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Information on EC 1.20.4.4 - arsenate reductase (thioredoxin) and Organism(s) Staphylococcus aureus and UniProt Accession P0A006
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1.20.4.4 arsenate reductase (thioredoxin)IUBMB Comments
The enzyme, characterized in bacteria of the Firmicutes phylum, is specific for thioredoxin . It has no activity with glutaredoxin [cf. EC 1.20.4.1, arsenate reductase (glutaredoxin)]. Although the arsenite formed is more toxic than arsenate, it can be extruded from some bacteria by EC 7.3.2.7, arsenite-transporting ATPase; in other organisms, arsenite can be methylated by EC 2.1.1.137, arsenite methyltransferase, in a pathway that produces non-toxic organoarsenical compounds. The enzyme also has the activity of EC 3.1.3.48, protein-tyrosine-phosphatase .
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The taxonomic range for the selected organisms is: Staphylococcus aureus
The expected taxonomic range for this enzyme is: Bacteria, Archaea
The expected taxonomic range for this enzyme is: Bacteria, Archaea
Synonyms
arsc3, sa_arsc, pi258 arsenate reductase, thioredoxin-coupled arsenate reductase, thioredoxin-arsenate reductase complex, trx-coupled arsenate reductase, bs_arsc, arsc1', 
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topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
ArsC
PATHWAY SOURCE
PATHWAYS
-
-
SYSTEMATIC NAME
IUBMB Comments
arsenate:thioredoxin oxidoreductase
The enzyme, characterized in bacteria of the Firmicutes phylum, is specific for thioredoxin [1]. It has no activity with glutaredoxin [cf. EC 1.20.4.1, arsenate reductase (glutaredoxin)]. Although the arsenite formed is more toxic than arsenate, it can be extruded from some bacteria by EC 7.3.2.7, arsenite-transporting ATPase; in other organisms, arsenite can be methylated by EC 2.1.1.137, arsenite methyltransferase, in a pathway that produces non-toxic organoarsenical compounds. The enzyme also has the activity of EC 3.1.3.48, protein-tyrosine-phosphatase [3].
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
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
-
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
-
-
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
-
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
the enzyme encoded by Staphylococcus aureus arsenic-resistance plasmid pI258 reduces intracellular arsenate to the more toxic arsenite, which is subsequently extruded from the cell
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
glutaredoxin and reduced glutathione does not stimulate arsenate reduction
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
the enzyme uses an intramolecular thiol pair (Cys82, Cys89) for the reduction of arsenate
-
-
?
additional information
?
-
assays are performed with different arsenate concentrations and arsenate reductase concentrations in the presence of 0.42 microM Escherichia coli thioredoxin, 0.14 microM Escherichia coli thioredoxin reductase and 125 microM NADPH
-
-
?
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
additional information
?
-
assays are performed with different arsenate concentrations and arsenate reductase concentrations in the presence of 0.42 microM Escherichia coli thioredoxin, 0.14 microM Escherichia coli thioredoxin reductase and 125 microM NADPH
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
-
-
-
?
arsenate + thioredoxin
arsenite + thioredoxin disulfide + H2O
the enzyme encoded by Staphylococcus aureus arsenic-resistance plasmid pI258 reduces intracellular arsenate to the more toxic arsenite, which is subsequently extruded from the cell
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
thioredoxin
interaction between thioredoxin and the enzyme pI258 arsenate reductase. Thioredoxin converts oxidized ArsC back towards its initial reduced state
additional information
ArsC couples to thioredoxin, thioredoxin reductase, and NADPH to be enzymatically active
-
additional information
purified enzyme reduces radioactive arsenate to arsenite when coupled to thioredoxin, thioredoxin reductase, and NADPH. All three protein components, arsenate reductase, thioredoxin, and thioredoxin reductase, are required for arsenate reduction. Glutaredoxin and reduced glutathione do not stimulate arsenate reduction
-
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
arsenite, tellurite, and antimonite [Sb(III)] are inhibitors for NADPH oxidation
-
additional information
the substrate, arsenate, is not inhibitory at concentrations up to 40 mM
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
mechanism by which potassium and tetrahedral oxyanions affect catalysis within the Trx-coupled family of arsenate reductases
-
additional information
-
mechanism by which potassium and tetrahedral oxyanions affect catalysis within the Trx-coupled family of arsenate reductases
