione
-
recombinant cytosolic isozyme, medium with Fe added, pH 7.2, 25°C
0.139
S-Lactoylglutathione
-
pH 7.2, 25°C, growth on minimal medium without in presence of zinc
0.209
S-Lactoylglutathione
-
pH 7.2, 25°C, growth on minimal medium without added metal ion
0.6
S-Lactoylglutathione
-
pH 7.2, 25°C, growth on minimal medium in presence of iron
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
136
S-D-lactoylglutathione
-
recombinant cytosolic isozyme, medium with Fe added, pH 7.2, 25°C
168
S-D-lactoylglutathione
recombinant enzyme grown in medium supplemented with Mn2+, pH 7.2, 25°C
240
S-D-lactoylglutathione
-
recombinant cytosolic isozyme, medium with Zn added, pH 7.2, 25°C
359
S-D-lactoylglutathione
recombinant enzyme grown in medium supplemented with Fe2+, pH 7.2, 25°C
370
S-D-lactoylglutathione
recombinant wild-type enzyme, pH 7.2, 25°C
408
S-D-lactoylglutathione
recombinant enzyme grown in medium not supplemented with metal ions, pH 7.2, 25°C
470
S-D-lactoylglutathione
-
recombinant cytosolic isozyme, minimal medium, pH 7.2, 25°C
136
S-Lactoylglutathione
-
pH 7.2, 25°C, growth on minimal medium in presence of iron
240
S-Lactoylglutathione
-
pH 7.2, 25°C, growth on minimal medium without in presence of zinc
470
S-Lactoylglutathione
-
pH 7.2, 25°C, growth on minimal medium without added metal ion
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0023
N,S-bis-(phenylmethoxycarbonyl)glutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.0017
N-(9-fluorenylmethoxycarbonyl)-S-phenylmethoxycarbonylglutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.015
N-(9-fluorenylmethoxycarbonyl)glutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.002
N-phenylmethoxycarbonyl-S-(9-fluorenylmethoxycarbonyl)glutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.0056
N-phenylmethoxycarbonylglutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.014
S-(9-fluorenylmethoxycarbonyl)glutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.0049
S-phenylmethoxycarbonylglutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.00089
N,S-bis-(9-fluorenylmethoxycarbonyl)glutathione
recombinant wild-type enzyme, pH 7.2, 25°C
0.0021
N,S-bis-(9-fluorenylmethoxycarbonyl)glutathione
recombinant mutant W57D, pH 7.2, 25°C
0.0027
N,S-bis-(9-fluorenylmethoxycarbonyl)glutathione
recombinant mutants W57N and K65A, pH 7.2, 25°C
0.0028
N,S-bis-(9-fluorenylmethoxycarbonyl)glutathione
recombinant mutant L9A, pH 7.2, 25°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
activity of wild-type and mutants in crude extracts
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.5
-
saturation kinetic behavior for the phosphate diester hydrolysis reaction
7.2
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7 - 9
-
saturation kinetic behavior for the thioester hydrolysis reaction. At pH 9.0 ligand exchange at the zinc center via deprotonation of the alpha-hydroxy group of the thioester to give an equilibrium amount of a zinc alkoxide species
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
25
pI VALUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
differential expression of mitochondrial and cytosolic gly II
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). GLO2N_ARATH
324
0
35841
Swiss-Prot
Chloroplast (Reliability: 2)
PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C140A
site-directed mutagenesis, increased kcat, unaltered Km compared to the wild-type enzyme
D58C
site-directed mutagenesis, highly reduced kcat, unaltered Km compared to the wild-type enzyme
H172R
site-directed mutagenesis, highly reduced activity compared to the wild-type enzyme
H54N
site-directed mutagenesis, highly reduced kcat, 2fold increased Km compared to the wild-type enzyme
H59C
