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Information on EC 1.14.13.167 - 4-nitrophenol 4-monooxygenase
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1.14.13.167 4-nitrophenol 4-monooxygenaseIUBMB Comments
Contains FAD. The enzyme catalyses the first step in a degradation pathway for 4-nitrophenol, the oxidation of 4-nitrophenol at position 4 with the concomitant removal of the nitro group as nitrite. The enzyme from the bacterium Pseudomonas sp. strain WBC-3 also catalyses EC 1.14.13.166, 4-nitrocatechol 4-monooxygenase.
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Word Map
- 1.14.13.167
-
sumo
-
ubiquitin-conjugating
-
sumoylation
- hydroxyquinol
-
sumo-conjugating
-
two-component
-
rhodococcus
-
ubiquitin-like
- para-nitrophenol
- hydroquinone
-
1,2-dioxygenase
The enzyme appears in viruses and cellular organisms
Synonyms
4-np/4-nca monooxygenase, 
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SYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
2C4NP 4-monooxygenase
4-NP/4-NCA monooxygenase
NpcA
para-nitrophenol 4-monooxygenase
PdcA
-
-
-
-
PNP 4-monooxygenase
pnpA
PnpA1
pnpA
-
-
-
-
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
4-nitrophenol + NADPH + H+ + O2 = 1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
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PATHWAY SOURCE
PATHWAYS
-
-
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SYSTEMATIC NAME
IUBMB Comments
4-nitrophenol,NAD(P)H:oxygen 4-oxidoreductase (4-hydroxylating, nitrite-forming)
Contains FAD. The enzyme catalyses the first step in a degradation pathway for 4-nitrophenol, the oxidation of 4-nitrophenol at position 4 with the concomitant removal of the nitro group as nitrite. The enzyme from the bacterium Pseudomonas sp. strain WBC-3 also catalyses EC 1.14.13.166, 4-nitrocatechol 4-monooxygenase.
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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
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
3-methyl-4-nitrophenol + NADPH + H+ + O2
2-methyl-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
4-nitrocatechol + NADPH + H+ + O2
?
lower activity compared to 4-nitrophenol
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NAD+ + H2O
-
-
-
?
2,4,6-trinitrophenol + NADH + H+ + O2
?
-
17% activity compared to 4-nitrophenol
-
-
?
2,4,6-trinitrophenol + NADH + H+ + O2
?
-
17% activity compared to 4-nitrophenol
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
Rhodococcus opacus RKJ300
-
-
-
-
?
4-nitrocatechol + NADH + H+ + O2
1,2,4-trihydroxybenzene + nitrite + NAD+ + H2O
-
96% activity compared to 4-nitrophenol
-
-
?
4-nitrocatechol + NADH + H+ + O2
1,2,4-trihydroxybenzene + nitrite + NAD+ + H2O
-
96% activity compared to 4-nitrophenol
-
-
?
4-nitrophenol + NADH + H+ + O2
1,4-benzoquinone + nitrite + NAD+ + H2O
-
-
-
?
4-nitrophenol + NADH + H+ + O2
1,4-benzoquinone + nitrite + NAD+ + H2O
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NADP+ + H2O
Rhodococcus opacus RKJ300
-
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
4-benzoquinone + nitrite + NADP+ + H2O
monooxygenation
-
-
?
4-nitrophenol + NADPH + H+ + O2
4-benzoquinone + nitrite + NADP+ + H2O
-
the enzyme completely degrades 0.24 mM 4-nitrophenol in 36 min
-
-
?
additional information
?
-
no activity with 3-nitrophenol, 2-nitrophenol, 4-nitrobenzoate, nitrobenzene, 3-dinitrobenzene, 2,6-dinitrotoluene, 3-hydroxy-4-nitrobenzoate, 2,4,6-trinitrophenol, and 4-chlorophenol
-
-
?
additional information
?
-
-
almost no activity towards 2,4,6-trinitrophenol
-
-
?
additional information
?
-
-
PnpA1A2 also convert 2-chloro-1,4-benzoquinone to 2-hydroxy-4-nitrophenol, the combined isozymes also show low activity with chlorohydroquinone forming 2-hydroxy-1,4-hydroquinone, metabolite identification by GC-MS analysis, overview. Isozyme PnpA2 is not active with 2-chloro-4-nitrophenol
-
-
?
additional information
?