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.009
arsenate
wild-type Staphylococcus aureus, condition: 150 mM KCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.022
arsenate
wild-type Staphylococcus aureus, condition: 150 mM NaCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.061
arsenate
wild-type Staphylococcus aureus, condition: 50 mM Na2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.08
arsenate
H62Q mutant Staphylococcus aureus, condition: 50 mM Na2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.081
arsenate
wild-type Staphylococcus aureus, condition: 50 mM K2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.087
arsenate
H62Q mutant Staphylococcus aureus, condition: 150 mM KCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.123
arsenate
H62Q mutant Staphylococcus aureus, condition: 150 mM NaCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.131
arsenate
H62Q mutant Staphylococcus aureus, condition: 50 mM K2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
additional information
arsenate
pH 7.5, 37°C, at low substrate concentrations the Km-value for arsenate is 0.0008 mM. Above 1 mM arsenate, a second increase in rate with increasing substrate is observed, with an apparent Km of 2 mM arsenate
additional information
additional information
NADPH oxidation shows Michaelis-Menten kinetics with a Km of 1 microM AsO43- and an apparent Vmax of 200 nmol/min per mg of protein. At high substrate concentration (above 1 mM AsO43-), a secondary rise in the reaction rate is observed, with a Km of 2 mM and an apparent Vmax of 450 nmol/min per mg of protein
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.061
arsenate
pH 7.5, 37°C, mutant enzyme C15A, at 0.002 mM arsenate
0.075
arsenate
pH 7.5, 37°C, wild-type enzyme, at 0.002 mM arsenate
0.08
arsenate
pH 7.5, 37°C, mutant enzyme C15A, at 10 mM arsenate
0.165
arsenate
pH 7.5, 37°C, wild-type enzyme, at 10 mM arsenate
0.58
arsenate
wild-type Staphylococcus aureus, condition: 150 mM NaCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.7
arsenate
H62Q mutant Staphylococcus aureus, condition: 150 mM KCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.75
arsenate
H62Q mutant Staphylococcus aureus, condition: 150 mM NaCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
0.9
arsenate
wild-type Staphylococcus aureus, condition: 150 mM KCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
1.98
arsenate
H62Q mutant Staphylococcus aureus, condition: 50 mM Na2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
2.9
arsenate
wild-type Staphylococcus aureus, condition: 50 mM Na2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
3.03
arsenate
H62Q mutant Staphylococcus aureus, condition: 50 mM K2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
3.65
arsenate
wild-type Staphylococcus aureus, condition: 50 mM K2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
6.1
arsenate
H62Q mutant Staphylococcus aureus, condition: 150 mM NaCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
8.1
arsenate
H62Q mutant Staphylococcus aureus, condition: 150 mM KCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
23.1
arsenate
H62Q mutant Staphylococcus aureus, condition: 50 mM K2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
24.8
arsenate
H62Q mutant Staphylococcus aureus, condition: 50 mM Na2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
26.4
arsenate
wild-type Staphylococcus aureus, condition: 150 mM NaCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
45.1
arsenate
wild-type Staphylococcus aureus, condition: 50 mM K2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
47.5
arsenate
wild-type Staphylococcus aureus, condition: 50 mM Na2SO4, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
100
arsenate
wild-type Staphylococcus aureus, condition: 150 mM KCl, study about the impact of potassium and the tetrahedral oxyanion sulfate on the steady-state kinetic parameters
441
arsenate
pH 7.5, 37°C, mutant enzyme C15A, at 0.002 mM arsenate
1170
arsenate
pH 7.5, 37°C, wild-type enzyme, at 0.002 mM arsenate
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
the enzyme encoded by Staphylococcus aureus arsenic-resistance plasmid pI258 reduces intracellular arsenate to the more toxic arsenite, which is subsequently extruded from the cell
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). MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
14400
14436
x * 14436, protein with a loss of the first three amino acid residues from part of the arsenate reductase may have occurred intracellularly or extracellularly during the purification process, mass spectral analysis
14500
electrospray mass spectrometry shows two molecular masses of 14810.5 and 14436.0 Da, suggesting that 70% of the purified protein lacks the N-terminal three amino acids
14810
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
?
