Degradation of 1,4-dioxane and cyclic ethers by an isolated fungus.
ABSTRACT: By using 1,4-dioxane as the sole source of carbon, a 1,4-dioxane-degrading microorganism was isolated from soil. The fungus, termed strain A, was able to utilize 1,4-dioxane and many kinds of cyclic ethers as the sole source of carbon and was identified as Cordyceps sinensis from its 18S rRNA gene sequence. Ethylene glycol was identified as a degradation product of 1,4-dioxane by the use of deuterated 1,4-dioxane-d8 and gas chromatography-mass spectrometry analysis. A degradation pathway involving ethylene glycol, glycolic acid, and oxalic acid was proposed, followed by incorporation of the glycolic acid and/or oxalic acid via glyoxylic acid into the tricarboxylic acid cycle.
Project description:A new simple and reliable spectrophotometric method is described to determine glyoxylic acid in its synthesis reaction mixture containing oxalic acid, glycolic acid, acetic acid, glyoxal, and ethylene glycol by means of a modified Hopkins-Cole reaction between glyoxylic acid and tryptophan in presence of ferric chloride and concentrated sulphuric acid. The linear range of glyoxylic acid concentration is 0-0.028?M. The limits of detection (LOD) and quantitation (LOQ) are 0.0019?M and 0.00577?M, respectively. The LOD, LOQ, standard deviation, relative standard deviation, and recovery ratio of the proposed method are comparable with a selected HPLC reference method. Both methods displayed same precision and credibility. Reaction stoichiometry between tryptophan and glyoxylic acid is assumed to be 2?:?3. Reaction mechanism has been postulated based on identified molar ratios of reactants. Glyoxal gave a negative test with tryptophan although it is a dialdehyde.
Project description:In this study, we investigated the metabolism of ethylene glycol in the Pseudomonas putida strains KT2440 and JM37 by employing growth and bioconversion experiments, directed mutagenesis, and proteome analysis. We found that strain JM37 grew rapidly with ethylene glycol as a sole source of carbon and energy, while strain KT2440 did not grow within 2 days of incubation under the same conditions. However, bioconversion experiments revealed metabolism of ethylene glycol by both strains, with the temporal accumulation of glycolic acid and glyoxylic acid for strain KT2440. This accumulation was further increased by targeted mutagenesis. The key enzymes and specific differences between the two strains were identified by comparative proteomics. In P. putida JM37, tartronate semialdehyde synthase (Gcl), malate synthase (GlcB), and isocitrate lyase (AceA) were found to be induced in the presence of ethylene glycol or glyoxylic acid. Under the same conditions, strain KT2440 showed induction of AceA only. Despite this difference, the two strains were found to use similar periplasmic dehydrogenases for the initial oxidation step of ethylene glycol, namely, the two redundant pyrroloquinoline quinone (PQQ)-dependent enzymes PedE and PedH. From these results we constructed a new pathway for the metabolism of ethylene glycol in P. putida. Furthermore, we conclude that Pseudomonas putida might serve as a useful platform from which to establish a whole-cell biocatalyst for the production of glyoxylic acid from ethylene glycol.
Project description:(DAF)Pd(OAc)2 (DAF = 4,5-diazafluorenone) catalyzes aerobic intramolecular aryl C-H amination with N-benzenesulfonyl-2-aminobiphenyl in dioxane to afford the corresponding carbazole product. Mechanistic studies show that the reaction involves in situ generation of peroxide species from 1,4-dioxane and O2, and the reaction further benefits from the presence of glycolic acid, an oxidative decomposition product of dioxane. An induction period observed for the formation of the carbazole product correlates with the formation of 1,4-dioxan-2-hydroperoxide via autoxidation of 1,4-dioxane, and the in situ-generated peroxide is proposed to serve as the reactive oxidant in the reaction. These findings have important implications for the palladium-catalyzed aerobic oxidation reactions conducted in ethereal solvents.
