Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

Franklin, F. Christopher H.; Bagdasarian, Michael; Timmis, Kenneth N.

doi:10.1073/pnas.78.12.7458

Cited by 502 publications

(295 citation statements)

References 17 publications

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“…P. putida F1 is reported to possess a toluene dioxygenase but not PAH dioxygenase (Zylstra & Gibson, 1989). Escherichia coli SURE (Promega), lacking any PAH degradation ability, and P. putida KT2440 ( = DSM 6125), described as a strain lacking C230 (Franklin et al, 1981), were used as negative controls for amplification of gene fragments of the initial dioxygenase and C230, respectively. Further strains described as PAH degraders were also examined : Pseudomonas stutzeri AN11 (kindly provided by R. Rossello-Mora; Max Planck Institute for Marine Microbiology, Bremen, Germany) ; Sphingomonas yanoikuyae DSM 6900 (Gibson et al, 1973) and Sphingomonas paucimobilis BA2 (Kastner et al, 1994).…”

Section: Methodsmentioning

confidence: 99%

Differential detection of key enzymes of polyaromatic-hydrocarbon-degrading bacteria using PCR and gene probes

Moser²,

et al. 1999

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Bacteria with ability to degrade polyaromatic hydrocarbons (PAHs), isolated from wastewater and soil samples, were investigated for their taxonomic, physiological and genetic diversity. Eighteen isolates able to metabolize naphthalene or phenanthrene as sole carbon source were taxonomically affiliated to different subclasses of the Proteobacteria (Sphingomonas spp., Acidovorax spp., Comamonas spp. and Pseudomonas spp.) and to phyla of Gram-positive bacteria with low and high DNA G+C content (Paenibacillus sp. and Rhodococcus spp., respectively). Representatives of the genera Pseudomonas and Sphingomonas formed a remarkably high fraction of these isolates; 9 out of 18 strains belonged to these groups. Tests for enzyme activities showed that the majority of the isolates growing with PAHs as sole sources of carbon and energy had an active catechol2,3-dioxygenase (C230). C230 specific activities were very diverse, ranging from 0-1 to 650 mU (mg protein)-'. Pseudomonas and Sphingomonas strains showed considerably higher activities than the other isolates. All PAH degraders were examined for the presence of an initial PAH dioxygenase and C230, which catalyse key steps of PAH degradation, by PCR amplification of gene fragments and subsequent hybridization. PCR primers and internal oligonucleotide probes were developed for the specific detection of the genes of Pseudomonas and Sphingomonas strains. (1996). The aim of the present investigation was to isolate bacterial PAH degraders from aquatic a n d terrestrial sites, to determine their taxonomic affiliation and to characterize them physiologically a n d genetically with respect to their ability to degrade PAHs. al., 1996) are specific for most genuine pseudornonads, distinct P. putida strains, and the genera Comamonas and Acidovorax, respectively. Keywords METHODSFurther examination of the isolates was done by physiological tests as reported previously (Kampfer et al., 1991). Fatty acid methyl ester (FAME) patterns were used as a further marker for identification. Preparation of FAME extracts, gas chromatographic analysis and numerical identification by the MIDI system (Sherlock 2.11) were performed as described previously (Kampfer et al., 1996).Determination of naphthalene and phenanthrene mineralization potential. The potential of the isolated bacteria for mineralization of PAHs was examined by gas chromatographically quantifying remaining PAH and sirnultaneously monitoring bacterial growth in batch cultures.For each tested strain, five airtight flasks were each filled with 10 rnl basal medium containing either 005 ' / o naphthalene or phenanthrene and were inoculated with approximately 10' cells. Cell number was estimated by measuring the optical density at 600nm, Remaining PAH in the cultures was extracted by vigorously shaking (30 rnin) with 5 rnl n-hexane for gas chromatographic analysis. The gas chromatograph HP 6890 was equipped with a flame-ionization detector and a 25 m capillary column (Ultra 2, cross-linked 5 % phenyl methyl siloxan ; Hewlett Packard) ....

