The Induction of the Enzymes of Naphthalene Metabolism in Pseudomonads by Salicylate and 2-Aminobenzoate

Barnsley, E. A.

doi:10.1099/00221287-88-1-193

Cited by 65 publications

(34 citation statements)

References 10 publications

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“…Strain 9816/11 was grown at 30ЊC in mineral salts basal medium (MSB) (49) with 0.2% (wt/vol) pyruvate as a carbon source in the presence of 0.05% (wt/vol) salicylate or anthranilate. These aromatic acids induce the synthesis of naphthalene catabolic enzymes in strain 9816 (2). Transformations of indan and indene, except where noted otherwise, were conducted with washed cell suspensions (turbidity, 2.0 to 2.5 at 600 nm) in 200 or 800 ml of 50 mM sodium-potassium phosphate buffer (pH 7.2) in 1.0-liter Erlenmeyer or 2.8-liter Fernbach flasks, respectively.…”

Section: Methodsmentioning

confidence: 99%

Desaturation, dioxygenation, and monooxygenation reactions catalyzed by naphthalene dioxygenase from Pseudomonas sp. strain 9816-4

et al. 1995

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Pseudomonas sp. strain 9816-4 grows with naphthalene as the sole source of carbon and energy (9). The initial reaction is catalyzed by a multicomponent enzyme system designated naphthalene dioxygenase (NDO) (11,12,23,24). NDO catalyzes the NAD(P)H-dependent enantiospecific incorporation of dioxygen into naphthalene to form (ϩ)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene (cis-naphthalene dihydrodiol) (26, 27) ( Fig. 1). An analogous reaction is catalyzed by toluene dioxygenase (TDO) from Pseudomonas putida F1, where enantiomerically pure (ϩ)-cis-(1S,2R)-dihydroxy-3-methylcyclohexa-3,5-diene (cis-toluene dihydrodiol) is the first detectable oxidation product (17,31,60). TDO also catalyzes the enantiospecific oxidation of naphthalene to (ϩ)-cis-naphthalene dihydrodiol (18,39).In addition to the enantiospecific oxidation of naphthalene and toluene, NDO and TDO from the above strains oxidize many related aromatic compounds to optically active dihydrodiols (10,18,28,30). Other bacterial dioxygenases show similar properties, and more than 130 chiral arene cis-dihydrodiols have been produced from a small number of strains (7,35,48). The high enantiomeric purity of these compounds has led to their use as chiral synthons in the enantiospecific synthesis of a wide variety of biologically active natural products (7,8,46,57). The present studies focus on another facet of this interesting group of dioxygenases, that is, their ability to catalyze reactions other than the formation of arene cis-dihydrodiols. For example, the TDO expressed by P. putida F39/D oxidizes indan to (1R)-indanol and oxidizes indene to cis-(1S,2R)-indandiol and (1S)-indenol (55). Similar reactions have been reported for TDO from P. putida UV4, although the 1-indenol produced by this strain is the (1R)-enantiomer (3, 5).We now report the identification and absolute stereochemistry of the products formed from indan and indene by NDO from Pseudomonas sp. strain 9816-4 and confirm earlier observations on the desaturation of indan to indene by NDO (22). MATERIALS AND METHODSOrganisms. Pseudomonas sp. strain 9816/11 is a mutant which oxidizes naphthalene stoichiometrically to (ϩ)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene (40). This organism is a derivative of Pseudomonas sp. strain 9816-4 (9, 59), which harbors the genes for naphthalene catabolism on an 83-kb NAH plasmid designated pDTG1 (45). Pseudomonas sp. strain 9816/C84, a cured strain, was used as a control in experiments with strain 9816/11. Escherichia coli strain JM109 (DE3)[pDTG141] contains the structural genes (nahAaAbAcAd) for NDO in plasmid pT7-5 (50). Expression of NDO in this strain is inducible by the addition of isopropylthiogalactopyranoside (IPTG). E. coli JM109(DE3)[pT7-5] was used as a control in experiments with strain JM109(DE3) [pDTG141].Biotransformation experiments. Strain 9816/11 was grown at 30ЊC in mineral salts basal medium (MSB) (49) with 0.2% (wt/vol) pyruvate as a carbon source in the presence of 0.05% (wt/vol) salicylate or anthranilate. These aromatic acids induce the s...

