Information on EC 1.14.12.23 - nitroarene dioxygenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.14.12.23
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RECOMMENDED NAME
GeneOntology No.
nitroarene dioxygenase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
nitrobenzene + NADH + O2 = catechol + nitrite + NAD+
show the reaction diagram
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
phenol degradation
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Aminobenzoate degradation
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Microbial metabolism in diverse environments
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2-nitrotoluene degradation
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2,6-dinitrotoluene degradation
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1-chloro-2-nitrobenzene degradation
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nitrobenzene degradation II
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SYSTEMATIC NAME
IUBMB Comments
nitrobenzene,NADH:oxygen oxidoreductase (1,2-hydroxylating, denitrifying)
This enzyme is a member of the naphthalene family of bacterial Rieske non-heme iron dioxygenases. It comprises a multicomponent system, containing a Rieske [2Fe-2S] ferredoxin, an NADH-dependent flavoprotein reductase (EC 1.18.1.3, ferredoxin---NAD+ reductase), and an alpha3beta3 oxygenase. The enzyme forms of a cis-dihydroxylated product that spontaneously rearranges to form a catechol with accompanying release of nitrite. It can typically act on many different nitroaromatic compounds, including chlorinated species. Enzymes found in different strains may have different substrate preferences. Requires Fe2+.
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
beta-subunit of oxidase
UniProt
Manually annotated by BRENDA team
M9PTH9 i.e. ferredoxin reductase subunit MntAa, M9PW10 i.e. oxygenase alpha subunit MntAc, M9PSS0 i.e. ferredoxin subunit MntAb, M9PV03 i.e. oxygenase beta subunit MntAd
M9PTH9 and M9PW10 and M9PSS0 and M9PV03
UniProt
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
F4Y9I5 i.e. ferredoxin reductase cnbAa, F4Y9I4 i.e. ferredoxin cnbAb, F4Y9J0 i.e. 2-chloronitrobenzene dioxygenase alpha subunit cnbAc, F4Y9J1 i.e. 2-chloronitrobenzene dioxygenase beta subunit cnbAd
F4Y9I5 and F4Y9I4 and F4Y9J0 and F4Y9J1
UniProt
Manually annotated by BRENDA team
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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
physiological function
F4Y9I5 and F4Y9I4 and F4Y9J0 and F4Y9J1
Escherichia coli cells expressing dioxygenase genes cnbAaAbAcAd convert 2-chloronitrobenzene to 3-chlorocatechol with concomitant nitrite release. The cnbCDEF gene cluster, homologous to a 3-chlorocatechol degradation cluster in Sphingomonas sp. strain TFD44, probably contains all of the genes necessary for the conversion of 3-chlorocatechol to 3-oxoadipate
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
1,3-dinitrobenzene + NADH + O2
4-nitrocatechol + nitrite + NAD+
show the reaction diagram
P95561 and Q8RTL5 and Q8RTL4 and Q8RTL3
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100% yield. 89% of the activity with ntrobenzene
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?
2,3-dinitrotoluene + NADH + O2
4-methyl-3-nitrocatechol + nitrite + NAD+
show the reaction diagram
2,4-dinitrotoluene nitrobenzene + NADH + O2
4-methyl-5-nitrocatechol + 4-methyl-3-nitrocatechol + nitrite + NAD+
show the reaction diagram
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products are 4-methyl-5-nitrocatechol + 25% 4-methyl-3-nitrocatechol
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?
