Evidence for the presence of a CmuA methyltransferase pathway in novel marine methyl halide‐oxidizing bacteria

Schäfer, Hendrik; McDonald, Ian R.; Nightingale, Philip D.; Murrell, J. Colin

doi:10.1111/j.1462-2920.2005.00757.x

Cited by 56 publications

(79 citation statements)

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“…These include the degradation of algal-derived sulfur compounds, such as dimethylsulfoniopropionate, (González et al, 1999), aerobic anoxygenic photosynthesis (Yurkov and Beatty, 1998), methyl halide metabolism (Schafer et al, 2005) and the oxidation of manganese (Hansel and Francis, 2006). The annotation of the genome of Ruegeria pomeroyi DSS-3 (basonym Silicibacter pomeroyi) lead to the discovery of carbon monoxide dehydrogenase (CODH) genes, suggesting yet another biogeochemical role for the MRC (Moran et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Correlating carbon monoxide oxidation with cox genes in the abundant Marine Roseobacter Clade

Cunliffe

2010

The ISME Journal

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The Marine Roseobacter Clade (MRC) is a numerically and biogeochemically significant component of the bacterioplankton. Annotation of multiple MRC genomes has revealed that an abundance of carbon monoxide dehydrogenase (CODH) cox genes are present, subsequently implying a role for the MRC in marine CO cycling. The cox genes fall into two distinct forms based on sequence analysis of the coxL gene; forms I and II. The two forms are unevenly distributed across the MRC genomes. Most (18/29) of the MRC genomes contain only the putative form II coxL gene. Only 10 of the 29 MRC genomes analysed have both the putative form II and the definitive form I coxL. None have only the form I coxL. Genes previously shown to be required for post-translational maturation of the form I CODH enzyme are absent from the MRC genomes containing only form II. Subsequent analyses of a subset of nine MRC strains revealed that only MRC strains with both coxL forms are able to oxidise CO.

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Section: Introductionmentioning

confidence: 99%

Correlating carbon monoxide oxidation with cox genes in the abundant Marine Roseobacter Clade

Cunliffe

2010

The ISME Journal

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“…Probe labeling and Southern hybridization were carried out as described previously (31). An EcoRI fragment of approximately 6.5 kb and a KpnI fragment of approximately 10 kb were identified as targets for cloning into pUC18 as described previously (31). Transformants containing inserts from size-fractionated EcoRI and KpnI digests were picked into 96-well plates containing 150 l LB broth (100 g ampicillin/ml) per well and incubated overnight at 37°C.…”

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“…Genomic DNA was digested with restriction endonucleases suitable for subsequent cloning of restriction fragments into pUC18. Digested DNA was electrophoresed and transferred onto HybondNϩ membrane (Amersham, Little Chalfont, United Kingdom) for Southern analysis essentially as described previously (31). A 25-ng sample of the PCR product obtained with primer combination dmoA6F/dmoARI was purified, and the probe was labeled using [␣-32 P]dCTP.…”

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“…Primers dmoARI (5Ј-GCCTGRAAGATNACNG GNGT-3Ј) and dmoARL (5Ј-GCCTGRAAVAGNACNGGNGT-3Ј) were based on the underlined part of an internal peptide sequence (TPV[I/L]FQAGASGR, where the fourth position could be a leucine or isoleucine residue). The primer combinations dmoA6F/dmoARI and dmoA6F/dmoARL were used to amplify gene fragments of the DMS monooxygenase subunit gene from 50 ng H. sulfonivorans DNA in a PCR consisting of an initial denaturation for 5 min at 95°C, followed by 35 cycles of 95°C for 1 min, 50°C for 1 min, and 72°C for 1 min, followed by a final extension step at 72°C for 10 min using a PCR setup which was otherwise as described previously (31). Genomic DNA was digested with restriction endonucleases suitable for subsequent cloning of restriction fragments into pUC18.…”

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confidence: 99%

“…A 25-ng sample of the PCR product obtained with primer combination dmoA6F/dmoARI was purified, and the probe was labeled using [␣-32 P]dCTP. Probe labeling and Southern hybridization were carried out as described previously (31). An EcoRI fragment of approximately 6.5 kb and a KpnI fragment of approximately 10 kb were identified as targets for cloning into pUC18 as described previously (31).…”

