Cloning of a gene cluster involved in the catabolism of p-nitrophenol by Arthrobacter sp. strain JS443 and characterization of the p-nitrophenol monooxygenase

Lynda L. Perry, Gerben Zylstra

Research output: Contribution to journalArticle

67 Citations (Scopus)

Abstract

The npd gene cluster, which encodes the enzymes of a p-nitrophenol catabolic pathway from Arthrobacter sp. strain JS443, was cloned and sequenced. Three genes, npdB, npdA1, and npdA2, were independently expressed in Escherichia coli in order to confirm the identities of their gene products. NpdA2 is a p-nitrophenol monooxygenase belonging to the two-component flavin-diffusible monooxygenase family of reduced flavin-dependent monooxygenases. NpdA1 is an NADH-dependent flavin reductase, and NpdB is a hydroxyquinol 1,2-dioxygenase. The npd gene cluster also includes a putative maleylacetate reductase gene, npdC. In an in vitro assay containing NpdA2, an E. coli lysate transforms p-nitrophenol stoichiometrically to hydroquinone and hydroxyquinol. It was concluded that the p-nitrophenol catabolic pathway in JS443 most likely begins with a two-step transformation of p-nitrophenol to hydroxy-1,4-benzoquinone, catalyzed by NpdA2. Hydroxy-1,4-benzoquinone is reduced to hydroxyquinol, which is degraded through the hydroxyquinol ortho cleavage pathway. The hydroquinone detected in vitro is a dead-end product most likely resulting from chemical or enzymatic reduction of the hypothetical intermediate 1,4-benzoquinone. NpdA2 hydroxylates a broad range of chloro- and nitro-substituted phenols, resorcinols, and catechols. Only p-nitro- or p-chloro-substituted phenols are hydroxylated twice. Other substrates are hydroxylated once, always at a position para to a hydroxyl group.

Original languageEnglish (US)
Pages (from-to)7563-7572
Number of pages10
JournalJournal of bacteriology
Volume189
Issue number21
DOIs
StatePublished - Nov 1 2007

Fingerprint

Arthrobacter
Multigene Family
Organism Cloning
Phenols
maleylacetate reductase
Mixed Function Oxygenases
Resorcinols
Catechols
Escherichia coli
Genes
Hydroxyl Radical
NAD
Oxidoreductases
4-nitrophenol monooxygenase
4-nitrophenol
Enzymes
4,6-dinitro-o-cresol
benzoquinone
hydroxyhydroquinone
In Vitro Techniques

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

Cite this

@article{aeecb61bfba24bbf99180d9fa4bb8ad1,
title = "Cloning of a gene cluster involved in the catabolism of p-nitrophenol by Arthrobacter sp. strain JS443 and characterization of the p-nitrophenol monooxygenase",
abstract = "The npd gene cluster, which encodes the enzymes of a p-nitrophenol catabolic pathway from Arthrobacter sp. strain JS443, was cloned and sequenced. Three genes, npdB, npdA1, and npdA2, were independently expressed in Escherichia coli in order to confirm the identities of their gene products. NpdA2 is a p-nitrophenol monooxygenase belonging to the two-component flavin-diffusible monooxygenase family of reduced flavin-dependent monooxygenases. NpdA1 is an NADH-dependent flavin reductase, and NpdB is a hydroxyquinol 1,2-dioxygenase. The npd gene cluster also includes a putative maleylacetate reductase gene, npdC. In an in vitro assay containing NpdA2, an E. coli lysate transforms p-nitrophenol stoichiometrically to hydroquinone and hydroxyquinol. It was concluded that the p-nitrophenol catabolic pathway in JS443 most likely begins with a two-step transformation of p-nitrophenol to hydroxy-1,4-benzoquinone, catalyzed by NpdA2. Hydroxy-1,4-benzoquinone is reduced to hydroxyquinol, which is degraded through the hydroxyquinol ortho cleavage pathway. The hydroquinone detected in vitro is a dead-end product most likely resulting from chemical or enzymatic reduction of the hypothetical intermediate 1,4-benzoquinone. NpdA2 hydroxylates a broad range of chloro- and nitro-substituted phenols, resorcinols, and catechols. Only p-nitro- or p-chloro-substituted phenols are hydroxylated twice. Other substrates are hydroxylated once, always at a position para to a hydroxyl group.",
author = "Perry, {Lynda L.} and Gerben Zylstra",
year = "2007",
month = "11",
day = "1",
doi = "10.1128/JB.01849-06",
language = "English (US)",
volume = "189",
pages = "7563--7572",
journal = "Journal of Bacteriology",
issn = "0021-9193",
publisher = "American Society for Microbiology",
number = "21",

}

TY - JOUR

T1 - Cloning of a gene cluster involved in the catabolism of p-nitrophenol by Arthrobacter sp. strain JS443 and characterization of the p-nitrophenol monooxygenase

AU - Perry, Lynda L.

