mpd Gene Cloning, Functional Expression of MPH in Multiple Hosts and Its Application in Bioremediation of Organophosphorus Pesticides
An effective chlorpyrifos-degrading bacterium was isolated from an organophosphorus pesticides manufacturer and identified as the genus Stenotrophomonas. The inoculation of the isolate (10 <6>cells/g) to soil treated with 100mg/kg chlorpyrifos resulted in a higher degradation rate than in noninoculated soils. The gene encoding the organophosphorus hydrolase was cloned from the strain and expressed in E. coli. Sequence blast analysis indicated that it has 98.5％ similar to methyl parathion hydrolase (MPH). Furthermore, MPH was displayed on the cell surface using N-and C-terminal domains of the ice nucleation protein (INPNC) from P. syringae as an anchoring motif. A shuttle vector pINCM coding for INPNC-MPH was constructed and used to target MPH onto the surface of P. putida JS444, overcoming the potential substrate uptake limitation. Over 90％ of the MPH activity was located on the cell surface as determined by cell fractionation experiments. The surface localization of the INPNC- MPH fusion was further verified by Western blot analysis and immunofluorescence microscopy. The engineered P. putida JS444 degraded organophosphates (OPs) as well as p-nitrophenol (PNP) rapidly. Functional expression of mpd gene in multiple hosts makes it a promising candidate for OP detoxification.
chlorpyrifos mpd gene methyl parathion hydrolase surface display ice nucleation protein whole-cell biocatalyst
Yang Chao Li Lin Qiao Chuanling
State Key Laboratory of Integrated Management of Pest Insects & Rodents, Institute of Zoology, Chine Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, P. R. C State Key Laboratory of Integrated Management of Pest Insects & Rodents, Institute of Zoology, Chine
国际会议
北京
英文
162-174
2007-10-09（万方平台首次上网日期，不代表论文的发表时间）