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Section: Methodsmentioning

confidence: 99%

Desaturation, dioxygenation, and monooxygenation reactions catalyzed by naphthalene dioxygenase from Pseudomonas sp. strain 9816-4

et al. 1995

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“…strain NCIB 9816/11 is a naphthalene dihydrodiol dehydrogenase mutant which oxidizes naphthalene to (ϩ)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene (22). Strain NCIB 9816/11 was grown as described above and was induced with 0.05% (wt/vol) salicylate (3). Induced cells were harvested in the late exponential phase of growth by centrifugation (9,000 ϫ g; 10 min; 4ЊC), washed once in MSB, and resuspended in 100 and 500 ml of MSB for kinetic experiments and isolation of metabolites, respectively.…”

Section: Methodsmentioning

confidence: 99%

Desaturation and oxygenation of 1,2-dihydronaphthalene by toluene and naphthalene dioxygenase

et al. 1995

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Bacterial strains expressing toluene and naphthalene dioxygenase were used to examine the sequence of reactions involved in the oxidation of 1,2-dihydronaphthalene. Toluene dioxygenase of Pseudomonas putida F39/D oxidizes 1,2-dihydronaphthalene to (؉)-cis-(1S,2R)-dihydroxy-1,2,3,4-tetrahydronaphthalene, (؉)-(1R)-hydroxy-1,2-dihydronaphthalene, and (؉)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene. In contrast, naphthalene dioxygenase of Pseudomonas sp. strain NCIB 9816/11 oxidizes 1,2-dihydronaphthalene to the opposite enantiomer, (؊)-cis-(1R,2S)-dihydroxy-1,2,3,4-tetrahydronaphthalene and the identical (؉)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene. Recombinant Escherichia coli strains expressing the structural genes for toluene and naphthalene dioxygenases confirmed the involvement of these enzymes in the reactions catalyzed by strains F39/D and NCIB 9816/11. 1-Hydroxy-1,2-dihydronaphthalene was not formed by strains expressing naphthalene dioxygenase. These results coupled with time course studies and deuterium labelling experiments indicate that, in addition to direct dioxygenation of the olefin, both enzymes have the ability to desaturate (dehydrogenate) 1,2-dihydronaphthalene to naphthalene, which serves as a substrate for cis dihydroxylation.Pseudomonas putida F1 and Pseudomonas sp. strain NCIB 9816-4 initiate the oxidation of toluene and naphthalene by the addition of both atoms of molecular oxygen and two hydrogen atoms to the aromatic nucleus to form (ϩ)-cis-(1S,2R)-dihydroxy-3-methylcyclohexa-3,5-diene (cis-toluene dihydrodiol) (14, 21) and (ϩ)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene (cis-naphthalene dihydrodiol) (19,20), respectively. These reactions are catalyzed by multicomponent enzyme systems designated toluene dioxygenase (TDO) (16, 31) and naphthalene dioxygenase (NDO) (13).Current interest in TDO and NDO stems from the fact that, in addition to the enantiospecific reactions shown above, both enzymes also oxidize a wide range of substrate analogs to optically active products (5,6,10,11,25). Many of these compounds have been used as chiral synthons to synthesize a number of compounds of interest to the pharmaceutical and specialty chemical industries (references 9, 10, and 17 and references cited therein).In addition to forming arene cis-dihydrodiols, the TDO expressed by strain F39/D catalyzes the monohydroxylation of indan to (1R)-indanol (7, 29). The same reaction is catalyzed by TDO expressed by P. putida UV4, a strain studied extensively by Boyd and his colleagues (5, 6). In contrast, NDO from strain NCIB 9816/11 oxidizes indan to (1S)-indanol and also catalyzes the desaturation of indan to indene. The latter compound is then oxidized by the enzyme to (1S)-indenol and cis-(1R,2S)-indandiol (15). The TDOs expressed by F39/D and UV4 do not catalyze the desaturation of indan to indene (6,29). Strain UV4 does, however, oxidize 1,2-dihydronaphthalene (compound I, Fig. 1) to (ϩ)-(1R)-hydroxy-1,2-dihydronaphthalene (compound II), (ϩ)-cis-(1S,2R)-dihydroxy-1,2,3,4-tetrahydronaphthalene (compo...