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Purification and Characterization of Dimethylsulfide Monooxygenase fromHyphomicrobium sulfonivorans

et al. 2011

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Dimethylsulfide (DMS) is a volatile organosulfur compound which has been implicated in the biogeochemical cycling of sulfur and in climate control. Microbial degradation is a major sink for DMS. DMS metabolism in some bacteria involves its oxidation by a DMS monooxygenase in the first step of the degradation pathway; however, this enzyme has remained uncharacterized until now. We have purified a DMS monooxygenase from Hyphomicrobium sulfonivorans, which was previously isolated from garden soil. The enzyme is a member of the flavin-linked monooxygenases of the luciferase family and is most closely related to nitrilotriacetate monooxygenases. ions. The purified enzyme had no activity with the substrates of related enzymes, including alkanesulfonates, aldehydes, nitrilotriacetate, or dibenzothiophenesulfone. The gene encoding the 53-kDa enzyme subunit has been cloned and matched to the enzyme subunit by mass spectrometry. DMS monooxygenase represents a new class of FMNH 2 -dependent monooxygenases, based on its specificity for dimethylsulfide and the molecular phylogeny of its predicted amino acid sequence. The gene encoding the large subunit of DMS monooxygenase is colocated with genes encoding putative flavin reductases, homologues of enzymes of inorganic and organic sulfur compound metabolism, and enzymes involved in riboflavin synthesis.Dimethylsulfide (DMS) is a volatile organosulfur compound, important in the biogeochemical cycling of sulfur and global climate regulation (4, 9). Bacterial metabolism of DMS is an important sink of the compound in nature and is thought to account for degradation of over 80% of the DMS produced in the marine environment. Although bacterial pathways of DMS degradation have been studied previously in Hyphomicrobium spp. and in Thiobacillus spp. (12, 36), they remain poorly characterized, and few enzymes of DMS metabolism have been purified (see reference 32). DMS monooxygenase was first reported from an assay of NADH-dependent oxygen uptake in the presence of DMS by cell extracts of Hyphomicrobium S (12), an activity also demonstrated in cell extracts of other Hyphomicrobium, Thiobacillus, and Arthrobacter isolates (6,7,34), with specific activities around 30 nmol NADH oxidized min Ϫ1 mg protein Ϫ1 . The enzyme has not previously been purified or characterized.The aims of this study were to purify and characterize the DMS monooxygenase enzyme from a member of the genus Hyphomicrobium. Since Hyphomicrobium S is no longer available, studies were undertaken using the type strain of H. sulfonivorans. The strain was originally isolated from garden soil and grows on DMS, as well as the related compounds dimethyl sulfoxide (DMSO) and dimethylsulfone (DMSO 2 ). During growth on DMSO 2 , H. sulfonivorans first reduces DMSO 2 to DMSO by a dimethylsulfone reductase, and subsequently a DMSO reductase converts DMSO to DMS, which is further oxidized to methanethiol and formaldehyde by a DMS mono-

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Hydrogen and carbon isotope fractionation during degradation of chloromethane by methylotrophic bacteria

et al. 2013

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Chloromethane (CH3Cl) is a widely studied volatile halocarbon involved in the destruction of ozone in the stratosphere. Nevertheless, its global budget still remains debated. Stable isotope analysis is a powerful tool to constrain fluxes of chloromethane between various environmental compartments which involve a multiplicity of sources and sinks, and both biotic and abiotic processes. In this study, we measured hydrogen and carbon isotope fractionation of the remaining untransformed chloromethane following its degradation by methylotrophic bacterial strains Methylobacterium extorquens CM4 and Hyphomicrobium sp. MC1, which belong to different genera but both use the cmu pathway, the only pathway for bacterial degradation of chloromethane characterized so far. Hydrogen isotope fractionation for degradation of chloromethane was determined for the first time, and yielded enrichment factors (ε) of −29‰ and −27‰ for strains CM4 and MC1, respectively. In agreement with previous studies, enrichment in 13C of untransformed CH3Cl was also observed, and similar isotope enrichment factors (ε) of −41‰ and −38‰ were obtained for degradation of chloromethane by strains CM4 and MC1, respectively. These combined hydrogen and carbon isotopic data for bacterial degradation of chloromethane will contribute to refine models of the global atmospheric budget of chloromethane.

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Evidence for the presence of a CmuA methyltransferase pathway in novel marine methyl halide‐oxidizing bacteria

Cited by 56 publications

References 53 publications

Correlating carbon monoxide oxidation with cox genes in the abundant Marine Roseobacter Clade

Correlating carbon monoxide oxidation with cox genes in the abundant Marine Roseobacter Clade

Purification and Characterization of Dimethylsulfide Monooxygenase fromHyphomicrobium sulfonivorans

Hydrogen and carbon isotope fractionation during degradation of chloromethane by methylotrophic bacteria

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