2,6-dinitrotoluene + NADH + O2
3-methyl-4-nitrocatechol + nitrite + NAD+
show the reaction diagram
2-amino-4,6-dinitrotoluenedinitrotoluene + NADH + O2
3-amino-4-methyl-5-nitrocatechol + nitrite + NAD+
show the reaction diagram
2-chloronitrobenzene + NADH + O2
3-chlorocatechol + nitrite + NAD+
show the reaction diagram
2-nitrotoluene + NADH + O2
3-methylcatechol + nitrite + NAD+
show the reaction diagram
2-nitrotoluene + NADH + O2
4-methylcatechol + nitrite + NAD+
show the reaction diagram
P95561 and Q8RTL5 and Q8RTL4 and Q8RTL3
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100% yield. 221% of the activity with ntrobenzene
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3-nitrotoluene + NADH + O2
3-methylcatechol + 4-methylcatechol + nitrite + NAD+
show the reaction diagram
M9PTH9 and M9PW10 and M9PSS0 and M9PV03
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products are 54% 3-methylcatechol, 39% 4-methylcatechol, 7% 3-nitrobenzyl alcohol
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3-nitrotoluene + NADH + O2
3-methylcatechol + nitrite + NAD+
show the reaction diagram
3-nitrotoluene + NADH + O2
4-methylcatechol + nitrite + NAD+
show the reaction diagram
4-nitrotoluene + NADH + O2
4-methylcatechol + nitrite + NAD+
show the reaction diagram
naphthalene + NADH + O2
? + nitrite + NAD+
show the reaction diagram
naphthalene + NADH + O2
cis-1,2-dihydroxy-1,2-dihydronaphthalene + NAD+
show the reaction diagram
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about 60% (-)-cis-1,2-dihydroxy-1,2-dihydronaphthalene
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naphthalene + NADH + O2
naphthalene cis-1,2-diol + nitrite + NAD+
show the reaction diagram
P95561 and Q8RTL5 and Q8RTL4 and Q8RTL3
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100% yield. 59% of the activity with ntrobenzene
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naphthalene + O2
cis-1,2-dihydroxy-1,2-dihydronaphthalene
show the reaction diagram
nitrobenzene + NADH + O2
catechol + nitrite + NAD+
show the reaction diagram
additional information
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
NADH
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0.5 mol of NADH is required to reduce each alphabeta heterodimer of oxygenase
[2Fe-2S]-center
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Iron
active site of alpha subunit contains a mononuclear iron atom
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.012 - 0.851
2,6-dinitrotoluene
0.0012 - 0.02
2-nitrotoluene
0.0039 - 0.04
3-nitrotoluene
0.121
Naphthalene
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pH 6.8, 30°C
0.0045 - 1.78
nitrobenzene
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.12 - 2.3
2-nitrotoluene
0.13 - 0.27
3-nitrotoluene
2.4
Naphthalene
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pH 6.8, 30°C
0.19 - 0.3
nitrobenzene
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
12
2-nitrotoluene
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pH 6.8, 30°C
65
3-nitrotoluene
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pH 6.8, 30°C
67
nitrobenzene
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pH 6.8, 30°C
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25300
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3 * 51200, alpha-subunit, 3 * 25300, beta-subunit, SDS-PAGE of oxygenase component
25500
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3 * 50400, alpha-subunit, 3 * 25500, beta-subunit, SDS-PAGE of oxygenase component
50400
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3 * 50400, alpha-subunit, 3 * 25500, beta-subunit, SDS-PAGE of oxygenase component
51200
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3 * 51200, alpha-subunit, 3 * 25300, beta-subunit, SDS-PAGE of oxygenase component
210000
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gel filtration, oxygenase component
215900
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dynamic light scattering, oxygenase component
216000
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gel filtration, oxygenase component
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexamer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
oxygenase component, to 1.2 A resolution, hexagonal space group P63 with cell dimensions a = b = 121.6 A and c = 84.4 A
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native enzyme and in complex with nitrobenzene and 3-nitrotoluene, to 1.2 A, 1.55 A and 1.5 A resolution, respectively. Enzyme shows a mushroom-shaped alpha3beta3 hexamer. The catalytic subunit contains a Rieske iron-sulfur center and an active-site mononuclear iron atom. The substrate pocket contains an asparagine residue that forms a hydrogen bond to the nitro-group of the substrate, and orients the substrate in relation to the active-site mononuclear iron atom
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
M9PTH9 and M9PW10 and M9PSS0 and M9PV03
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A405G
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synthesizes 17% 3-methylcatechol, 83% 4-methylcatechol from substrate 3-nitrotoluene
