Isolation, Cloning, Sequence Analysis and Localization of the Operon Encoding Dimethyl Sulfoxide/Trimethylamine N-oxide Reductase fromRhodobacter capsulatus

Knäblein, Jörg; Mann, Karlheinz; Ehlert, Stephan; Fonstein, Michael; Huber, Robert; Schneider, Frank

doi:10.1006/jmbi.1996.0554

Cited by 17 publications

(17 citation statements)

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“…The dorCBA genes are almost identical to the previously sequenced dmsCBA genes of R. sphaeroides f. sp. denitrificans, and the dorA gene product has extensive homology to the dorA gene product of R. capsulatus (22,37,45,51). Mutations in the dorCBA genes resulted in the inability to use DMSO or TMAO as the terminal electron acceptor in anaerobic respiration and in greatly diminished in vitro DMSOR activity.…”

Section: Discussionmentioning

confidence: 99%

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Characterization of genes encoding dimethyl sulfoxide reductase of Rhodobacter sphaeroides 2.4.1T: an essential metabolic gene function encoded on chromosome II

1997

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Rhodobacter sphaeroides is a gram-negative purple nonsulfur bacterium belonging to the alpha-3 subdivision of the Proteobacteria (49). This organism is capable of a wide range of metabolic lifestyles, exhibiting growth chemoautotrophically, photoautotrophically, photoheterotrophically, chemoheterotrophically, and diazotrophically (reviewed in reference 24). The ability of purple nonsulfur bacteria, as well as other bacterial species, to utilize dimethyl sulfoxide (DMSO) and trimethylamine N-oxide (TMAO) as terminal electron acceptors in the absence of oxygen for chemoheterotrophic growth has been well documented (12,23,52). Both DMSO and TMAO are commonly occurring compounds in nature and play roles in sulfur and nitrogen cycling (1).Under anoxic conditions in the presence of DMSO or TMAO and in the absence of light, R. sphaeroides can grow via a periplasmic DMSO reductase (DMSOR), which serves as the terminal electron acceptor in a cascade in which electrons are transferred from quinol to DMSO or TMAO (24). This process does not require the cytochrome bc 1 complex, and electrons are transferred from quinol to DMSOR via a membranebound b-type cytochrome and a periplasmic c-type cytochrome (25). DMSOR from both R. sphaeroides and the closely related bacterium R. capsulatus have been crystallized and shown to contain the molybdopterin cofactor (Moco) as the sole prosthetic group (for a review, see reference 21).Studies on the genetics of DMSO reduction in both R. sphaeroides and R. capsulatus have been limited mainly to several reports which describe the sequence of the gene encoding DMSOR (2, 22, 37, 51). The dmsA or dorA gene product has extensive homology to other molybdoenzymes such as biotin sulfoxide reductase and TMAO reductase (50). Recently, the sequences of two genes upstream of the dmsA gene of R. sphaeroides f. sp. denitrificans were reported (45). The dmsC and dmsB gene products were shown to be homologous to the torC and torD gene products, which encode a c-type cytochrome and a membrane protein, respectively, and it was suggested that the dmsCBA genes form a transcriptional unit, similar to the torCDA genes in E. coli (27). Studies from our laboratory revealed that R. sphaeroides 2.4.1 T possesses two different circular chromosomes, of ϳ3.0 Mbp (CI) and ϳ0.9 Mbp (CII) (42,43). A number of genes have been shown to be duplicated between CI and CII (5). In contrast, we have previously shown by Tn5 mutagenesis that some pathways, e.g., p-aminobenzoic acid, uracil, histidine, and thymine biosynthesis, are partitioned rather than duplicated between CI and CII (4). These results suggested that CII is an essential genomic element, having unique as well as shared and duplicated functions.Recently, we used a low-redundancy sequencing strategy for analysis of the genetic content of CII (5). We analyzed ϳ300 kb of unique DNA which identified approximately 200 putative open reading frames (ORFs) representing a wide variety of functions, e.g., amino acid biosynthesis, nutrient transporters, redox-active systems, ...

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Section: Discussionmentioning

confidence: 99%

“…Studies on the genetics of DMSO reduction in both R. sphaeroides and R. capsulatus have been limited mainly to several reports which describe the sequence of the gene encoding DMSOR (2,22,37,51). The dmsA or dorA gene product has extensive homology to other molybdoenzymes such as biotin sulfoxide reductase and TMAO reductase (50).…”

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confidence: 99%

Characterization of genes encoding dimethyl sulfoxide reductase of Rhodobacter sphaeroides 2.4.1T: an essential metabolic gene function encoded on chromosome II

1997

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“…During this redox reaction the molybdenum ion changes its oxidation state between Mo(IV) and Mo(VI). DMSOR from R. capsulatus is a single-chain protein consisting of 781 amino acid residues with a molecular mass of 86,574 Da (Knäblein et al, 1996). The enzyme contains a cofactor of 1541 Da, consisting of two molybdopterin guanine dinucleotide (MGD) molecules arranged around a molybdenum center.…”

Section: Introductionmentioning

confidence: 99%

Crystal Structure of Dimethyl Sulfoxide Reductase fromRhodobacter capsulatusat 1.88 Å Resolution