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“…The 14 genes encoding the enzymes for this metabolism are organized in two operons: nah (nahA-F), encoding six enzymes required for metabolism of naphthalene to salicylate and pyruvate, and sal (nahG-M), encoding eight enzymes which metabolize salicylate to pyruvate and acetaldehyde (43). Expression of these enzymes is increased over 20-fold by growth in the presence of the inducer salicylate or the nonmetabolizable inducer 2-aminobenzoate (2). Induction requires the product of only one regulatory gene, nahR, encoding a 36-kilodalton (kDa) polypeptide (33,36,44); it mediates induction by activating transcription from both the nah and sal promoters (33).…”

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confidence: 99%

Evidence that the transcription activator encoded by the Pseudomonas putida nahR gene is evolutionarily related to the transcription activators encoded by the Rhizobium nodD genes

Schell

Sukordhaman

1989

J Bacteriol

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The nahR gene of the 83-kilobase naphthalene degradation plasmid NAH7 of Pseudomonas putida encodes a 34-kilodalton polypeptide which binds to the nah and sal promoters to activate transcription of the degradation genes in response to the inducer salicylate. The DNA sequence of the nahR gene was determined, and a derived amino acid sequence of the NahR protein was obtained. A computer search for homologous proteins showed that within the first 124 amino-terminal residues, NahR has approximately 35% identity with the transcriptional activator proteins encoded by the nodD genes of Rhizobium species. Allowing for ultraconservative amino acid substitutions, greater than 47% overall similarity was found between NahR and NodD, while 32% similarity was found between NahR and another transcription activator, LysR of Escherichia coli. The region of greatest similarity among all three proteins contained a probable helix-turn-helix DNA-binding motif as suggested by homology with the proposed consensus sequence for Cro-like DNA-binding domains. The high level of amino acid identity between NahR and NodD, in conjunction with the observations that nahR and nodD are 45% homologous in DNA sequence, are divergently transcribed from homologous promoters near the structural genes they control, and have similar DNA-binding sites, strongly suggests that these two genes evolved from a common ancestor.The 83-kilobase (kb) NAH7 plasmid from the soil bacterium Pseudomonas putida encodes enzymes for the metabolism of naphthalene or salicylate as the sole carbon and energy source (8). The 14 genes encoding the enzymes for this metabolism are organized in two operons: nah (nahA-F), encoding six enzymes required for metabolism of naphthalene to salicylate and pyruvate, and sal (nahG-M), encoding eight enzymes which metabolize salicylate to pyruvate and acetaldehyde (43). Expression of these enzymes is increased over 20-fold by growth in the presence of the inducer salicylate or the nonmetabolizable inducer 2-aminobenzoate (2). Induction requires the product of only one regulatory gene, nahR, encoding a 36-kilodalton (kDa) polypeptide (33,36,44); it mediates induction by activating transcription from both the nah and sal promoters (33). NahR activates transcription from the sal promoter in trans, only in the presence of salicylate. Deletion experiments have shown that the sal promoter sequences between -83 and -45 are required for both DNA binding and transcription activation by NahR (35,36). Other experiments have shown that the NahR protein protects a highly conserved region (-82 to -47) of both the nah and sal promoters from DNase I digestion in the presence or absence of salicylate and that this binding is necessary for transcription activation (35). It has been suggested that the promoter-bound NahR protein, upon binding salicylate, undergoes a conformational change which results in increased transcription from that promoter (35).An analogous system is found in another genus of soil bacteria, Rhizobium. Members of this genus harbor larg...

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The Induction of the Enzymes of Naphthalene Metabolism in Pseudomonads by Salicylate and 2-Aminobenzoate

Cited by 65 publications

References 10 publications

Desaturation, dioxygenation, and monooxygenation reactions catalyzed by naphthalene dioxygenase from Pseudomonas sp. strain 9816-4

Desaturation, dioxygenation, and monooxygenation reactions catalyzed by naphthalene dioxygenase from Pseudomonas sp. strain 9816-4

Desaturation and oxygenation of 1,2-dihydronaphthalene by toluene and naphthalene dioxygenase

Evidence that the transcription activator encoded by the Pseudomonas putida nahR gene is evolutionarily related to the transcription activators encoded by the Rhizobium nodD genes

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