Sequence of the gene (pheA) encoding phenol monooxygenase from Pseudomonas sp. EST1001: expression in Escherichia coli and Pseudomonas putida

Nurk, Allan; Kasak, Lagle; Kivisaar, Maia

doi:10.1016/0378-1119(91)90531-f

Cited by 98 publications

(70 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Washing stringency was as follows: two rinses of 10 and 20 min duration in 2 x SSC/O.l YO SDS at room temperature, two washes of 15 min in 1 x SSC/O.l YO SDS at 60 "C, followed by two 10 min washes in 0-25 x SSC/O.l YO SDS at 60 "C. Filters were stripped for re-probing by boiling for 3 min in 0.5% SDS, followed by cooling to room temperature in the SDS solution. Nurk et al (1991) This study * PCR fragments were generated using the following primers : PCRl primers 5'-CGCCATATGCCGATCA-AGTACAAG-3' and 5'-GCTGAAAACTGCAGCAAC AGC-3' ; PCR2 primers 5'-CGCCAT ATGCCGATTGC-TCAGCT-3' and 5'-CAAACTGTTAACGAACACTCTC-3' ; PCR3 primers 5'-ATGCACGAATCGACCATT-TC-3' and 5'-TACTTGGCGACGGTGCGACG-3'. f The primers used were designed on the basis of conserved regions found upon sequence comparisons of rpoN genes which encode d4 factors (Inouye et ul., 1989;Kullik et al, 1991).…”

Section: Methodsmentioning

confidence: 99%

“…As a general rule, mono-hydroxylated ring structures such as phenol are oxygenated by single component flavoprotein monooxygenases (Ballou, 1982). For example, genes encoding flavoprotein hydroxylases (monooxygenases) induced by growth on phenol have recently been identified and characterized in a number of micro-organisms, including Pseudomonas pickettii (Kukor & Olsen, 1992), Pseudomonas EST1001 (Nurk et al, 1991) and the yeast Trichosporon cutaneum (Kalin et al, 1992). The 2,4-dichlorophenol hydroxylases of Acinetobacter and Alcaligenes species have also been shown to be flavoproteins, although these enzymes show no activity with unsubstituted phenol (Beadle & Smith, 1982;Liu & Chapman, 1984).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Conservation of regulatory and structural genes for a multi-component phenol hydroxylase within phenol-catabolizing bacteria that utilize a meta-cleavage pathway

Nordlund

Powlowski

Hagström³

et al. 1993

Journal of General Microbiology

View full text Add to dashboard Cite

Pseudomonas sp. strain CF600 can degrade phenol and some of its methylated derivatives via a plasmid (pVIlSO)-encoded pathway. The metabolic route involves hydroxylation by a multi-component phenol hydroxylase and a subsequent meta-cleavage pathway. All 15 structural genes involved are clustered in an operon that is regulated by a divergently transcribed transcriptional activator. The multi-component nature of the phenol hydroxylase is unusual since reactions of this type are usually accomplished by single component flavoproteins. We have isolated and analysed a number of marine bacterial isolates capable of degrading phenol and a range of other aromatic compounds as sole carbon and energy sources. Southern hybridization and enzyme assays were used to compare the catabolic pathways of these strains and of the archetypal phenol-degrader Pseudomonas U, with respect to known catabolic genes encoded by Pseudomonas CF600. All the strains tested that degraded phenol via a meta-cleavage pathway were found to have DNA highly homologous to each of the components of the multicomponent phenol hydroxylase. Moreover, DNA of the same strains also strongly hybridized to probes specific for pVI150-encoded meta-pathway genes and the specific regulator of its catabolic operon. These results demonstrate conservation of structural and regulatory genes involved in aromatic catabolism within strains isolated from diverse geographical locations (UK, Norway and USA) and a range of habitats that include activated sludge, sea water and fresh-water mud.

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Conservation of regulatory and structural genes for a multi-component phenol hydroxylase within phenol-catabolizing bacteria that utilize a meta-cleavage pathway

Nordlund

Powlowski

Hagström³

et al. 1993

Journal of General Microbiology

View full text Add to dashboard Cite

show abstract

“…I, might not be a simple flavoprotein. Plasmid-encoded 2,4-in 0.05 M sodium phosphate (pH 7.6) at a protein concendichlorophenol hydroxylase purified fromAlcaligenes eutro-2.9 mg/ml; II, as in spectrum I but containing 8.7 mg of phus(pJP4) (28) (37), PheA (34), TodA (49), and PobA (9). (B) Comparison of a portion of the deduced sequence of ThuD with sequences of other flavoprotein hydroxylases.…”

Section: Methodsmentioning

confidence: 99%

“…Symbols: *, amino acid residues conserved among all of the sequences shown; j, amino acid residues conserved among the four phenol hydroxylases (TfdB, PheA, PhyA, and ThuD); :, residue identical with that found in TbuD; ., conservative change from the TbuD residue. Dashes (18), xylS (19), and piL4 (20 [38] VNAND NVVIVGTGLAGVEVAFGL RASGWEGNI IRLVGD I 1 [37] AT HVAIIGNGVGGFTTANALIRAEGFEGRI ISLIGD I [34] MKT QVAIIGAGPSGLLTGQLL I BKAGID NVILE I 1 [32] …”

Section: Methodsmentioning

confidence: 99%

Complete nucleotide sequence of tbuD, the gene encoding phenol/cresol hydroxylase from Pseudomonas pickettii PKO1, and functional analysis of the encoded enzyme

Kukor

Olsen

1992

J Bacteriol

View full text Add to dashboard Cite

The gene (tbuD) encoding phenol hydroxylase, the enzyme that converts cresols or phenol to the corresponding catechols, has been cloned from Pseudomonaspickettii PKO1 as a 26.5-kbp BamHI-cleaved DNA fragment, designated pRO1957, which allowed the heterogenetic recipient Pseudomonas aeruginosa PA01c to grow on phenol as the sole source of carbon. Two subclones of pRO1957 carried in trans have shown phenol hydroxylase activity in cell extracts ofP. aeruginosa. The nucleotide sequence was determined for one of these subclones, a 3.1-kbp Hindm fragment, and an open reading frame that would encode a peptide of 73 kDa was found. The size of this deduced peptide is consistent with the size of a novel peptide that had been detected in extracts of phenol-induced cells of P. aeruginosa carrying pRO1959, a partial HindI deletion subclone of pRO1957. Phenol hydroxylase purified from phenol-plus-Casamino Acid-grown cells ofP. aeruginosa carrying pRO1959 has an absorbance spectrum characteristic of a simple flavoprotein; moreover, the enzyme exhibits a broad substrate range, accommodating phenol and the three isomers of cresol equally well. Sequence comparisons revealed little overall homology with other flavoprotein hydroxylases, supporting the novelty of this enzyme, although three conserved domains were apparent.Soil bacteria, including members of the genus Pseudomonas, are able to degrade a wide variety of aromatic hydrocarbons. In oxygenated environments, this degradation proceeds via enzymatic mono-or dihydroxylation of the aromatic nucleus, leading to ortho-dihydroxy-substituted products (catechols) which are the substrates for ring cleavage and subsequent entry into central metabolism.We have previously reported (11) on the isolation and characterization of Pseudomonas pickettii PKO1, a strain that metabolizes benzene and toluene via phenolic intermediates (phenol and m-cresol, respectively) that are further hydroxylated to catechol or methylcatechols prior to metaring cleavage. The genetic material carrying this toluene-and benzene-degradative capability was cloned from the chromosome of P. pickettii PKO1 as a 26.5-kbp DNA fragment designated pRO1957. The individual genes encoding these degradative enzymes have been mapped, and the genes have been expressed in Pseudomonas aeruginosa PAO1 (24).The phenol/cresol hydroxylase-encoding structural gene, tbuD, was subcloned from pRO1957 as a 3.1-kbp HindIII fragment (25). We report here the nucleotide sequence of a 2,019-bp region of the 3.1-kbp DNA fragment that carries tbuD. The peptide encoded by tbuD was purified and shown to be a flavoprotein capable of hydroxylating phenol as well as a broad range of substituted phenols. MATERIALS AND METHODSBacterial strains and culture conditions. P. aeruginosa PAO1 (15) and Escherichia coli DH5a (13), used for construction and maintenance of plasmids, were cultured at 37°C on plate count complex medium (36) or Luria-Bertani medium (42), respectively. Plasmids were introduced into E.coli by the procedure of Hanahan (13) by the proc...

show abstract

“…The aromatic hydroxylases are designated as UbiH, 2-octaprenyl-6-methoxyphenol hydroxylase from E. coli (GenBank TM accession number P25534) (43); PobARh, 4-hydroxybenzoate hydroxylase (PobA) from Rhizobium leguminosarum (GenBank TM accession number AAA73519) (58); PhyATc, phenol hydroxylase from Trichosporon cutaneum (GenBank TM accession number AAA34202) (59); ShPs, salicylate hydroxylase from Pseudomonas putida (GenBank TM accession number d1011754) (60); PcpB, pentachlorophenol 4-monooxygenase (pcpB) from Flavobacterium sp. (GenBank TM accession number AAF15368) (61); PobAPS, p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens (GenBank TM accession number WHPSBF) (62); PobAAc, p-hydroxybenzoate hydroxylase (pobA) from Acinetobacter calcoaceticus (GenBank TM accession number AAC37163) (63); DnrF, 11-aklavinone hydroxylase from Streptomyces peucetius (GenBank TM accession number AAC43342) (64); and PheA, phenol monooxygenase from Pseudomonas sp (GenBank TM accession number AAC64901) (65).…”

Section: Coq6 Is a Flavin-dependent Monooxygenase In Q Biosynthesismentioning

confidence: 99%

The Saccharomyces cerevisiae COQ6 Gene Encodes a Mitochondrial Flavin-dependent Monooxygenase Required for Coenzyme Q Biosynthesis

Gin

Hsu

Rothman

et al. 2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

Coenzyme Q (Q) is a lipid that functions as an electron carrier in the mitochondrial respiratory chain in eukaryotes. There are eight complementation groups of Q-deficient Saccharomyces cerevisiae mutants, designated coq1-coq8. Here we have isolated the COQ6 gene by functional complementation and, in contrast to a previous report, find it is not an essential gene. coq6 mutants are unable to grow on nonfermentable carbon sources and do not synthesize Q but instead accumulate the Q biosynthetic intermediate 3-hexaprenyl-4-hydroxybenzoic acid. The Coq6 polypeptide is imported into the mitochondria in a membrane potential-dependent manner. Coq6p is a peripheral membrane protein that localizes to the matrix side of the inner mitochondrial membrane. Based on sequence homology to known proteins, we suggest that COQ6 encodes a flavindependent monooxygenase required for one or more steps in Q biosynthesis.

show abstract

Sequence of the gene (pheA) encoding phenol monooxygenase from Pseudomonas sp. EST1001: expression in Escherichia coli and Pseudomonas putida

Cited by 98 publications

References 19 publications

Conservation of regulatory and structural genes for a multi-component phenol hydroxylase within phenol-catabolizing bacteria that utilize a meta-cleavage pathway

Conservation of regulatory and structural genes for a multi-component phenol hydroxylase within phenol-catabolizing bacteria that utilize a meta-cleavage pathway

Complete nucleotide sequence of tbuD, the gene encoding phenol/cresol hydroxylase from Pseudomonas pickettii PKO1, and functional analysis of the encoded enzyme

The Saccharomyces cerevisiae COQ6 Gene Encodes a Mitochondrial Flavin-dependent Monooxygenase Required for Coenzyme Q Biosynthesis

Contact Info

Product

Resources

About