Characterization in Pseudomonas putida Cg1 of nahR and its role in bacterial survival in soil

Park, W.; Madsen, Eugene L.

doi:10.1007/s00253-004-1630-6

Cited by 9 publications

(3 citation statements)

References 39 publications

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“…4); (iii) the phenotype of a nagR mutant (strain CJN110; Fig. 5) matches that of other naphthalene degraders with mutations in LysRtype regulatory genes (33,38); and (iv) the expression pattern of a portion of the genes in strain CJ2 resembles that of other naphthalene degraders (Fig. 3).…”

Section: Discussionmentioning

confidence: 72%

The Naphthalene Catabolic ( nag ) Genes of Polaromonas naphthalenivorans CJ2: Evolutionary Implications for Two Gene Clusters and Novel Regulatory Control

Jeon

Park

et al. 2006

Appl Environ Microbiol

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Polaromonas naphthalenivorans CJ2, found to be responsible for the degradation of naphthalene in situ at a coal tar waste-contaminated site (C.-O. Jeon et al., Proc. Natl. Acad. Sci. USA 100:13591-13596, 2003), is able to grow on mineral salts agar media with naphthalene as the sole carbon source. Beginning from a 484-bp nagAc-like region, we used a genome walking strategy to sequence genes encoding the entire naphthalene degradation pathway and additional flanking regions. We found that the naphthalene catabolic genes in P. naphthalenivorans CJ2 were divided into one large and one small gene cluster, separated by an unknown distance. The large gene cluster (nagRAaGHAbAcAdBFCQEDJIORF1tnpA) is bounded by a LysR-type regulator (nagR). The small cluster (nagR2ORF2I؆KL) is bounded by a MarR-type regulator (nagR2). The catabolic genes of P. naphthalenivorans CJ2 were homologous to many of those of Ralstonia U2, which uses the gentisate pathway to convert naphthalene to central metabolites. However, three open reading frames (nagY, nagM, and nagN), present in Ralstonia U2, were absent. Also, P. naphthalenivorans carries two copies of gentisate dioxygenase (nagI) with 77.4% DNA sequence identity to one another and 82% amino acid identity to their homologue in Ralstonia sp. strain U2. Investigation of the operons using reverse transcription PCR showed that each cluster was controlled independently by its respective promoter. Insertional inactivation and lacZ reporter assays showed that nagR2 is a negative regulator and that expression of the small cluster is not induced by naphthalene, salicylate, or gentisate. Association of two putative Azoarcus-related transposases with the large cluster and one Azoarcus-related putative salicylate 5-hydroxylase gene (ORF2) in the small cluster suggests that mobile genetic elements were likely involved in creating the novel arrangement of catabolic and regulatory genes in P. naphthalenivorans.The degradation of naphthalene has been studied extensively in two Pseudomonas species that carry the archetypal catabolic plasmids, NAH7 (in P. putida G7) and pDTG1 (in P. putida NCIB9816-4) (6,11,41). In both strains, the nah dissimilatory genes are organized into two operons: one coding for the enzymes involved in the conversion of naphthalene to salicylate (naphthalene degradation upper pathway) and another coding for the conversion of salicylate to pyruvate and acetyl coenzyme A via meta-cleavage (naphthalene degradation lower pathway) (2,42,42,54). In contrast to the nah naphthalene catabolic pathway in Pseudomonas species, the nag genes of Ralstonia sp. strain U2 encode the alternative gentisate pathway, which converts naphthalene to fumarate and pyruvate via salicylate and gentisate (14). The nag genes are organized in a single operon (56,57). Despite the contrasts in structural genes and their arrangements, the nah and nag systems exhibit strikingly similar regulation. Transcription of nag and both nah operons is controlled by a single Lys-R-type regulatory protein (NagR and NahR, respe...

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Section: Discussionmentioning

confidence: 72%

The Naphthalene Catabolic ( nag ) Genes of Polaromonas naphthalenivorans CJ2: Evolutionary Implications for Two Gene Clusters and Novel Regulatory Control

Jeon

Park

et al. 2006

Appl Environ Microbiol

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“…5 ). Several naphthalene carbolic proteins have been identified in Pseudomonas sp., including NahR, NahG, NahU, and NahW 15 , 38 – 40 , and these proteins imparted resistance to the toxic effects of naphthalene, hence these bacterial cells can persistent in soils where naphthalene occurs at high concentrations 41 .…”

Section: Discussionmentioning

confidence: 99%

The naphthalene catabolic protein NahG plays a key role in hexavalent chromium reduction in Pseudomonas brassicacearum LZ-4