I204A
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synthesizes 2% 3-methylcatechol, 97% 4-methylcatechol and 1% 3-nitrobenzyl alcohol from substrate 3-nitrotoluene
I204T
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synthesizes 5% 3-methylcatechol, 93% 4-methylcatechol and 2% 3-nitrobenzyl alcohol from substrate 3-nitrotoluene
I204V
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synthesizes 13% 3-methylcatechol, 83% 4-methylcatechol and 5% 3-nitrobenzyl alcohol from substrate 3-nitrotoluene
A405G
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synthesizes 17% 3-methylcatechol, 83% 4-methylcatechol from substrate 3-nitrotoluene
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I204A
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synthesizes 2% 3-methylcatechol, 97% 4-methylcatechol and 1% 3-nitrobenzyl alcohol from substrate 3-nitrotoluene
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I204T
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synthesizes 5% 3-methylcatechol, 93% 4-methylcatechol and 2% 3-nitrobenzyl alcohol from substrate 3-nitrotoluene
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I204V
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synthesizes 13% 3-methylcatechol, 83% 4-methylcatechol and 5% 3-nitrobenzyl alcohol from substrate 3-nitrotoluene
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F293H
mutation in alpha-subunit of oxygenase, residue is important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene
F293I
mutation in alpha-subunit of oxygenase, residue is important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene
F293Q
mutation in alpha-subunit of oxygenase, residue is important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene. Mutant is 2.5 times faster than wild-type at oxidizing 2,6-dinitrotoluene while retaining a similar Km for the substrate
I350F
mutation in alpha-subunit of oxygenase, residue is important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene. Most mononitrotoluene oxidation products formed by the N258V mutant are nitrobenzyl alcohols rather than catechols
I350T
mutation in alpha-subunit of oxygenase, residue is important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene
N258V
mutation in alpha-subunit of oxygenase, residue is important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene. Up to 99% of the mononitrotoluene oxidation products formed by the N258V mutant are nitrobenzyl alcohols rather than catechols
F350T
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contrary to wild-type, mutant produces 3-methyl-4-nitrocatechol from 2,6-dinitrotoluene and 3-amino-4-methyl-5-nitrocatechol and 2-amino-4,6-dinitrobenzyl alcohol from 2-amino-4,6-dinitrotoluene, and mutant releases nitrite from 2,3-dinitrotoluene sixfold faster than wild-type
F350T/G407S
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double mutant increases the rate of oxidation of 2,3-dinitrotoluene, 2,6-dinitrotoluene , and 2-amino-4,6-dinitrotoluene threefold relative to variant F350T
G50S/L225R/A269T
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mutant displays increased rate of 4-amino-2-nitrotoluene oxidation and additionally produces oxidation product 4-amino-2-nitrocresol (enhanced 11fold relative to wild-type) as well as 4-amino-2-nitrobenzyl alcohol
L225R
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12fold faster generation of 4-amino-2-nitrocresol and production of 4-amino-2-nitrobenzyl alcohol from 4-amino-2-nitrotoluene as well as 24fold faster generation of nitrite and 15fold faster generation of 2,3-dinitrobenzyl alcohol from 2,3-dinitrotoluene
F350T
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contrary to wild-type, mutant produces 3-methyl-4-nitrocatechol from 2,6-dinitrotoluene and 3-amino-4-methyl-5-nitrocatechol and 2-amino-4,6-dinitrobenzyl alcohol from 2-amino-4,6-dinitrotoluene, and mutant releases nitrite from 2,3-dinitrotoluene sixfold faster than wild-type
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F350T/G407S
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double mutant increases the rate of oxidation of 2,3-dinitrotoluene, 2,6-dinitrotoluene , and 2-amino-4,6-dinitrotoluene threefold relative to variant F350T
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G50S/L225R/A269T
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mutant displays increased rate of 4-amino-2-nitrotoluene oxidation and additionally produces oxidation product 4-amino-2-nitrocresol (enhanced 11fold relative to wild-type) as well as 4-amino-2-nitrobenzyl alcohol
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L225R
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12fold faster generation of 4-amino-2-nitrocresol and production of 4-amino-2-nitrobenzyl alcohol from 4-amino-2-nitrotoluene as well as 24fold faster generation of nitrite and 15fold faster generation of 2,3-dinitrobenzyl alcohol from 2,3-dinitrotoluene
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additional information