title: Functional heterologous production of reductive dehalogenases from Desulfitobacterium hafniense strains.
abstract: The anaerobic dehalogenation of organohalides is catalyzed by the reductive dehalogenase (RdhA) enzymes produced in phylogenetically diverse bacteria. These enzymes contain a cobamide cofactor at the active site and two iron-sulfur clusters. In this study, the tetrachloroethene (PCE) reductive dehalogenase (PceA) of the Gram-positive Desulfitobacterium hafniense strain Y51 was produced in a catalytically active form in the nondechlorinating, cobamide-producing bacterium Shimwellia blattae (ATCC 33430), a Gram-negative gammaproteobacterium. The formation of recombinant catalytically active PceA enzyme was significantly enhanced when its dedicated PceT chaperone was coproduced and when 5,6-dimethylbenzimidazole and hydroxocobalamin were added to the S. blattae cultures. The experiments were extended to D. hafniense DCB-2, a reductively dehalogenating bacterium harboring multiple rdhA genes. To elucidate the substrate spectrum of the rdhA3 gene product of this organism, the recombinant enzyme was tested for the conversion of different dichlorophenols (DCP) in crude extracts of an RdhA3-producing S. blattae strain. 3,5-DCP, 2,3-DCP, and 2,4-DCP, but not 2,6-DCP and 3,4-DCP, were reductively dechlorinated by the recombinant RdhA3. In addition, this enzyme dechlorinated PCE to trichloroethene at low rates.
authors.affiliation: Department of Applied and Ecological Microbiology, Institute of Microbiology, Friedrich Schiller University, Jena, Germany.
authors.forename: Anita
authors.initials: A
authors.lastname: Mac Nelly
authors.affiliation: Research Group Mass Spectrometry, Max Planck Institute for Chemical Ecology, Jena, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Kai
authors.affiliation: Research Group Mass Spectrometry, Max Planck Institute for Chemical Ecology, Jena, Germany.
authors.forename: Aleš
authors.initials: A
authors.lastname: Svatoš
authors.affiliation: Department of Applied and Ecological Microbiology, Institute of Microbiology, Friedrich Schiller University, Jena, Germany.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Diekert
authors.affiliation: Department of Applied and Ecological Microbiology, Institute of Microbiology, Friedrich Schiller University, Jena, Germany torsten.schubert@uni-jena.de.
authors.forename: Torsten
authors.initials: T
authors.lastname: Schubert
mesh_terms: D000498:Allyl Compounds / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001562:Benzimidazoles / Q000378:metabolism; D002384:Catalysis; D002733:Chlorophenols / Q000378:metabolism; D003001:Cloning, Molecular; D003038:Cobamides / Q000096:biosynthesis; D003470:Culture Media; D004269:DNA, Bacterial / Q000235:genetics; D045858:Desulfitobacterium / Q000201:enzymology* / Q000235:genetics; D020563:Gammaproteobacteria / Q000201:enzymology; D054879:Halogenation / Q000502:physiology*; D006843:Hydrocarbons, Chlorinated; D006879:Hydroxocobalamin / Q000378:metabolism; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D014241:Trichloroethylene / Q000378:metabolism
chemical_list: D000498:Allyl Compounds; D001426:Bacterial Proteins; D001562:Benzimidazoles; D002733:Chlorophenols; D003038:Cobamides; D003470:Culture Media; D004269:DNA, Bacterial; D006843:Hydrocarbons, Chlorinated; C028879:sulfonated 2,4-dichlorophenol; D014241:Trichloroethylene; C043065:2,3-dichloro-1-propene; C015158:5,6-dimethylbenzimidazole; D010088:Oxidoreductases; C112990:tetrachloroethene dehalogenase; C116190:3,5-dichlorophenol; D006879:Hydroxocobalamin