Huang

Tao

Jiang

et al. 2017

Sci Rep

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Soil contamination by PAH and heavy metals is a growing problem. Here, we showed that a new isolate, Pseudomonas brassicacearum strain LZ-4, can simultaneously degrade 98% of 6 mM naphthalene and reduce 92.4% of 500 μM hexavalent chromium [Cr (VI)] within 68 h. A draft genome sequence of strain LZ-4 (6,219,082 bp) revealed all the genes in the naphthalene catabolic pathway and some known Cr (VI) reductases. Interestingly, genes encoding naphthalene pathway components were upregulated in the presence of Cr (VI), and Cr (VI) reduction was elevated in the presence of naphthalene. We cloned and expressed these naphthalene catabolic genes and tested for Cr (VI) reduction, and found that NahG reduced 79% of 100 μM Cr (VI) in 5 minutes. Additionally, an nahG deletion mutant lost 52% of its Cr (VI) reduction ability compared to that of the wild-type strain. As nahG encodes a salicylate hydroxylase with flavin adenine dinucleotide (FAD) as a cofactor for electron transfer, Cr (VI) could obtain electrons from NADH through NahG-associated FAD. To the best of our knowledge, this is the first report of a protein involved in a PAH-degradation pathway that can reduce heavy metals, which provides new insights into heavy metal-PAH contamination remediation.

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“…Indeed, both the nah and bph operons for naphthalene and biphenyl degradation, respectively, contain multiple LysR-type regulator binding sites which allow for independent transcription of upper and lower pathway genes (Huang & Schell, 1991;Watanabe et al, 2003Watanabe et al, , 2000. The importance of regulating relative levels of catechol degradation activity was hinted at by Park & Madsen (2004) who have shown that regulation by NahR, which preferentially upregulates the lower pathway genes encoding degradation of catechol, was critical for the survival of naphthalene degraders in naphthalene-contaminated soil. This contrasts with the monocistronic nature of the tod operon and the absence of a supplementary promoter directly upstream of todE.…”

Section: Discussionmentioning

confidence: 99%

Growth of Pseudomonas putida F1 on styrene requires increased catechol-2,3-dioxygenase activity, not a new hydrolase

et al. 2011

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Pseudomonas putida F1 cannot grow on styrene despite being able to degrade it through the toluene degradation (tod) pathway. Previous work had suggested that this was because TodF, the meta-fission product (MFP) hydrolase, was unable to metabolize the styrene MFP 2-hydroxy-6-vinylhexa-2,4-dienoate. Here we demonstrate via kinetic and growth analyses that the substrate specificity of TodF is not the limiting factor preventing F1 from growing on styrene. Rather, we found that the metabolite 3-vinylcatechol accumulated during styrene metabolism and that micromolar concentrations of this intermediate inactivated TodE, the catechol-2, 3-dioxygenase (C23O) responsible for its cleavage. Analysis of cells growing on styrene suggested that inactivation of TodE and the subsequent accumulation of 3-vinylcatechol resulted in toxicity and cell death. We found that simply overexpressing TodE on a plasmid (pTodE) was all that was necessary to allow F1 to grow on styrene. Similar results were also obtained by expressing a related C23O, DmpB from Pseudomonas sp. CF600, in tandem with its plant-like ferredoxin, DmpQ (pDmpQB). Further analysis revealed that the ability of F1 (pDmpQB) and F1 (pTodE) to grow on styrene correlated with increased C23O activity as well as resistance of the enzyme to 3-vinylcatechol-mediated inactivation. Although TodE inactivation by 3-halocatechols has been studied before, to our knowledge, this is the first published report demonstrating inactivation by a 3-vinylcatechol. Given the ubiquity of catechol intermediates in aromatic hydrocarbon metabolism, our results further demonstrate the importance of C23O inactivation as a determinant of growth substrate specificity.

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Characterization in Pseudomonas putida Cg1 of nahR and its role in bacterial survival in soil

Cited by 9 publications

References 39 publications

The Naphthalene Catabolic ( nag ) Genes of Polaromonas naphthalenivorans CJ2: Evolutionary Implications for Two Gene Clusters and Novel Regulatory Control

The Naphthalene Catabolic ( nag ) Genes of Polaromonas naphthalenivorans CJ2: Evolutionary Implications for Two Gene Clusters and Novel Regulatory Control

The naphthalene catabolic protein NahG plays a key role in hexavalent chromium reduction in Pseudomonas brassicacearum LZ-4

Growth of Pseudomonas putida F1 on styrene requires increased catechol-2,3-dioxygenase activity, not a new hydrolase

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