Dataset Information

Genomic, proteomic, and biochemical analysis of the organohalide respiratory pathway in Desulfitobacterium dehalogenans.

ABSTRACT: Desulfitobacterium dehalogenans is able to grow by organohalide respiration using 3-chloro-4-hydroxyphenyl acetate (Cl-OHPA) as an electron acceptor. We used a combination of genome sequencing, biochemical analysis of redox active components, and shotgun proteomics to study elements of the organohalide respiratory electron transport chain. The genome of Desulfitobacterium dehalogenans JW/IU-DC1(T) consists of a single circular chromosome of 4,321,753 bp with a GC content of 44.97%. The genome contains 4,252 genes, including six rRNA operons and six predicted reductive dehalogenases. One of the reductive dehalogenases, CprA, is encoded by a well-characterized cprTKZEBACD gene cluster. Redox active components were identified in concentrated suspensions of cells grown on formate and Cl-OHPA or formate and fumarate, using electron paramagnetic resonance (EPR), visible spectroscopy, and high-performance liquid chromatography (HPLC) analysis of membrane extracts. In cell suspensions, these components were reduced upon addition of formate and oxidized after addition of Cl-OHPA, indicating involvement in organohalide respiration. Genome analysis revealed genes that likely encode the identified components of the electron transport chain from formate to fumarate or Cl-OHPA. Data presented here suggest that the first part of the electron transport chain from formate to fumarate or Cl-OHPA is shared. Electrons are channeled from an outward-facing formate dehydrogenase via menaquinones to a fumarate reductase located at the cytoplasmic face of the membrane. When Cl-OHPA is the terminal electron acceptor, electrons are transferred from menaquinones to outward-facing CprA, via an as-yet-unidentified membrane complex, and potentially an extracellular flavoprotein acting as an electron shuttle between the quinol dehydrogenase membrane complex and CprA.

SUBMITTER: Kruse T

PROVIDER: S-EPMC4325113 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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Genomic, proteomic, and biochemical analysis of the organohalide respiratory pathway in Desulfitobacterium dehalogenans.

Kruse Thomas T van de Pas Bram A BA Atteia Ariane A Krab Klaas K Hagen Wilfred R WR Goodwin Lynne L Chain Patrick P Boeren Sjef S Maphosa Farai F Schraa Gosse G de Vos Willem M WM van der Oost John J Smidt Hauke H Stams Alfons J M AJ

Journal of bacteriology 20141215 5

Desulfitobacterium dehalogenans is able to grow by organohalide respiration using 3-chloro-4-hydroxyphenyl acetate (Cl-OHPA) as an electron acceptor. We used a combination of genome sequencing, biochemical analysis of redox active components, and shotgun proteomics to study elements of the organohalide respiratory electron transport chain. The genome of Desulfitobacterium dehalogenans JW/IU-DC1(T) consists of a single circular chromosome of 4,321,753 bp with a GC content of 44.97%. The genome co ...[more]

PMID: 25512312

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Project description:Desulfitobacterium hafniense was originally isolated for its ability to use organohalogens as terminal electron acceptors in the process of organohalide respiration (OHR). However, in contrast to obligate OHR bacteria, Desulfitobacterium spp. show a highly versatile energy metabolism with the capacity to use many possible electron donors and acceptors and to grow fermentatively also. The sequencing of several Desulfitobacterium genomes displaying numerous and apparently redundant members of redox enzyme families has confirmed their large metabolic potential. Nonetheless, the enzymes responsible for many metabolic traits are not yet identified. In the present work, we conducted an extended proteomic study by comparing the proteomes of D. hafniense strain DCB-2 cultivated in different combinations of electron donors and acceptors, triggering five alternative respiratory metabolisms that include OHR, as well as fermentation. The Tandem Mass Tag labelling proteomic approach allowed us to identify almost 60% of the predicted proteome of strain DCB-2 in all six growth conditions and to quantify the relative abundance of 2796 proteins. This dataset was analysed in order to highlight the proteins that were up-regulated in one or a subset of growth conditions to identify possible key players in the different energy metabolisms. The addition of sodium sulfide as reducing agent in the medium – a very widespread practice in the cultivation of strictly anaerobic bacteria – triggered the expression of the dissimilatory sulfite reduction pathway in relatively less favourable conditions such as fermentative growth on pyruvate, respiration with H2 as electron donor and OHR conditions. The presence of H2, CO2 and acetate in the medium induced several metabolic pathways involved in carbon metabolism including the Wood-Ljungdahl pathway and two pathways related to the fermentation of butyrate that rely on electron-bifurcating enzymes. While the predicted fumarate reductase appears to be constitutively expressed, a new lactate dehydrogenase and lactate transporters were identified. Finally, the OHR metabolism with 3-chloro-4-hydroxyphenylacetate as electron acceptor strongly induced proteins encoded in several reductive dehalogenase gene clusters including four new proteins, the function of which is discussed. Overall, we believe that this extended proteomic database represents a new landmark in understanding the metabolic versatility of Desulfitobacterium spp. and provides a solid basis for addressing present and future research questions.

Project description:IntroductionDesulfitobacterium hafniense was isolated for its ability to use organohalogens as terminal electron acceptors via organohalide respiration (OHR). In contrast to obligate OHR bacteria, Desulfitobacterium spp. show a highly versatile energy metabolism with the capacity to use different electron donors and acceptors and to grow fermentatively. Desulfitobacterium genomes display numerous and apparently redundant members of redox enzyme families which confirm their metabolic potential. Nonetheless, the enzymes responsible for many metabolic traits are not yet identified.MethodsIn the present work, we conducted an extended proteomic study by comparing the proteomes of Desulfitobacterium hafniense strain DCB-2 cultivated in combinations of electron donors and acceptors, triggering five alternative respiratory metabolisms that include OHR, as well as fermentation. Tandem Mass Tag labelling proteomics allowed us to identify and quantify almost 60% of the predicted proteome of strain DCB-2 (2,796 proteins) in all six growth conditions. Raw data are available via ProteomeXchange with identifier PXD030393.Results and discussionThis dataset was analyzed in order to highlight the proteins that were significantly up-regulated in one or a subset of growth conditions and to identify possible key players in the different energy metabolisms. The addition of sodium sulfide as reducing agent in the medium - a very widespread practice in the cultivation of strictly anaerobic bacteria - triggered the expression of the dissimilatory sulfite reduction pathway in relatively less favorable conditions such as fermentative growth on pyruvate, respiration with H2 as electron donor and OHR conditions. The presence of H2, CO2 and acetate in the medium induced several metabolic pathways involved in carbon metabolism including the Wood-Ljungdahl pathway and two pathways related to the fermentation of butyrate that rely on electron-bifurcating enzymes. While the predicted fumarate reductase appears to be constitutively expressed, a new lactate dehydrogenase and lactate transporters were identified. Finally, the OHR metabolism with 3-chloro-4-hydroxyphenylacetate as electron acceptor strongly induced proteins encoded in several reductive dehalogenase gene clusters, as well as four new proteins related to corrinoid metabolism. We believe that this extended proteomic database represents a new landmark in understanding the metabolic versatility of Desulfitobacterium spp. and provides a solid basis for addressing future research questions.

Dataset Information

Genomic, proteomic, and biochemical analysis of the organohalide respiratory pathway in Desulfitobacterium dehalogenans.

Publications

Genomic, proteomic, and biochemical analysis of the organohalide respiratory pathway in Desulfitobacterium dehalogenans.

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