Project description:The groundwater contaminant 1,4-dioxane (dioxane) is transformed by several monooxygenase-expressing microorganisms, but only a few of these, including Pseudonocardia dioxanivorans strain CB1190, can metabolize the compound as a sole carbon and energy source. However, nothing is yet known about the genetic basis of dioxane metabolism. In this study, we used a microarray to study differential expression of genes in strain CB1190 grown on dioxane, glycolate (a previously identified intermediate of dioxane degradation), or pyruvate. Of eight multicomponent monooxygenase gene clusters carried by the strain CB1190 genome, only the monooxygenase gene cluster located on plasmid pPSED02 was upregulated with dioxane relative to pyruvate. Plasmid-borne genes for putative aldehyde dehydrogenases, an aldehyde reductase, and an alcohol oxidoreductase were also induced during growth with dioxane. With both dioxane and glycolate, a chromosomal gene cluster encoding a putative glycolate oxidase was upregulated, as were chromosomal genes related to glyoxylate metabolism through the glyoxylate carboligase pathway. Glyoxylate carboligase activity in cell extracts from cells pregrown with dioxane and in Rhodococcus jostii strain RHA1 cells expressing the putative strain CB1190 glyoxylate carboligase gene further demonstrated the role of glyoxylate metabolism in the degradation of dioxane. Finally, we used (13)C-labeled dioxane amino acid isotopomer analysis to provide additional evidence that metabolites of dioxane enter central metabolism as three-carbon compounds, likely as phosphoglycerate. The routing of dioxane metabolites via the glyoxylate carboligase pathway helps to explain how dioxane is metabolized as a sole carbon and energy source for strain CB1190.
Project description:The bacterium Pseudonocardia dioxanivorans CB1190 grows on the cyclic ethers 1,4-dioxane (dioxane) and tetrahydrofuran (THF) as sole carbon and energy sources. Prior transcriptional studies indicated that an annotated THF monooxygenase (THF MO) gene cluster, thmADBC, located on a plasmid in CB1190 is upregulated during growth on dioxane. In this work, transcriptional analysis demonstrates that upregulation of thmADBC occurs during growth on the dioxane metabolite β-hydroxyethoxyacetic acid (HEAA) and on THF. Comparison of the transcriptomes of CB1190 grown on THF and succinate (an intermediate of THF degradation) permitted the identification of other genes involved in THF metabolism. Dioxane and THF oxidation activity of the THF MO was verified in Rhodococcus jostii RHA1 cells heterologously expressing the CB1190 thmADBC gene cluster. Interestingly, these thmADBC expression clones accumulated HEAA as a dead-end product of dioxane transformation, indicating that despite its genes being transcriptionally upregulated during growth on HEAA, the THF MO enzyme is not responsible for degradation of HEAA in CB1190. Similar activities were also observed in RHA1 cells heterologously expressing the thmADBC gene cluster from Pseudonocardia tetrahydrofuranoxydans K1.
Project description:Tetrahydrofuran (THF) is known to induce the biodegradation of 1,4-dioxane (dioxane), an emerging contaminant, but the mechanisms by which THF affects dioxane biodegradation in microbial communities are not well understood. To fill this knowledge gap, changes in the microbial community structure in microcosm experiments with synthetic medium and landfill leachate were examined over time using 16S rRNA gene amplicon sequencing and functional gene quantitative PCR assays. The overarching hypothesis being tested was that THF promoted dioxane biodegradation by increasing the abundance of dioxane-degrading bacteria in the consortium. The data revealed that in experiments with synthetic medium, the addition of THF significantly increased the abundance of Pseudonocardia, a genus with several representatives that can grow on both dioxane and THF, and of Rhodococ cus ruber, a species that can use THF as the primary growth substrate while cometabolizing dioxane. However, in similar experiments with landfill leachate, only R. ruber was significantly enriched. When the THF concentration was higher than the dioxane concentration, THF competitively inhibited dioxane degradation since dioxane degradation was negligible, while the dioxane-degrading bacteria and the corresponding THF/dioxane monooxygenase gene copies increased by a few orders of magnitude.IMPORTANCE Widespread in groundwater and carcinogenic to humans, 1,4-dioxane (dioxane) is attracting significant attention in recent years. Advanced oxidation processes can effectively remove dioxane but require high energy consumption and operation costs. Biological removal of dioxane is of particular interest due to the ability of some bacteria to mineralize dioxane at a low energy cost. Although dioxane is generally considered recalcitrant to biodegradation, more than 20 types of bacteria can degrade dioxane as the sole electron donor substrate or the secondary electron donor substrate. In the latter case, tetrahydrofuran (THF) is commonly studied as the primary electron donor substrate. Previous work has shown that THF promotes dioxane degradation at a low THF concentration but inhibits dioxane degradation at a high THF concentration. Our work expanded on the previous work by mechanically examining the effects of THF on dioxane degradation in a microbial community context.