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Section: Methodsmentioning

confidence: 99%

Differential detection of key enzymes of polyaromatic-hydrocarbon-degrading bacteria using PCR and gene probes

Moser²,

et al. 1999

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“…6 Best results were achieved for the IPTG inducible promoter which were more than twice those obtained by constructs where the Ptac/lacIq 18,22 is replaced by a Pm/xylS promoter/regulator gene system. 23 The genome sequence of Pseudomonas putida KT2440 is characterized by a high average GC content (~61.5%). The impact of the scFv coding sequence GCcontent and codon usage on production efficiencies was investigated using synthetic codon adapted sequences designed for anti-CRP and anti-MUC1 scFv genes.…”

Section: Assessment Of a New Scfv Production Hostmentioning

confidence: 99%

Biomedicals from a soil bug

Dammeyer

2012

Bioengineered

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“…putida RKJ1) and Ps. putida KT2442 (Franklin et al 1981), and two indole-producing The mineral salts medium (MSM) used in the present study was same as described earlier (Rani et al 1996). Naphthalene, salicylate and isopropyl-(-D-thiogalactopyranoside (IPTG) were added to the basal medium either alone or in combination, so as to induce the plasmidencoded genes.…”

Section: Bacteria Media and Culture Conditionsmentioning

confidence: 99%

“…putida RKJ1) and Ps. putida KT2442 (Franklin et al 1981), and two indole-producing strains of E. coli, i.e. TB1 (Li et al 1979) and AB1157 (Bachmann 1972) by conjugation using E. coli pRK2013 as the helper strain (Jain et al 1984).…”

Section: Bacteria Media and Culture Conditionsmentioning

confidence: 99%

Indigo production by naphthalene-degrading bacteria

Bhushan

Samanta

Jain

2000

Letters in Applied Microbiology

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A I N . 2000. A wild-type naphthalene-degrading strain Pseudomonas putida RKJ1 and two recombinant strains each of Ps. putida and Escherichia coli carrying the genes for naphthalene degradation on a recombinant plasmid pRKJ3, produced indigo and indirubin pigments from indole. Naphthalene, salicylate and IPTG induced cells of naphthalene-degrading recombinant bacteria produced up to two times higher indigo compared with the uninduced cells. The maximum rates of indigo formation by Ps. putida RKJ1, Ps. putida RKJ5/pRKJ3, Ps. putida KT2442/pRKJ3, E. coli TB1/pRKJ3 and E. coli AB1157/pRKJ3 were 0Á60, 0Á80, 0Á60, 1Á20 and 1Á50 nmol min À1 mg dry biomass À1 , respectively, using indole as the substrate. The apparent K m values of indigo formation by these same bacteria were 0Á22, 0Á15, 0Á10, 0Á21 and 0Á20 mmol l À1 , respectively, again using indole as the substrate. The present study revealed that E. coli AB1157 was the most ef®cient of the hosts tested for the expression of the plasmid encoded genes (pRKJ3) from the wild-type strain Ps. putida RKJ1. In addition, both recombinant E. coli strains were capable of producing indigo directly from nutrient medium. INTRODUCTIONIndigo is one of the oldest textile dyes and was traditionally produced from an extract of the plants of the genus Indigofera. Presently, it is being produced by chemical synthesis and is marketed by many leading companies. Very few reports are available regarding the microbial biosynthesis of indigo. Among the microbial indigo producers, the majority of micro-organisms are aromatic hydrocarbondegrading bacteria such as naphthalene-degrading Pseudomonas putida NDO (Murdock et al. 1993), Ps. putida PpG7 (O'Connor and Hartmans 1998), p-cumate and mand p-toluate-degrading Ps. putida F1 and Ps. putida mt-2, respectively (Eaton and Chapman 1995), styrene-degrading Ps. putida S12 and CA-3 (O'Connor et al. 1997) and toluene-degrading Ps. mendocina KR1 (Yen et al. 1991). The enzyme system responsible for indigo formation generally consists of one or more enzymes, typically monooxygenases, dioxygenases or hydroxylases (O'Connor et al. 1997;Doukyu et al. 1998). The genes encoding these enzymes have been cloned from Pseudomonas and Rhodococcus spp. into Escherichia coli strains (Yen et al. 1991;Hart et al. 1992;Eaton and Chapman 1995) so as to produce indigo directly from either nutrient medium (Ensley et al. 1983) or from glucose (Murdock et al. 1993).Recently, a method has been patented by O'riel and Kim (1998) for producing indigo and indirubin dyes using a recombinant Escherichia coli containing a gene encoding a phenol hydroxylase from Bacillus stearothermophilus. In the present study, we report on the formation and identi®ca-tion of indigo and indirubin pigments by one wild-type and four recombinant bacterial strains. MATERIALS AND METHODS Bacteria, media and culture conditionsOf the ®ve bacteria used in the present study, Ps. putida RKJ1 (Samanta et al. 1998) was a wild-type strain and was capable of degrading naphthalene through salicylate. Four recombi...

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Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

Cited by 502 publications

References 17 publications

Differential detection of key enzymes of polyaromatic-hydrocarbon-degrading bacteria using PCR and gene probes

Differential detection of key enzymes of polyaromatic-hydrocarbon-degrading bacteria using PCR and gene probes

Biomedicals from a soil bug

Indigo production by naphthalene-degrading bacteria

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