title: Export of methyl parathion hydrolase to the periplasm by the twin-arginine translocation pathway in Escherichia coli.
abstract: The uptake of organophosphates (OPs) is a rate-limiting factor in whole-cell biocatalysis systems. Here, we report the periplasmic secretion of methyl parathion hydrolase (MPH) by employing the twin-arginine translocation (Tat) pathway in Escherichia coli. The twin-arginine signal peptide of trimethylamine N-oxide reductase (TorA) from E. coli was used for exporting MPH to the periplasm of E. coli, alleviating the substrate uptake limitation. A periplasmic expression vector, pUTM18, coding for TorA-MPH was constructed, and the periplasmic secretion and functionality of MPH were demonstrated by cell fractionation, immunoblotting, and enzyme activity assays. The strain expressing periplasmic MPH showed 3-fold higher whole-cell activity than the control strain expressing cytoplasmic MPH. Suspended cultures also exhibited good stability, retaining almost 100% activity over a period of 2 weeks. Owing to their high activity and superior stability, these "live biocatalysts" are ideal for large-scale detoxification of OPs.
authors.affiliation: State Key Laboratory of Integrated Management of Pest Insects & Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Yang
authors.forename: Roland
authors.initials: R
authors.lastname: Freudl
authors.forename: Chuanling
authors.initials: C
authors.lastname: Qiao
mesh_terms: D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000648:ultrastructure; D015870:Gene Expression; D005822:Genetic Vectors; D008743:Methyl Parathion / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000235:genetics / Q000378:metabolism; D019897:Periplasm / Q000201:enzymology; D010575:Pesticides / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D016981:Plesiomonas / Q000201:enzymology; D011993:Recombinant Fusion Proteins / Q000235:genetics
chemical_list: D009943:Organophosphorus Compounds; D010575:Pesticides; D011993:Recombinant Fusion Proteins; D008743:Methyl Parathion; D010089:Oxidoreductases, N-Demethylating; C023186:trimethylamine dehydrogenase; D010744:Phosphoric Monoester Hydrolases