Project description:Bacterial multicomponent monooxygenase gene targets in Pseudonocardia dioxanivorans CB1190 were evaluated for their use as biomarkers to identify the potential for 1,4-dioxane biodegradation in pure cultures and environmental samples. Our studies using laboratory pure cultures and industrial activated sludge samples suggest that the presence of genes associated with dioxane monooxygenase, propane monooxygenase, alcohol dehydrogenase, and aldehyde dehydrogenase are promising indicators of 1,4-dioxane biotransformation; however, gene abundance was insufficient to predict actual biodegradation. A time course gene expression analysis of dioxane and propane monooxygenases in Pseudonocardia dioxanivorans CB1190 and mixed communities in wastewater samples revealed important associations with the rates of 1,4-dioxane removal. In addition, transcripts of alcohol dehydrogenase and aldehyde dehydrogenase genes were upregulated during biodegradation, although only the aldehyde dehydrogenase was significantly correlated with 1,4-dioxane concentrations. Expression of the propane monooxygenase demonstrated a time-dependent relationship with 1,4-dioxane biodegradation in P. dioxanivorans CB1190, with increased expression occurring after over 50% of the 1,4-dioxane had been removed. While the fraction of P. dioxanivorans CB1190-like bacteria among the total bacterial population significantly increased with decrease in 1,4-dioxane concentrations in wastewater treatment samples undergoing active biodegradation, the abundance and expression of monooxygenase-based biomarkers were better predictors of 1,4-dioxane degradation than taxonomic 16S rRNA genes. This study illustrates that specific bacterial monooxygenase and dehydrogenase gene targets together can serve as effective biomarkers for 1,4-dioxane biodegradation in the environment.
Project description:We report here the complete genome sequence of Mycobacterium dioxanotrophicus PH-06, which is capable of using 1,4-dioxane as a sole source of carbon and energy. The reported sequence will enable the elucidation of this novel metabolic pathway and the development of molecular biomarkers to assess bioremediation potential at contaminated sites.
Project description:The asymmetric unit of the title compound,, C(4)H(4)N(2)O(5)·0.5C(4)H(8)O(2), contains one molecule of 5,5-dihydroxybarbituric acid with a nearly planar barbiturate ring and half a molecule of 1,4-dioxane. The geometry of the centrosymmetric dioxane molecule is close to an ideal chair conformation. The crystal structure exhibits a complex three-dimensional hydrogen-bonded network. Barbiturate mol-ecules are connected to one another via N-H?O=C, O-H?O=C and N-H?O(hydr-oxy) inter-actions, while the barbituric acid mol-ecule is linked to dioxane by an O-H?O contact.
Project description:THE ASYMMETRIC UNIT OF THE TITLE COMPOUND [SYSTEMATIC NAME: 5-(1-bromo-prop-2-en-1-yl)-5-sec-butyl-pyrimidine-2,4,6-trione 1,4-dioxane hemisolvate], C(11)H(15)BrN(2)O(3)·0.5C(4)H(8)O(2), contains one half-mol-ecule of 1,4-dioxane and one mol-ecule of butallyl-onal, with an almost planar barbiturate ring [largest deviation from the mean plane = 0.049?(5)?Å]. The centrosymmetric dioxane mol-ecule adopts a nearly ideal chair conformation. The barbiturate mol-ecules are linked together by an N-H?O hydrogen bond, giving a single-stranded chain. Additionally, each dioxane mol-ecule acts as a bridge between two anti-parallel strands of hydrogen-bonded barbiturate mol-ecules via two hydrogen bonds, N-H?O(dioxane)O?H-N. Thus, a ladder structure is obtained, with the connected barbiturate mol-ecules forming the 'stiles' and the bridging dioxane mol-ecules the 'rungs'.