-
Rhodococcus opacus RKJ300
-
PnpA1A2 also convert 2-chloro-1,4-benzoquinone to 2-hydroxy-4-nitrophenol, the combined isozymes also show low activity with chlorohydroquinone forming 2-hydroxy-1,4-hydroquinone, metabolite identification by GC-MS analysis, overview. Isozyme PnpA2 is not active with 2-chloro-4-nitrophenol
-
-
?
additional information
?
-
-
almost no activity towards 2,4,6-trinitrophenol
-
-
?
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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
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
3-methyl-4-nitrophenol + NADPH + H+ + O2
2-methyl-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
2-chloro-4-nitrophenol + NADPH + H+ + O2
2-chloro-1,4-benzoquinone + nitrite + NADP+ + H2O
Rhodococcus opacus RKJ300
-
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NADP+ + H2O
-
-
-
-
?
4-nitrophenol + NADPH + H+ + O2
1,4-benzoquinone + nitrite + NADP+ + H2O
Rhodococcus opacus RKJ300
-
-
-
-
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
FAD
dependent on, negligible activity is observed when FMN replaces FAD
FAD
the open position of the isoalloxazine ring is anchored by directed or water-mediated hydrogen bonds to the side chains of Ser311 and Arg234. The ribityl moiety of FAD forms hydrogen bonds to Arg52 and Gln115. The phosphate moiety of FAD interacts with the conserved GXGXXG motif and GD motif. The ribose moiety of ADP is held by the hydrogen bonds to Glu42, Ser43 and Glu44. The adenine ring of FAD interacts with Val140 and Thr177
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
addition of Mg2+ or Mn2+ ions has no effect on the activity
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0203
3-Methyl-4-nitrophenol
-
pH and temperature not specified in the publication
0.1374
NADPH
His-tagged enzyme, in 20 mM phosphate buffer (pH 7.0), temperature not specified in the publication
0.012
4-nitrophenol
His-tagged enzyme, in 20 mM phosphate buffer (pH 7.0), temperature not specified in the publication
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.0000046
-
using 2,4,6-trinitrocatechol as substrate, pH and temperature not specified in the publication
0.000025
-
using 4-nitrocatechol as substrate, pH and temperature not specified in the publication
0.0000264
-
using 4-nitrophenol as substrate, pH and temperature not specified in the publication
1.73
-
mutant strain SJ98DELTApnpA1 cell extract, pH 7.5, 30°C, substrate 2-chloro-4-nitrophenol
3.74
-
mutant strain SJ98DELTApnpA1 cell extract, pH 7.5, 30°C, substrate 4-nitrophenol
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
30
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
metabolism
physiological function
malfunction
-
the npcA mutant strain SDA1 completely loses the ability to grow on 4-nitrophenol as the sole carbon source
malfunction
-
the npcA mutant strain SDA1 completely loses the ability to grow on 4-nitrophenol as the sole carbon source
-
metabolism
-
the enzyme is involved in 3-methyl-4-nitrophenol degradation in Burkholderia sp. strain SJ98, phathway overview. Methyl-1,4-benzoquinone (MBQ) and methylhydroquinone (MHQ), rather than catechol proposed previously, are identified as the intermediates before ring cleavage during 3-methyl-4-nitrophenol degradation by Burkholderia sp. strain SJ98. By sequential catalysis assays, PnpCD, PnpE, and PnpF are likely involved in the further pathway of 3-methyl-4-nitrophenol catabolism
metabolism
-
the enzymes responsible for 4-nitrophenol catabolism are also likely to be involved in 2-chloro-4-nitrophenol degradation. Catabolism of 2-chloro-4-nitrophenol and of 4-nitrophenolshare the same gene cluster in strain SJ98
physiological function
para-nitrophenol 4-monooxygenase gene pnpA plays an essential role in 4-nitrophenol mineralization in strain Pseudomonas putida strain WBC-3
physiological function
-
