Information on EC 1.20.99.1 - arsenate reductase (donor)

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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota

EC NUMBER
COMMENTARY hide
1.20.99.1
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RECOMMENDED NAME
GeneOntology No.
arsenate reductase (donor)
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
arsenite + acceptor = arsenate + reduced acceptor
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
redox reaction
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
arsenate reduction (respiratory)
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arsonoacetate degradation
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SYSTEMATIC NAME
IUBMB Comments
arsenate:acceptor oxidoreductase
Benzyl viologen can act as an acceptor. Unlike EC 1.20.4.1, arsenate reductase (glutaredoxin), reduced glutaredoxin cannot serve as a reductant.
CAS REGISTRY NUMBER
COMMENTARY hide
146907-46-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
enzyme is expressed under chemolithoautotrophic conditions and exhibits both arsenite oxidase and arsenate reductase activity
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Manually annotated by BRENDA team
isolated from arsenic contaminated soil. 30% of arsenic-resistant isolates are found to carry an arsenate reductase encoded by the arsC gene, suggesting that the arsC gene may be diverse in these isolates or that these isolates have other arsenic-resistant mechanisms rather than containing arsC
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Manually annotated by BRENDA team
isolated from arsenic contaminated soil. 30% of arsenic-resistant isolates are found to carry an arsenate reductase encoded by the arsC gene, suggesting that the arsC gene may be diverse in these isolates or that these isolates have other arsenic-resistant mechanisms rather than containing arsC
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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UniProt
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
strain ANA-3
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Manually annotated by BRENDA team
strain ANA-3
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
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anaerobic growth of Pseudomonas putida WB on glucose is shown by arsenate reductase ArsC with energy derived only from substrate level phosphorylation. Two mol of acetate are generated intermediary and the reducing equivalents of glycolysis and pyruvate decarboxylation serve for arsenate reduction or are released as H2; anaerobic growth on acetate and lactate is shown by arsenate reductase ArrA coupled to respiratory electron chain energy conservation. In the presence of arsenate, both substrates are totally oxidized to CO2 and H2 with part of the H2 serving for respiratory arsenate reduction to deliver energy for growth
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
arsenate + ?
arsenite + ?
show the reaction diagram
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?
arsenate + acetate
arsenite + ?
show the reaction diagram
arsenate + benzyl viologen
arsenite + oxidized benzyl viologen
show the reaction diagram
arsenate + methyl viologen
arsenite + oxidized methyl viologen
show the reaction diagram
arsenate + reduced acceptor
arsenite + acceptor
show the reaction diagram
arsenate + reduced glutaredoxin
arsenite + acceptor
show the reaction diagram
arsenite + acceptor
arsenate + reduced acceptor
show the reaction diagram
arsenite + azurin + H2O
arsenate + reduced azurin
show the reaction diagram
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?
arsenite + cytochrome c + H2O
arsenate + reduced cytochrome c
show the reaction diagram
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?
arsenite + cytochrome c-551
arsenate + cytochrome c-551
show the reaction diagram
arsenite + cytochrome c-551
arsenate + reduced cytochrome c-551
show the reaction diagram
arsenite + oxidized 2,6-dichlorophenolindophenol
arsenate + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
sodium stibo(V)gluconate + reduced acceptor
sodium stibo(III)gluconate + acceptor
show the reaction diagram
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i.e. pentostam, reduction of antimony(V) to antimony(III)
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?
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
arsenate + acetate
arsenite + ?
show the reaction diagram
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anaerobic respiration using arsenate as terminal electron acceptor and acetate as the electron donor
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?
arsenate + reduced acceptor
arsenite + acceptor
show the reaction diagram
arsenite + acceptor
arsenate + reduced acceptor
show the reaction diagram
arsenite + azurin + H2O
arsenate + reduced azurin
show the reaction diagram
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?
arsenite + cytochrome c + H2O
arsenate + reduced cytochrome c
show the reaction diagram
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?
arsenite + cytochrome c-551
arsenate + reduced cytochrome c-551
show the reaction diagram
additional information
?