Schneider

Löwe

Huber

et al. 1996

Journal of Molecular Biology

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261

209

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“…Based on sequence homologies, TorA belongs to the Me 2 SO/ TMAO reductase family which includes the M 2 SO/TMAO reductases from Rhodobacter capsulatus and Rhodobacter sphaeroides (5) and TMAO reductase of Shewanella species. 2 These enzymes share several properties: (i) they are all molybdoenzymes located in the periplasm of the bacterium and (ii) in each case, it has been proposed that a membrane anchored pentahemic c type cytochrome feeds electrons to the terminal enzyme (6,7). In E. coli, this cytochrome, TorC, is encoded by torC, the first gene of the torCAD operon (4,8).…”

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confidence: 99%

“…While the role of the TorC and TorA proteins is well documented, the role of the third protein, TorD, predicted in the torCAD operon is not (4). However, the presence of TorD homologue proteins, not only in R. capsulatus and R. sphaeroides Tor systems (6,7), but also in Shewanella species 2 is intriguing and suggests a similar role for this protein in these systems. As TorD contains two small hydrophobic segments, at its amino and carboxyl ends, respectively, it was proposed to be a membranous b type cytochrome involved in the electron transfer pathway for TMAO reduction (4,9).…”

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confidence: 99%

TorD, A Cytoplasmic Chaperone That Interacts with the Unfolded Trimethylamine N-Oxide Reductase Enzyme (TorA) in Escherichia coli

Pommier

Méjean

Giordano

et al. 1998

Journal of Biological Chemistry

124

127

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Reduction of trimethylamine N-oxide (TMAO) in Escherichia coli involves the terminal molybdoreductase TorA, located in the periplasm, and the membrane anchored c type cytochrome TorC. In this study, the role of the TorD protein, encoded by the third gene of torCAD operon, is investigated. Construction of a mutant, in which the torD gene is interrupted, showed that the absence of TorD protein leads to a two times decrease of the final amount of TorA enzyme. However, specific activity and biochemical properties of TorA enzyme were similar to those of the enzyme produced in the wild type. Excess of TorD protein restores the normal level of TorA enzyme, and also, leads to the appearance of a new cytoplasmic form of TorA on SDS-polyacrylamide gel electrophoresis using gentle conditions. This probably indicates a new folding state of the cytoplasmic TorA protein when TorD is overexpressed. BIAcore techniques demonstrated direct specific interaction between the TorA and TorD proteins. This interaction was enhanced when TorA was previously unfolded by heating. Finally, as TorA is a molybdoenzyme, we demonstrated that TorD can interact with TorA before the molybdenum cofactor has been inserted. As TorD homologue encoding genes are found in various TMAO reductase loci, we propose that TorD is a chaperone protein specific for the TorA enzyme. It belongs to a family of TorD-like chaperones present in several bacteria, and, probably, involved in TMAO reductase folding.In Escherichia coli, the main anaerobic respiratory pathway responsible for reduction of trimethylamine N-oxyde (TMAO) 1 to TMA (trimethylamine) requires the products of the torCAD operon which, in anaerobiosis, is induced in the presence of TMAO or related compounds via the two-component regulatory system, TorS/TorR (1, 2).TMAO reductase (TorA), the terminal reductase of this system, is a 97-kDa molybdoprotein encoded by the torA gene (3, 4). Based on sequence homologies, TorA belongs to the Me 2 SO/ TMAO reductase family which includes the M 2 SO/TMAO reductases from Rhodobacter capsulatus and Rhodobacter sphaeroides (5) and TMAO reductase of Shewanella species.2 These enzymes share several properties: (i) they are all molybdoenzymes located in the periplasm of the bacterium and (ii) in each case, it has been proposed that a membrane anchored pentahemic c type cytochrome feeds electrons to the terminal enzyme (6, 7). In E. coli, this cytochrome, TorC, is encoded by torC, the first gene of the torCAD operon (4,8). While the role of the TorC and TorA proteins is well documented, the role of the third protein, TorD, predicted in the torCAD operon is not (4). However, the presence of TorD homologue proteins, not only in R. capsulatus and R. sphaeroides Tor systems (6, 7), but also in Shewanella species 2 is intriguing and suggests a similar role for this protein in these systems. As TorD contains two small hydrophobic segments, at its amino and carboxyl ends, respectively, it was proposed to be a membranous b type cytochrome involved in the electron transfer path...

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Isolation, Cloning, Sequence Analysis and Localization of the Operon Encoding Dimethyl Sulfoxide/Trimethylamine N-oxide Reductase fromRhodobacter capsulatus

Cited by 17 publications

References 50 publications

Characterization of genes encoding dimethyl sulfoxide reductase of Rhodobacter sphaeroides 2.4.1T: an essential metabolic gene function encoded on chromosome II

Characterization of genes encoding dimethyl sulfoxide reductase of Rhodobacter sphaeroides 2.4.1T: an essential metabolic gene function encoded on chromosome II

Crystal Structure of Dimethyl Sulfoxide Reductase fromRhodobacter capsulatusat 1.88 Å Resolution

TorD, A Cytoplasmic Chaperone That Interacts with the Unfolded Trimethylamine N-Oxide Reductase Enzyme (TorA) in Escherichia coli

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