PnpA, a PNP 4-monooxygenase, is able to catalyze the monooxygenation of 2-chloro-4-nitrophenol to 2-chloro-1,4-benzoquinone. PnpB, a 1,4-benzoquinone reductase, has the ability to catalyze the reduction of 2-chloro-1,4-benzoquinone to chlorohydroquinone. Moreover, PnpB is also able to enhance PnpA activity in vitro in the conversion of 2-chloro-4-nitrophenol to 2-chloro-1,4-benzoquinone. Gene pnpA plays an essential role in the degradation of both 2-chloro-4-nitrophenol and 4-nitrophenol. Catabolism of 2-chloro-4-nitrophenol and of 4-nitrophenolshare the same gene cluster in strain SJ98
physiological function
-
PnpA, a PNP 4-monooxygenase, is able to catalyze the monooxygenation of 3-methyl-4-nitrophenol to 3-methyl-1,4-benzoquinone. PnpB, a 1,4-benzoquinone reductase, has the ability to catalyze the reduction of 3-methyl-1,4-benzoquinone to methylhydroquinone. Moreover, PnpB is also able to enhance PnpA activity in vitro in the conversion of 3-methyl-4-nitrophenol to 3-methyl-1,4-benzoquinone
physiological function
-
PnpA1, a PNP 4-monooxygenase, is able to catalyze the monooxygenation of 2-chloro-4-nitrophenol to 2-chloro-1,4-benzoquinone
physiological function
2-chloro-4-nitrophenol serves as a substrate for the 4-monooxygenase PnpA, but wild-type is unable to utilize 2-chloro-4-nitrophenol. Transcriptional regulator PnpR variant S6C drives the 4-nitrophenol-independent expression of PnpA and PnpB operons in an almost constitutive manner and allows higher levels of induction upon addition of inducers. When the pnpRS6C allele is introduced into a PnpR-deleted mutant, the corresponding strain acquires the ability to grow on 2-chloro-4-nitrophenol but is still able to utilize 4-nitrophenol for growth
physiological function
Rhodococcus opacus RKJ300
-
PnpA1, a PNP 4-monooxygenase, is able to catalyze the monooxygenation of 2-chloro-4-nitrophenol to 2-chloro-1,4-benzoquinone
-
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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). A0A2S6UGG3_9PROT
396
1
43331
TrEMBL
-
A0A2S6QEC9_9PROT
409
0
46444
TrEMBL
-
A0A2S6ULC9_9PROT
408
0
47074
TrEMBL
-
A0A2S6QDC1_9PROT
407
0
45197
TrEMBL
-
A0A2S6QCL1_9PROT
399
0
44791
TrEMBL
-
A0A2S6TX41_9PROT
399
1
45192
TrEMBL
-
A0A2P5VJF4_9PROT
399
0
45432
TrEMBL
-
A0A2S6ULT8_9PROT
320
0
36740
TrEMBL
-
A0A2S6TX56_9PROT
402
1
45650
TrEMBL
-
A0A2S6T7D3_9PROT
402
1
46011
TrEMBL
-
A0A2S6Q9S7_9PROT
405
1
45630
TrEMBL
-
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MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
sitting drop vapor diffusion method, using 20 mM HEPES pH 7.0, 15% (w/v) PEG 4000 and 15% (v/v) 2-propanol
structure in its apo and FAD-complex to a resolution of 2.04 A and 2.48 A, respectively. The 4-nitrophenol substrate lies near the N5 edge of isoalloxazine ring in the protein interior. The bound substrate is rotated 60° with respect to pOHB deviating from the isoalloxazine. The nitro group of 4-nitrophenol is directly hydrogen bonded to Val54 and Arg234, and the hydroxyl group is hydrogen bonded to Val223 and Cys23
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
-
enzyme deletion strain SJ98DELTApnpA1 exhibits specific activities for 2-chloro-4-nitrophenol approximately 20% lower than those of wild-type SJ98
additional information
-
generation of pnpA deletion mutant SJ981pnpA1, the mutant shows that pnpA is absolutely necessary for the catabolism of 3-methyl-4-nitrophenol
additional information
-
construction of pK18mobsacB-pnpA1 for gene knockout of pnpA1 in strain RKJ300
additional information
Rhodococcus opacus RKJ300
-
construction of pK18mobsacB-pnpA1 for gene knockout of pnpA1 in strain RKJ300
-
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
0°C, storage medium, no loss of activity after several hours, 30% loss of activity after 12 h, 80% loss of activity after 40 h