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the arsRDABC plasmid operon codes for an ATP-dependent anion pump and is responsible for resistance against arsenate, arsenite, antimonite, and tellurite
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cytochrome c-551
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Molybdenum
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cofactor
pterin
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inequivalent P- and Q-pterins coordinated to the molybdenum ion, structure analysis
Pterin molybdenum cofactor
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
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increases activity
Cd2+
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increases activity
Cu2+
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increases activity
Fe2+
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increases activity
Molybdenum
sulfate
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binds at the active site as arsenate analogue
sulfite
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binds at the active site as arsenite analogue
Zinc
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it is possible that one zinc atom represents an additional cofactor
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
arsenate
citrate
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dithiothreitol
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glutathione and dithiothreitol in combination enhance arsenate reduction in vitro more than glutathione alone
galactose
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glucose
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glutathione
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glutathione and dithiothreitol in combination enhance arsenate reduction in vitro more than glutathione alone
selenate
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thiol
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required for activity, thiol requirement can be satisfied by dithiothreitol, stimulation by glutathione, thioredoxin or reduced lipoc acid is negligible compared to that of dithiothreitol
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.3 - 24.4
arsenate
0.035 - 0.061
arsenite
0.1 - 0.2
reduced glutaredoxin
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additional information
additional information
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electrochemical enzyme properties determination by noncatalytic and catalytic voltrammetry, temperature and pH-dependeny
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.012 - 14400
arsenate
30.4
arsenite
Hydrogenophaga sp. NT-14
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pH 5.5, calculated with an Mr of 309000 Da
0.19 - 0.26
reduced glutaredoxin
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0034 - 0.027
arsenate
255
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0007
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5.6
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purified enzyme
6.68
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in the presence of 100 micromol arsenate, 30C, pH 7
12.15
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in the presence of 200 micromol arsenate, 30C, pH 7
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.2
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optimal pH
7
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assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 7.5
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
14000
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2 * 98000, AroA, + 2 * 14000, AroB, alpha2beta2, SDS-PAGE
14330
sequence analysis
14500
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x * 14500, calculated
14960
sequence analysis
16000
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3 * 86000, AroA, + 3 * 16000, AroB, alpha3beta3, SDS-PAGE
29000
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alpha,beta, 1 * 87000, ArrA, + 1 * 29000, ArrB, SDS-PAGE
71800
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gel filtration
86000
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3 * 86000, AroA, + 3 * 16000, AroB, alpha3beta3, SDS-PAGE
87000
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alpha,beta, 1 * 87000, ArrA, + 1 * 29000, ArrB, SDS-PAGE
98000
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2 * 98000, AroA, + 2 * 14000, AroB, alpha2beta2, SDS-PAGE
123000
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gel filtration
219000
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gel filtration
306000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 14500, calculated
hexamer
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3 * 86000, AroA, + 3 * 16000, AroB, alpha3beta3, SDS-PAGE
tetramer
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2 * 98000, AroA, + 2 * 14000, AroB, alpha2beta2, SDS-PAGE
additional information
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crystal structure and active site structure analysis
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
X-ray diffraction structure determination and analysis of 2 different crystal forms at 1.64-2.03 A using multiple isomorphous replacement with anomalous scattering and multiple-wavelength anomalous dispersion methods
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purified recombinant wild-type and SeMet-labeled enzyme, 4C, PEG or PEG methyl ether is utilized as main precipitant, 2 crystal forms, X-ray diffraction structure determination and analysis at 2.2 A, flash-annealing technique
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purified wild-type and selenomethionine-labeled enzyme, hanging drop vapour diffusion method, enzyme in 0.1 M sodium acetate, pH 4.4-4.6, 0.2 M ammonium sulfate, in presence of 30-35% PEG methyl ether, and 5 mM DTT, crystal structure in reduced form, X-ray diffraction structure determination and analysis at 1.6-2.4 A resolution
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solution structure and backbone dynamics of both the reduced and oxidized forms of enzyme. Reduced from undergoes millisecond conformational changes in the functional P-loop and C82-C89. In the oxidized form, C82-C89 shows motional flexibility on both picosecond-to-nanosecond and possibly millisecond scales
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solution structure of enzyme in complex with thioarsenate represents the transiently formed intermediate during the intermolecualr thiol-disulfide exchange reaction. Substantial conformational changes are coupled to the reaction process
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free enzyme or enzyme complexed with arsenate and arsenite, X-ray diffraction structure determination and analysis at 1.65 A and 1.26 A, respecively; in complex with arsenitate and arsenite and enzyme alone. Native structure shows sulfate and sulfite ions binding in the active site as analogs of arsenate and arsenite. Arsenate forms a covlaent adduct with C12 in the active site, showing tetrahedral geometry. The corresponding adduct with arsenite binds as thiarsahydroxy adduct. High number of 385 water molecules bound to crystal structure
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wild-type and mutant R60K and R60A enzymes in complex with product arsenite, X-ray diffraction structure determination and analysis at 1.3-1.8 A resolution
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recombinant protein, space group P321
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
57.5fold to homogeneity from the periplasm by preparation of spheroplasts, ammonium sulfate fractionation, ultrafiltration, cation exchange chromatography, and gel filtration, purification via anion exchange or hydrophobic interaction chromatography is not successful
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by nickel-nitrilotriacetic acid chromatography; by nickel-nitrilotriacetic acid chromatography
partial
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recombinant soluble wild-type and SeMet-labeled enzyme from Escherichia coli
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using Ni-NTA chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
catalytic domain is expressed in Escherichia coli as a His-tagged fusion protein
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expressed in an Escherichia coli strain in which the arsC gene is deleted and in a yeast (Saccharomyces cerevisiae) strain with a disrupted ACR2 gene; expressed in an Escherichia coli strain in which the arsC gene is deleted and in a yeast (Saccharomyces cerevisiae) strain with a disrupted ACR2 gene
expression of wild-type and SeMet-labeled enzyme in Escherichia coli
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
catalytic domain of Cdc25C is expressed in bacteria. Cdc25B shows arsenate reductase activity. Enzyme reduces inorganic arsenate but not methylated pentavalent arsenicals
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
R60A
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site-directed muatgenesis, crystal structure determination and analysis with bound arsenite, formation of a covalent and more stableCys12-thiol-dihydroxyarsenite intermediate compared to the wild-type enzyme, resulting in slow release of product and loss of activity
R60K
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site-directed muatgenesis, crystal structure determination and analysis with bound arsenite, formation of a covalent and more stableCys12-thiol-dihydroxyarsenite intermediate compared to the wild-type enzyme, resulting in slow release of product and loss of activity
C120A
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inactive mutant
R126A
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inactive mutant
C71S
mutagenesis of cysteine residues in the putative active site HC(X) 5 R motif leads to nearly complete loss of both phosphatase and arsenate reductase activities; mutagenesis of cysteine residues in the putative active site HC(X) 5 R motif leads to nearly complete loss of both phosphatase and arsenate reductase activities
additional information
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catalytic domain of Cdc25B is expressed in bacteria. Cdc25B shows arsenate reductase activity. Enzyme reduces inorganic arsenate but not methylated pentavalent arsenicals. Alteration of either the cysteine and arginine residues of the Cys-X5-Arg motif leads to the loss of both reductase and phosphatase activities