Production of epoxides from gaseous alkenes by resting-cell suspensions and immobilized cells of alkene-utilizing bacteria

Habets-Crützen, A. Q. H.; Brink, L.E.S.; Ginkel, C.G. van; Bont, J.A.M. de; Tramper, J.

doi:10.1007/bf00250633

Cited by 87 publications

(34 citation statements)

References 17 publications

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“…The VCand ethene-assimilating bacteria may be useful for the bioremediation of sites contaminated with chlorinated solvents (38). In addition, several ethene-assimilating strains have been evaluated as biocatalysts for the production of epoxides (17,46). Much research has focused on the kinetics of VC and ethene oxidation and on the cometabolism of related substrates (7,24,49), while fundamental questions concerning the catabolic pathways and enzymes involved have been somewhat neglected.…”

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confidence: 99%

Epoxyalkane:Coenzyme M Transferase in the Ethene and Vinyl Chloride Biodegradation Pathways of Mycobacterium Strain JS60

2003

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Mycobacterium strains that grow on ethene and vinyl chloride (VC) are widely distributed in the environment and are potentially useful for biocatalysis and bioremediation. The catabolic pathway of alkene assimilation in mycobacteria is not well characterized. It is clear that the initial step is a monooxygenase-mediated epoxidation that produces epoxyethane from ethene and chlorooxirane from VC, but the enzymes involved in subsequent transformation of the epoxides have not been identified. We investigated epoxyethane metabolism in Mycobacterium strain JS60 and discovered a coenzyme M (CoM)-dependent enzyme activity in extracts from VCand ethene-grown cells. PCR amplifications using primers targeted at epoxyalkane:CoM transferase (EaCoMT) genes yielded part of the JS60 EaCoMT gene, which was used to clone an 8.4-kb genomic DNA fragment. The complete EaCoMT gene (etnE) was recovered, along with genes (etnABCD) encoding a fourcomponent monooxygenase and two genes possibly involved in acyl-CoA ester metabolism. Reverse transcription-PCR indicated that the etnE and etnA genes were cotranscribed and inducible by ethene and VC. Heterologous expression of the etnE gene in Mycobacterium smegmatis mc 2 155 using the pMV261 vector gave a recombinant strain capable of transforming epoxyethane, epoxypropane, and chlorooxirane. A metabolite identified by mass spectrometry as 2-hydroxyethyl-CoM was produced from epoxyethane. The results indicate that the EaCoMT and monooxygenase enzymes encoded by a single operon (etnEABCD) catalyze the initial reactions in both the VC and ethene assimilation pathways. CoM-mediated reactions appear to be more widespread in bacteria than was previously believed.

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confidence: 99%

Epoxyalkane:Coenzyme M Transferase in the Ethene and Vinyl Chloride Biodegradation Pathways of Mycobacterium Strain JS60

2003

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Distribution of the Coenzyme M Pathway of Epoxide Metabolism among Ethene- and Vinyl Chloride-Degrading Mycobacterium Strains