AU - Zylstra, Gerben

PY - 2007/11/1

Y1 - 2007/11/1

N2 - The npd gene cluster, which encodes the enzymes of a p-nitrophenol catabolic pathway from Arthrobacter sp. strain JS443, was cloned and sequenced. Three genes, npdB, npdA1, and npdA2, were independently expressed in Escherichia coli in order to confirm the identities of their gene products. NpdA2 is a p-nitrophenol monooxygenase belonging to the two-component flavin-diffusible monooxygenase family of reduced flavin-dependent monooxygenases. NpdA1 is an NADH-dependent flavin reductase, and NpdB is a hydroxyquinol 1,2-dioxygenase. The npd gene cluster also includes a putative maleylacetate reductase gene, npdC. In an in vitro assay containing NpdA2, an E. coli lysate transforms p-nitrophenol stoichiometrically to hydroquinone and hydroxyquinol. It was concluded that the p-nitrophenol catabolic pathway in JS443 most likely begins with a two-step transformation of p-nitrophenol to hydroxy-1,4-benzoquinone, catalyzed by NpdA2. Hydroxy-1,4-benzoquinone is reduced to hydroxyquinol, which is degraded through the hydroxyquinol ortho cleavage pathway. The hydroquinone detected in vitro is a dead-end product most likely resulting from chemical or enzymatic reduction of the hypothetical intermediate 1,4-benzoquinone. NpdA2 hydroxylates a broad range of chloro- and nitro-substituted phenols, resorcinols, and catechols. Only p-nitro- or p-chloro-substituted phenols are hydroxylated twice. Other substrates are hydroxylated once, always at a position para to a hydroxyl group.

AB - The npd gene cluster, which encodes the enzymes of a p-nitrophenol catabolic pathway from Arthrobacter sp. strain JS443, was cloned and sequenced. Three genes, npdB, npdA1, and npdA2, were independently expressed in Escherichia coli in order to confirm the identities of their gene products. NpdA2 is a p-nitrophenol monooxygenase belonging to the two-component flavin-diffusible monooxygenase family of reduced flavin-dependent monooxygenases. NpdA1 is an NADH-dependent flavin reductase, and NpdB is a hydroxyquinol 1,2-dioxygenase. The npd gene cluster also includes a putative maleylacetate reductase gene, npdC. In an in vitro assay containing NpdA2, an E. coli lysate transforms p-nitrophenol stoichiometrically to hydroquinone and hydroxyquinol. It was concluded that the p-nitrophenol catabolic pathway in JS443 most likely begins with a two-step transformation of p-nitrophenol to hydroxy-1,4-benzoquinone, catalyzed by NpdA2. Hydroxy-1,4-benzoquinone is reduced to hydroxyquinol, which is degraded through the hydroxyquinol ortho cleavage pathway. The hydroquinone detected in vitro is a dead-end product most likely resulting from chemical or enzymatic reduction of the hypothetical intermediate 1,4-benzoquinone. NpdA2 hydroxylates a broad range of chloro- and nitro-substituted phenols, resorcinols, and catechols. Only p-nitro- or p-chloro-substituted phenols are hydroxylated twice. Other substrates are hydroxylated once, always at a position para to a hydroxyl group.

UR - http://www.scopus.com/inward/record.url?scp=35649004922&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=35649004922&partnerID=8YFLogxK

U2 - 10.1128/JB.01849-06

DO - 10.1128/JB.01849-06

M3 - Article

C2 - 17720792

AN - SCOPUS:35649004922

VL - 189

SP - 7563

EP - 7572

JO - Journal of Bacteriology

JF - Journal of Bacteriology

SN - 0021-9193

IS - 21

ER -