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Ni-NTA agarose column chromatography and Sephadex G-100 gel filtration
recombinant N-terminally His6-tagged enzyme PnpA from Escherichia coli strain Rosetta(DE3)pLysS by nickel affinity chromatography
recombinant N-terminally His6-tagged enzyme PnpA from Escherichia coli strain Rosetta(DE3)pLysS by nickel affinity chromatography
-
recombinant N-terminally His6-tagged enzyme PnpA from Escherichia coli strain Rosetta(DE3)pLysS by nickel affinity chromatography
-
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) cells
gene pnpA, expression of the pnp cluster is detected only under inducing conditions, and pnpABA1CDEFG is a single transcriptional operon, genetic structure, RT-PCR and real-time PCR enzyme expression analysis, recombinant expression of N-terminally His6-tagged enzyme PnpA in Escherichia coli strain Rosetta(DE3)pLysS
-
gene pnpA1, recombinant expression of N-terminally His6-tagged enzyme PnpA in Escherichia coli strain Rosetta(DE3)pLysS
-
the enzyme is encoded in the pnpABA1CDEF cluster, which is involved in para-nitrophenol (PNP) and 2-chloro-4-nitrophenol (2C4NP) catabolism and is also likely responsible for 3-methyl-4-nitrophenol (3M4NP) degradation in strain SJ98, real-time quantitative PCR enzyme expression analysis, recombinant expression of N-terminally His6-tagged enzyme PnpA in Escherichia coli strain Rosetta(DE3)pLysS
-
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
0.0018 mg/ml of para-benzoquinone reductase PnpB enhances the 4-nitrophenol degradation activity by para-nitrophenol 4-monooxygenase by about 2fold
2-chloro-4-nitrophenol degradation is induced by either 2-chloro-4-nitrophenol or 4-nitrophenol, negligible activity in uninduced cells
-
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Kitagawa, W.; Kimura, N.; Kamagata, Y.
A novel p-nitrophenol degradation gene cluster from a gram-positive bacterium, Rhodococcus opacus SAO101
J. Bacteriol.
186
4894-4902
2004
Rhodococcus opacus, Rhodococcus opacus SAO101
brenda
Zhang, J.J.; Liu, H.; Xiao, Y.; Zhang, X.E.; Zhou, N.Y.
Identification and characterization of catabolic para-nitrophenol 4-monooxygenase and para-benzoquinone reductase from Pseudomonas sp. strain WBC-3
J. Bacteriol.
191
2703-2710
2009
Pseudomonas sp. (C1I201)
brenda
Wei, M.; Zhang, J.J.; Liu, H.; Zhou, N.Y.
para-Nitrophenol 4-monooxygenase and hydroxyquinol 1,2-dioxygenase catalyze sequential transformation of 4-nitrocatechol in Pseudomonas sp. strain WBC-3
Biodegradation
21
915-921
2010
Pseudomonas sp.
brenda
Liu, W.; Shen, W.; Zhao, X.; Cao, H.; Cui, Z.
Expression, purification, crystallization and preliminary X-ray analysis of para-nitrophenol 4-monooxygenase from Pseudomonas putida DLL-E4
Acta Crystallogr. Sect. F
65
1004-1006
2009
Pseudomonas putida (C6FI48), Pseudomonas putida DLL-E4 (C6FI48)
brenda
Min, J.; Zhang, J.J.; Zhou, N.Y.
The gene cluster for para-nitrophenol catabolism is responsible for 2-chloro-4-nitrophenol degradation in Burkholderia sp. strain SJ98
Appl. Environ. Microbiol.
80
6212-6222
2014
Burkholderia sp., Rhodococcus opacus, Rhodococcus opacus RKJ300
brenda
Min, J.; Lu, Y.; Hu, X.; Zhou, N.Y.
Biochemical characterization of 3-methyl-4-nitrophenol degradation in Burkholderia sp. strain SJ98
Front. Microbiol.
7
791
2016
Burkholderia sp.
brenda
Chen, Q.; Huang, Y.; Duan, Y.; Li, Z.; Cui, Z.; Liu, W.
Crystal structure of p-nitrophenol 4-monooxygenase PnpA from Pseudomonas putida DLL-E4 The key enzyme involved in p-nitrophenol degradation
Biochem. Biophys. Res. Commun.
504
715-720
2018
Pseudomonas putida (C6FI48)
brenda
Deng, S.; Zhang, W.; Wang, J.; Gao, Y.; Xu, Y.; Zhou, N.
Single point mutation in the transcriptional regulator PnpR renders Pseudomonas sp. strain WBC-3 capable of utilizing 2-chloro-4-nitrophenol
Int. Biodeter. Biodegrad.
143
104732
2019
Pseudomonas sp. WBC-3 (C1I201)
-
brenda
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