title: A Pseudomonas putida strain genetically engineered for 1,2,3-trichloropropane bioremediation.
abstract: 1,2,3-Trichloropropane (TCP) is a toxic compound that is recalcitrant to biodegradation in the environment. Attempts to isolate TCP-degrading organisms using enrichment cultivation have failed. A potential biodegradation pathway starts with hydrolytic dehalogenation to 2,3-dichloro-1-propanol (DCP), followed by oxidative metabolism. To obtain a practically applicable TCP-degrading organism, we introduced an engineered haloalkane dehalogenase with improved TCP degradation activity into the DCP-degrading bacterium Pseudomonas putida MC4. For this purpose, the dehalogenase gene (dhaA31) was cloned behind the constitutive dhlA promoter and was introduced into the genome of strain MC4 using a transposon delivery system. The transposon-located antibiotic resistance marker was subsequently removed using a resolvase step. Growth of the resulting engineered bacterium, P. putida MC4-5222, on TCP was indeed observed, and all organic chlorine was released as chloride. A packed-bed reactor with immobilized cells of strain MC4-5222 degraded >95% of influent TCP (0.33 mM) under continuous-flow conditions, with stoichiometric release of inorganic chloride. The results demonstrate the successful use of a laboratory-evolved dehalogenase and genetic engineering to produce an effective, plasmid-free, and stable whole-cell biocatalyst for the aerobic bioremediation of a recalcitrant chlorinated hydrocarbon.
authors: Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands Department of Chemistry, University of Engineering and Technology Lahore, Faisalabad Campus, Faisalabad, Pakistan.
authors: Ghufrana
authors: G
authors: Samin
authors: Loschmidt Laboratories and Research Centre for Toxic Compounds in the Environment RECETOX, Faculty of Science, Masaryk University, Brno, Czech Republic.
authors: Martina
authors: M
authors: Pavlova
authors: Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.
authors: M Irfan
authors: MI
authors: Arif
authors: Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.
authors: Christiaan P
authors: CP
authors: Postema
authors: Loschmidt Laboratories and Research Centre for Toxic Compounds in the Environment RECETOX, Faculty of Science, Masaryk University, Brno, Czech Republic.
authors: Jiri
authors: J
authors: Damborsky
authors: Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands d.b.janssen@rug.nl.
authors: Dick B
authors: DB
authors: Janssen
mesh_terms: D001673:Biodegradation, Environmental; D001711:Biotransformation; D004251:DNA Transposable Elements; D004785:Environmental Pollutants / Q000378:metabolism*; D015870:Gene Expression; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism; D060847:Metabolic Engineering; D053858:Metabolic Networks and Pathways / Q000235:genetics; D010957:Plasmids; D011407:Propane / Q000031:analogs & derivatives* / Q000378:metabolism; D016958:Pseudomonas putida / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012641:Selection, Genetic
chemical_list: D004251:DNA Transposable Elements; D004785:Environmental Pollutants; D011994:Recombinant Proteins; C009536:1,2,3-trichloropropane; D006867:Hydrolases; C046137:haloalkane dehalogenase; D011407:Propane