Information on EC 1.14.12.3 - benzene 1,2-dioxygenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.14.12.3
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RECOMMENDED NAME
GeneOntology No.
benzene 1,2-dioxygenase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
benzene + NADH + H+ + O2 = cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
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redox reaction
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reduction
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
benzene degradation
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Benzoate degradation
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Chlorocyclohexane and chlorobenzene degradation
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Microbial metabolism in diverse environments
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SYSTEMATIC NAME
IUBMB Comments
benzene,NADH:oxygen oxidoreductase (1,2-hydroxylating)
A system, containing a reductase which is an iron-sulfur flavoprotein (FAD), an iron-sulfur oxygenase and ferredoxin. Requires Fe2+.
CAS REGISTRY NUMBER
COMMENTARY hide
9075-66-5
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
1-methylcyclohexene + O2 + NADH
?
show the reaction diagram
2 cyclohexene + 2 O2 + 2 NADH + 2 H+
2-cyclohexen-1-ol + cis-1,2-cyclohexanediol + 2 NAD+ + H2O
show the reaction diagram
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failure of transformed bacteria to grow on cyclohexene is attributed to the toxicity of metabolic intermediates accumulating from the 2-cyclohexen-1-ol metabolism, cyclohexene metabolism pathways, overview
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?
3-methylcyclohexene + O2 + NADH
?
show the reaction diagram
benzene + NADH + H+ + O2
cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
benzene + NADH + O2
(1R,2S)-cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
benzene + NADH + O2
?
show the reaction diagram
benzene + NADH + O2
cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
benzene + NADPH + O2
cis-cyclohexa-3,5-diene-1,2-diol + NADP+
show the reaction diagram
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activity with NADPH is 10% of the activity with NADH
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?
benzene + O2 + NADH
cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
ethylbenzene + NADH + O2
cis-2,3-dihydroxy-1-ethyl-cyclohexa-4,6-diene + 1-phenethyl alcohol + NAD+
show the reaction diagram
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?
ethylbenzene + O2 + NADH
cis-2,3-dihydroxy-1-ethyl-cyclohexa-4,6-diene + NAD+
show the reaction diagram
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-
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?
toluene + NADH + O2
cis-2,3-dihydroxy-1-methyl-cyclohexa-4,6-diene + NAD+
show the reaction diagram
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?
toluene + O2 + NADH
?
show the reaction diagram
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?
trifluoromethylbenzene + NADH + O2
?
show the reaction diagram
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
1-methylcyclohexene + O2 + NADH
?
show the reaction diagram
2 cyclohexene + 2 O2 + 2 NADH + 2 H+
2-cyclohexen-1-ol + cis-1,2-cyclohexanediol + 2 NAD+ + H2O
show the reaction diagram
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failure of transformed bacteria to grow on cyclohexene is attributed to the toxicity of metabolic intermediates accumulating from the 2-cyclohexen-1-ol metabolism, cyclohexene metabolism pathways, overview
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?
3-methylcyclohexene + O2 + NADH
?
show the reaction diagram
benzene + NADH + H+ + O2
cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
benzene + NADH + O2
(1R,2S)-cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
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bioremediation of aromatic environmental pollutants, initial step of benzene degradation
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?
benzene + NADH + O2
?
show the reaction diagram
benzene + O2 + NADH
cis-cyclohexa-3,5-diene-1,2-diol + NAD+
show the reaction diagram
toluene + O2 + NADH
?
show the reaction diagram
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?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
FAD
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contains FAD
Ferredoxin
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iron-sulfur centre
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[2Fe-2S] cluster
NADPH
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activity with NADPH is 10% of the activity with NADH
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00782 - 0.0112
NADH
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0049
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recombinant enzyme expressed in Escherichia coli, substrate cyclohexene
0.0077
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recombinant enzyme expressed in Escherichia coli, substrate 1-methylcyclohexene
0.0086
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recombinant enzyme expressed in Escherichia coli, substrate 3-methylcyclohexene
0.0172
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recombinant enzyme expressed in Escherichia coli, substrate toluene
0.0236
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recombinant enzyme expressed in Escherichia coli, substrate benzene
0.025
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with ethylbenzene as substrate
0.071
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with toluene as substrate
0.115
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with benzene as substrate
additional information
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
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assay at room temperature
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Pseudomonas putida (strain ATCC 700007 / DSM 6899 / BCRC 17059 / F1)
Pseudomonas putida (strain ATCC 700007 / DSM 6899 / BCRC 17059 / F1)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
11860
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ferredoxin component of the benzene dioxygenase, fast atom bombardment mass spectrometry
12000
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intermediate electron-carrying protein, gel filtration
12300
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intermediate electron-carrying protein, meniscus depletion method
42000
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alpha2, beta2, 2 * 54500 + 2 * 23500, terminal dioxygenase component. 2 * 42000, reductase component. The ferredoxin component has a MW of 12300 Da
55000
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55000 Da is the MW of the alpha-subunit of the terminal dioxygenase
168000
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gel filtration
186000
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terminal dioxygenase component, meniscus depletion method
215000
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terminal dioxygenase component, meniscus depletion method
215300
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terminal dioxygenase component, meniscus depletion method