Coleman¹,

Spain²

2003

Appl Environ Microbiol

109

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Aerobic bacteria that grow on ethene and vinyl chloride (VC) are widely distributed in the environment and have attracted interest because of their potential applications in bioremediation and biocatalysis (5,6,11,12,32,33). The first step in ethene and VC assimilation is known to be a monooxygenase reaction yielding epoxyethane from ethene (5, 7) and chlorooxirane from VC (12, 33), but the downstream pathways are not well understood. Our recent work (5a) has revealed that an epoxyalkane:coenzyme M (CoM) transferase (EaCoMT) enzyme is involved in epoxyethane and chlorooxirane metabolism in Mycobacterium strain JS60, a strain isolated from contaminated groundwater by enrichment on VC as the sole carbon source (5). The EaCoMT reaction is a key step that channels the initial intermediates into central metabolic pathways and also guards against the accumulation of highly toxic and reactive epoxides in the cytoplasm.EaCoMT enzymes have previously been found only in the propene-oxidizing bacteria Xanthobacter strain Py2 and Rhodococcus strain B-276. In such strains, EaCoMT is part of an unusual epoxide carboxylase enzyme complex consisting of EaCoMT, two stereoselective dehydrogenases, and an oxidoreductase/carboxylase (1, 2, 9, 16). In addition to unique biochemical reactions, the propene assimilation pathway is also distinguished by unusual genetic elements. In both strains Py2 and B-276, the propene monooxygenase genes are carried on linear megaplasmids, and in strain Py2, the epoxide carboxylase system and CoM biosynthesis genes are also plasmid borne (18,26).As part of the study that yielded strain JS60, we isolated many other mycobacteria that grew on VC (5). It is not known whether these isolates or similar Mycobacterium strains isolated on ethene (6, 7) possess EaCoMT enzymes. The relationship between strains isolated on VC and ethene is unclear, and the role of factors such as site contamination and geography in the dissemination and evolution of both groups is unknown. On the basis of the recent finding that the EaCoMT gene in Xanthobacter Py2 is carried by a linear plasmid, it might be speculated that similar elements are involved in ethene and VC metabolism. To address these questions, we investigated 10 mycobacteria isolated on VC (6 strains) or ethene (4 strains) from a diverse range of environmental samples. EaCoMT genes and enzyme activities were found in all of the strains. In the VC degraders, the EaCoMT enzymes appear to be encoded on linear megaplasmids. MATERIALS AND METHODSIsolation and growth of bacteria. The growth methods and media used were described previously (5, 5a). Six Mycobacterium strains (JS60, JS61, JS616, JS617, JS619, and JS621) that grow on VC and ethene were previously isolated (5) from groundwater (Plaquemine, La.), activated sludge (Ithaca, N.Y.), pond sediment (Carlyss, La.), activated carbon (Dortmund, Germany), aquifer sediment (Travis Air Force Base, Calif.), and groundwater (Moody Air Force Base, Ga.), respectively. Several mycobacteria that grow on ethene (but not VC) wer...

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“…Alkene-utilizing bacteria immobilized in calcium alginate or K-carrageenan retained 60-100% of their epoxide production activity (Habets-Crutzen et al, 1984).…”

Section: Epoxidesmentioning

confidence: 98%

“…For example, for immobilization of Lavandula vera cells, Nakajima et al (1985) first removed the calcium ions present in the cell suspension by washing with 3% sucrose solution, then mixed the suspension with 5% K-carrageenan solution at 50 C, and finally dropped the mixture rapidly through a wide-mouthed pipette into 0.3 M KCI solution cooled on ice. Alternatively, Habets-Crutzen et al (1984) extruded a carrageenan cell mixture into cold 2% CaC12 solution first and then after 20 minutes replaced the Caa, solution with cold 2% KCI solution. Karkare, Dean and Venkatasubramanian (1985) prepared immobilized yeast beads with K-carrageenan by the following procedure: K-carrageenan, 1.78% (w/ w), was dissolved at 60 C in deionized triple-distilled water with NaCl (8-76 g/1) and silica gel (150 g/1) and then cooled to 40 C; Saccharomyces cerevisiae Ethanol inoculum was added at approximately 5 x 107 cells/ml in growth medium, and beads were immediately cast under sterile conditions by extrusion of the mixture from a vibrating needle into 0.3 m KC1 solution held at 10 C. This procedure produced beads with diameters as small as 0 3 mm.…”

Section: Gels and Other Polymers For Entrapment Or Encapsulation Of Cmentioning

confidence: 99%

Biocatalysis with Immobilized Cells

Akin¹

1987

Biotechnology and Genetic Engineering Reviews

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Production of epoxides from gaseous alkenes by resting-cell suspensions and immobilized cells of alkene-utilizing bacteria

Cited by 87 publications

References 17 publications

Epoxyalkane:Coenzyme M Transferase in the Ethene and Vinyl Chloride Biodegradation Pathways of Mycobacterium Strain JS60

Epoxyalkane:Coenzyme M Transferase in the Ethene and Vinyl Chloride Biodegradation Pathways of Mycobacterium Strain JS60

Distribution of the Coenzyme M Pathway of Epoxide Metabolism among Ethene- and Vinyl Chloride-Degrading Mycobacterium Strains

Biocatalysis with Immobilized Cells

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