x * 14436, protein with a loss of the first three amino acid residues from part of the arsenate reductase may have occurred intracellularly or extracellularly during the purification process, mass spectral analysis
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C10A
C10S/C15A
C10S/C15A/C82S
C10S/C15A/C89L
mutant, determination of the redox potential of the Cys82-Cys89 redox couple
C15A
C82A
C82S
C89A
additional information
C10A
site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes
C10S/C15A/C82S
mutant, determination of the redox potential of the Cys82-Cys89 redox couple, thioredoxin is unable to reduce the Cys10-Cys15 disulfide in oxidized ArsC C82S
C10S/C15A/C82S
site-directed mutagenesis, triple mutant, structure very similar to that of the reduced form of wild-type ArsC
C15A
site-directed mutagenesis, only ArsC wild type and ArsC C15A show enzymatic activity
C15A
as compared to wild-type enzyme the affinity is reduced ba a factor of 2
C82A
site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes
C82S
site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes
C89A
site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes
additional information
ArsC triple mutants and the Trx C32S mutant formate a ArsC-TrxC32S complex
additional information
-
ArsC triple mutants and the Trx C32S mutant formate a ArsC-TrxC32S complex
additional information
commonly occurring mutation of a histidine (H62), located about 6 A from the potassium-binding site in Sa_ArsC, to a glutamine uncouples the kinetic dependency on potassium. Mutations within the Trx-coupled family of arsenate reductases lead to subtly different ion-dependent kinetic features
additional information
-
commonly occurring mutation of a histidine (H62), located about 6 A from the potassium-binding site in Sa_ArsC, to a glutamine uncouples the kinetic dependency on potassium. Mutations within the Trx-coupled family of arsenate reductases lead to subtly different ion-dependent kinetic features
additional information
essential cysteinyl residues and redox couple in arsenate reductase are identified by a combination of site-specific mutagenesis and endoprotease-digest mass spectroscopy analysis
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
C10S/C15A/C89L mutant and C10S/C15A/C82S mutant purified by Ni2+-NTA Superflow
wild-type ArsC and the Cys mutants, gel filtration more than 97% pure
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
C10S/C15A/C89L mutant and C10S/C15A/C82S mutant expressed in Escherichia coli
expressed in Escherichia coli
expression in Escherichia coli. Wild-type enzyme and the Cys mutants (C15A, C10A, C82A, C82S, C89A, C10SC15S, C10SC15A) are expressed in Escherichia coli. Wild-type enzyme, mutant enzyme C15A, mutant enzyme C10A, mutant enzyme C82S, mutant enzyme C89A, and mutant enzyme C10SC15A are expressed soluble and with high yields. Mutant enzyme C82A is found in inclusion bodies, and the double mutant C10S/C15S is not expressed
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Ji, G.; Garber, E.A.E.; Armes, L.G.; Chen, C.M.; Fuchs, J.A.; Silver, S.
Arsenate reductase of Staphylococcus aureus plasmid pI258
Biochemistry
33
7294-7299
1994
Staphylococcus aureus (P0A006)
Messens, J.; Hayburn, G.; Brosens, E.; Laus, G.; Wyns, L.
Development of a downstream process for the isolation of Staphylococcus aureus arsenate reductase overproduced in Escherichia coli
J. Chromatogr. B
737
167-178
2000
Staphylococcus aureus (P0A006)
Messens, J.; Hayburn, G.; Desmyter, A.; Laus, G.; Wyns, L.
The essential catalytic redox couple in arsenate reductase from Sataphylococcus aureus
Biochemistry
38
16857-16865
1999
Staphylococcus aureus (P0A006)
Messens, J.; Van Molle, I.; Vanhaesebrouck, P.; Van Belle, K.; Wahni, K.; Martins, J.C.; Wyns, L.; Loris, R.
The structure of a triple mutant of pI258 arsenate reductase from Staphylococcus aureus and its 5-thio-2-nitrobenzoic acid adduct
Acta Crystallogr. Sect. D
60
1180-1184
2004
Staphylococcus aureus (P0A006), Staphylococcus aureus
Messens, J.; Van Molle, I.; Vanhaesebrouck, P.; Limbourg, M.; Van Belle, K.; Wahni, K.; Martins, J.C.; Loris, R.; Wyns, L.
How thioredoxin can reduce a buried disulphide bond
J. Mol. Biol.
339
527-537
2004
Staphylococcus aureus (P0A006), Staphylococcus aureus
Roos, G.; Buts, L.; Van Belle, K.; Brosens, E.; Geerlings, P.; Loris, R.; Wyns, L.; Messens, J.
Interplay between ion binding and catalysis in the thioredoxin-coupled arsenate reductase family
J. Mol. Biol.
360
826-838
2006
Bacillus subtilis (P45947), Bacillus subtilis, Staphylococcus aureus (P0A006), Staphylococcus aureus
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