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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alpha, beta, iron sulfur protein component; the enzyme consist of two dissimilar alpha and beta subunits, alpha subunit structure modeling
tetramer
additional information
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, stable for 6 months to 1 year
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
iron-sulfur proteins of the benzene dioxygenase system: 1.intermediate electron-carrying protein and terminal dioxygenase
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli JM109; expression of wild-type and mutant enzymes in Escherichia coli strains JM109 and CJ236
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expression in Escherichia coli JM109; genes bedC1C2BA, under the control of the tac promoter, subcloned into pLAFR5, successfully conjugated into seven of the Gram-negative cis-1,2-cyclohexanediol-degrading isolates and stably maintained and expressed in three of them, strain CHD1CHD3, the strains grow on cis-1,2-cyclohexanediol as sole carbon source, express an active BDO and oxidise cyclohexene, but none of the three strains is able to grow on cyclohexene as sole carbon source, overview, expression in Escherichia coli strain JM109
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expression in Escherichia coli strain DH5alpha and JM109, co-expression with benzene dihydrodiol dehydrogenase under the control of the Ptac promoter or without any induction. The recombinant strains expressing the BED and the BDDH enzymes transform benzene into dihydrodiol with corresponding consumption of oxygen and regenerate NADH by converting dihydrodiol to catechol
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expression of the alpha-subunit and the beta-subunit of terminal dioxygenase in Escherichia coli
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genes bedC1 and bedC2 encoding the terminal oxygenase alpha-subunit and beta-subunit, expression in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A291S
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reduced activity with ethylbenzene
E444D
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no effect on activity
G404D
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reduced activity with ethylbenzene
H119C
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the mutant alpha-subunit of the terminal dioxygenase is unable to coordinate an EPR-detectable Rieske [2Fe-2S] cluster with the characteristic g factors
H222M
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in the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase the mutant alpha-subunit is unable to reconstitute dioxygenase activity
H228C
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in the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase the mutant alpha-subunit is unable to reconstitute dioxygenase activity
H98C
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the mutant alpha-subunit of the terminal dioxygenase is unable to coordinate an EPR-detectable Rieske [2Fe-2S] cluster with the characteristic g factors, detection of a novel EPR spectrum, the intensity of the spectrum is approximately 8% from the wild-type
I301V
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the mutation in the C-terminal part of subunit alpha enhances the substrate specificity for ethylbenzene, the mutant shows altered patterns of products formed from toluene and ethylbenzene, including monohydroxylated side chains
I301V/T305S/I307L/L309V
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increased activity with ethylbenzene; the mutations in the C-terminal part of subunit alpha enhance the substrate specificity for ethylbenzene, the quadruple mutant also shows a high uncoupled rate of electron transfer without product formation
I307L
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the mutation in the C-terminal part of subunit alpha enhances the substrate specificity for ethylbenzene, the mutant shows altered patterns of products formed from toluene and ethylbenzene, including monohydroxylated side chains
I412V
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reduced activity with ethylbenzene
K436R
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reduced activity with ethylbenzene
L285W
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reduced activity with ethylbenzene
L285W/A291S/G404D
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slightly reduced activity with ethylbenzene
L28W/A291S
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reduced activity with ethylbenzene
L309V
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the mutation in the C-terminal part of subunit alpha enhances the substrate specificity for ethylbenzene, the mutant shows altered patterns of products formed from toluene and ethylbenzene, including monohydroxylated side chains
T305S
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the mutation in the C-terminal part of subunit alpha enhances the substrate specificity for ethylbenzene, the mutant shows altered patterns of products formed from toluene and ethylbenzene, including monohydroxylated side chains
V324I/I327V
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reduced activity with ethylbenzene
Y118S
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the mutant alpha-subunit of the terminal dioxygenase shows an EPR spectrum of half the intensity of that of the wild-type. In the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase it shows significantly reduced activities
Y221A
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in the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase the mutant alpha-subunit shows significantly reduced activity
H119C
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the mutant alpha-subunit of the terminal dioxygenase is unable to coordinate an EPR-detectable Rieske [2Fe-2S] cluster with the characteristic g factors
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H222M
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in the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase the mutant alpha-subunit is unable to reconstitute dioxygenase activity
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H98C
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the mutant alpha-subunit of the terminal dioxygenase is unable to coordinate an EPR-detectable Rieske [2Fe-2S] cluster with the characteristic g factors, detection of a novel EPR spectrum, the intensity of the spectrum is approximately 8% from the wild-type
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Y118S
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the mutant alpha-subunit of the terminal dioxygenase shows an EPR spectrum of half the intensity of that of the wild-type. In the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase it shows significantly reduced activities
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Y221A
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in the reconstitution assay with the reductase component, the ferredoxin component and the beta-subunit of terminal dioxygenase the mutant alpha-subunit shows significantly reduced activity
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additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis