Molecular detection and diversity of novel diterpenoid dioxygenase DitA1 genes from proteobacterial strains and soil samples

Witzig, Robert; Aly, Hamdy A. H.; Strömpl, Carsten; Wray, Victor; Junca, Howard; Pieper, Dietmar H.

doi:10.1111/j.1462-2920.2007.01242.x

Cited by 14 publications

(14 citation statements)

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“…During exponential growth, 25 ml of each culture was mixed with the same volume of RNAprotect Bacteria reagent (Qiagen) and incubated for 5 min at room temperature, and cells were harvested by centrifugation. Harvested cells were resuspended in 100 ml water and immediately processed as previously described (Witzig et al, 2007). cDNA was synthesized from 10 mg total RNA using SuperScript III reverse transcriptase (Invitrogen) according to the procedure from the manufacturer, followed by purification of cDNA using a Qiaquick PCR purification kit (Qiagen).…”

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Degradation of chloroaromatics by Pseudomonas putida GJ31: assembled route for chlorobenzene degradation encoded by clusters on plasmid pKW1 and the chromosome

et al. 2009

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Pseudomonas putida GJ31 has been reported to grow on chlorobenzene using a meta-cleavage pathway with chlorocatechol 2,3-dioxygenase (CbzE) as a key enzyme. The CbzE-encoding gene was found to be localized on the 180 kb plasmid pKW1 in a cbzTEXGS cluster, which is flanked by transposases and encodes only a partial (chloro)catechol meta-cleavage pathway comprising ferredoxin reductase, chlorocatechol 2,3-dioxygenase, an unknown protein, 2-hydroxymuconic semialdehyde dehydrogenase and glutathione S-transferase. Downstream of cbzTEXGS are located cbzJ, encoding a novel type of 2-hydroxypent-2,4-dienoate hydratase, and a transposon region highly similar to Tn5501. Upstream of cbzTEXGS, traNEOFG transfer genes were found. The search for gene clusters possibly completing the (chloro)catechol metabolic pathway of GJ31 revealed the presence of two additional catabolic gene clusters on pKW1. The mhpRBCDFETP cluster encodes enzymes for the dissimilation of 2,3-dihydroxyphenylpropionate in a novel arrangement characterized by the absence of a gene encoding 3-(3-hydroxyphenyl)propionate monooxygenase and the presence of a GntR-type regulator, whereas the nahINLOMKJ cluster encodes part of the naphthalene metabolic pathway. Transcription studies supported their possible involvement in chlorobenzene degradation. The upper pathway cluster, comprising genes encoding a chlorobenzene dioxygenase and a chlorobenzene dihydrodiol dehydrogenase, was localized on the chromosome. A high level of transcription in response to chlorobenzene revealed it to be crucial for chlorobenzene degradation. The chlorobenzene degradation pathway in strain GJ31 is thus a mosaic encoded by four gene clusters.

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Degradation of chloroaromatics by Pseudomonas putida GJ31: assembled route for chlorobenzene degradation encoded by clusters on plasmid pKW1 and the chromosome

et al. 2009

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“…Product accumulation was quantified using 60% (vol/vol) methanol. HPLC-mass spectrometry (HPLC-MS) was performed as previously described (59). The one-dimensional 1 H nuclear magnetic resonance (NMR) spectra were recorded as previously described (59).…”

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“…HPLC-mass spectrometry (HPLC-MS) was performed as previously described (59). The one-dimensional 1 H nuclear magnetic resonance (NMR) spectra were recorded as previously described (59). For respective transformation experiments, the substrates were added from a 100 mM stock solution in deuterated DMSO in order to avoid interference from protonated DMSO-derived signals.…”

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“…In the case of DBF-grown cells, residual crystals of DBF were removed by filtration (595 1/2 filter paper; Schleicher & Schuell) before harvesting. Harvested cells were resuspended in 100 l water and immediately processed as previously described (59). The RNA was subsequently purified using the RNeasy kit (Qiagen), and 2 l of the eluted RNA (60 l) was separated in 1% agarose gels and stained with ethidium bromide.…”

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Two Angular Dioxygenases Contribute to the Metabolic Versatility of Dibenzofuran-Degrading Rhodococcus sp. Strain HA01

Aly¹,

Huu²,

Wray

et al. 2008

Appl Environ Microbiol

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Rhodococcus sp. strain HA01, isolated through its ability to utilize dibenzofuran (DBF) as the sole carbon and energy source, was also capable, albeit with low activity, of transforming dibenzo-p-dioxin (DD). This strain could also transform 3-chlorodibenzofuran (3CDBF), mainly by angular oxygenation at the ether bond-carrying carbon (the angular position) and an adjacent carbon atom, to 4-chlorosalicylate as the end product. Similarly, 2-chlorodibenzofuran (2CDBF) was transformed to 5-chlorosalicylate. However, lateral oxygenation at the 3,4-positions was also observed and yielded the novel product 2-chloro-3,4-dihydro-3,4-dihydroxydibenzofuran. Two gene clusters encoding enzymes for angular oxygenation (dfdA1A2A3A4 and dbfA1A2) were isolated, and expression of both was observed during growth on DBF. Heterologous expression revealed that both oxygenase systems catalyze angular oxygenation of DBF and DD but exhibited complementary substrate specificity with respect to CDBF transformation. While DfdA1A2A3A4 oxygenase, with high similarity to DfdA1A2A3A4 oxygenase from Terrabacter sp. strain YK3, transforms 3CDBF by angular dioxygenation at a rate of 29% ؎ 4% that of DBF, 2CDBF was not transformed. In contrast, DbfA1A2 oxygenase, with high similarity to the DbfA1A2 oxygenase from Terrabacter sp. strain DBF63, exhibited complementary activity with angular oxygenase activity against 2CDBF but negligible activity against 3CDBF. Thus, Rhodococcus sp. strain HA01 constitutes the first described example of a bacterial strain where coexpression of two angular dioxygenases was observed. Such complementary activity allows for the efficient transformation of chlorinated DBFs.Biarylethers, comprising dibenzo-p-dioxin (DD), dibenzofuran (DBF), diphenyl ether, and their halogenated derivatives, are widespread environmental pollutants. Polychlorinated DDs and DBFs, the contaminating by-products formed during the manufacture of pesticides or the incineration of industrial and domestic wastes, can cause a wide range of serious health effects (1,4,13).Microorganisms play important roles in the degradation and mineralization of xenobiotic and aromatic compounds in natural environments. Their aerobic degradation is frequently initiated by Rieske nonheme iron oxygenases which catalyze the incorporation of two oxygen atoms into the aromatic ring to form arene cis-diols (15). This is then followed by a dehydrogenation reaction catalyzed by a cis-dihydrodiol dehydrogenase to give catechol or substituted catechols, which serve as substrates for oxygenolytic aromatic ring cleavage. Rieske nonheme iron oxygenases are typically composed of a terminal oxygenase (iron-sulfur protein) and different electron transport proteins (8). The catalytic iron-sulfur proteins are homoor heteromultimers with the ␣-subunit containing a Riesketype [2Fe-2S] cluster, a mononuclear nonheme iron oxygen activation center, and a substrate-binding site (8) that is responsible for substrate specificity (15). Comparison of the amino acid sequences of the ␣-subunits ...

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“…Because of their abundance in wood and their toxicity to fish, resin acids are important pollutants resulting from pulp and paper production (1). A wide range of bacteria are known to grow on abietane resin acids (26,42), and recent studies have provided evidence for the involvement of P450s in bacterial catabolism of these compounds. Knockout mutagenesis in Pseudomonas diterpeniphila A19-6a indicated that the P450 TdtD (CYP226A3) is involved in, but is not essential for, catabolism of dehydroabietic acid (DhA) and abietic acid (AbA) (27).…”

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Distinct Roles for Two CYP226 Family Cytochromes P450 in Abietane Diterpenoid Catabolism by Burkholderia xenovorans LB400

et al. 2008

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The 80-kb dit cluster of Burkholderia xenovorans LB400 encodes the catabolism of abietane diterpenoids. This cluster includes ditQ and ditU, predicted to encode cytochromes P450 (P450s) belonging to the poorly characterized CYP226A subfamily. Using proteomics, we identified 16 dit-encoded proteins that were significantly more abundant in LB400 cells grown on dehydroabietic acid (DhA) or abietic acid (AbA) than in succinategrown cells. A key difference in the catabolism of DhA and AbA lies in the differential expression of the P450s; DitU was detected only in the AbA-grown cells, whereas DitQ was expressed both during growth on DhA and during growth on AbA. Analyses of insertion mutants showed that ditQ was required for growth on DhA, ditU was required for growth on AbA, and neither gene was required for growth on the central intermediate

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Molecular detection and diversity of novel diterpenoid dioxygenase DitA1 genes from proteobacterial strains and soil samples

Cited by 14 publications

References 68 publications

Degradation of chloroaromatics by Pseudomonas putida GJ31: assembled route for chlorobenzene degradation encoded by clusters on plasmid pKW1 and the chromosome

Degradation of chloroaromatics by Pseudomonas putida GJ31: assembled route for chlorobenzene degradation encoded by clusters on plasmid pKW1 and the chromosome

Two Angular Dioxygenases Contribute to the Metabolic Versatility of Dibenzofuran-Degrading Rhodococcus sp. Strain HA01

Distinct Roles for Two CYP226 Family Cytochromes P450 in Abietane Diterpenoid Catabolism by Burkholderia xenovorans LB400

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