site-directed mutagenesis, reduced activity compared to the wild-type enzyme
K142A
site-directed mutagenesis, 50% reduced kcat, 2fold increased Km compared to the wild-type enzyme
K65A
site-directed mutagenesis, reduced sensitivity to 9-fluoromethoxycarbonyl- or phenylmethoxycarbonyl-protected glutathione derivative inhibitors
L9A
site-directed mutagenesis, reduced sensitivity to 9-fluoromethoxycarbonyl- or phenylmethoxycarbonyl-protected glutathione derivative inhibitors
N178A
site-directed mutagenesis, reduced kcat, slightly increased Km compared to the wild-type enzyme
R225A
site-directed mutagenesis, increased activity compared to the wild-type enzyme
R248W
W57D
site-directed mutagenesis, reduced sensitivity to 9-fluoromethoxycarbonyl- or phenylmethoxycarbonyl-protected glutathione derivative inhibitors
W57N
site-directed mutagenesis, reduced sensitivity to 9-fluoromethoxycarbonyl- or phenylmethoxycarbonyl-protected glutathione derivative inhibitors
R248W
mutant shows enhanced zinc content and reduced iron content compared to the wild-type enzyme
R248W
site-directed mutagenesis, about 8fold reduced kcat, about 10fold increased Km compared to the wild-type enzyme
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
overexpression of wild-type and mutants in Escherichia coli BL21(DE3)
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pharmacology
enzyme is a potential anti-cancer and/or anti-protozoal target, rational design of inhibitors
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Crowder, M.W.; Maiti, M.K.; Banovic, L.; Makaroff, C.A.
Glyoxalase II from A. thaliana requires Zn(II) for catalytic activity
FEBS Lett.
418
351-354
1997
Arabidopsis thaliana
Ridderstroem, M.; Mannervik, B.
Molecular cloning and charaterization of the thiolesterase glyoxalase II from Arabidopsis thaliana
Biochem. J.
322
449-454
1997
Arabidopsis thaliana
-
Yang, K.W.; Sobieski, D.N.; Carenbauer, A.L.; Crawford, P.A.; Makaroff, C.A.; Crowder, M.W.
Explaining the inhibition of glyoxalase II by 9-fluorenylmethoxycarbonyl-protected glutathione derivatives
Arch. Biochem. Biophys.
414
271-278
2003
Arabidopsis thaliana (O24496), Arabidopsis thaliana
Schilling, O.; Wenzel, N.; Naylor, M.; Vogel, A.; Crowder, M.; Makaroff, C.; Meyer-Klaucke, W.
Flexible metal binding of the metallo-beta-lactamase domain: glyoxalase II incorporates iron, manganese, and zinc in vivo
Biochemistry
42
11777-11786
2003
Arabidopsis thaliana (O24496), Arabidopsis thaliana
Zang, T.M.; Hollman, D.A.; Crawford, P.A.; Crowder, M.W.; Makaroff, C.A.
Arabidopsis glyoxalase II contains a zinc/iron binuclear metal center that is essential for substrate binding and catalysis
J. Biol. Chem.
276
4788-4795
2001
Arabidopsis thaliana (O24496)
Wenzel, N.F.; Carenbauer, A.L.; Pfiester, M.P.; Schilling, O.; Meyer-Klaucke, W.; Makaroff, C.A.; Crowder, M.W.
The binding of iron and zinc to glyoxalase II occurs exclusively as di-metal centers and is unique within the metallo-beta-lactamase family
J. Biol. Inorg. Chem.
9
429-438
2004
Arabidopsis thaliana
Marasinghe, G.P.K.; Sander, I.M.; Bennett, B.; Periyannan, G.; Yang, K.W.; Makaroff, C.A.; Crowder, M.W.
Structural Studies on a mitochondrial glyoxalase II
J. Biol. Chem.
280
40668-40675
2005
Arabidopsis thaliana (Q9SID3), Arabidopsis thaliana
Sukdeo, N.; Honek, J.F.
Microbial glyoxalase enzymes: metalloenzymes controlling cellular levels of methylglyoxal
Drug Metabol. Drug Interact.
23
29-50
2008
Arabidopsis thaliana, Escherichia coli
Yadav, S.K.; Singla-Pareek, S.L.; Sopory, S.K.
An overview on the role of methylglyoxal and glyoxalases in plants
Drug Metabol. Drug Interact.
23
51-68
2008
Arabidopsis thaliana, Brassica sp., Glycine max, Solanum tuberosum, Zea mays
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