title: Development of a whole-cell biocatalyst/biosensor by display of multiple heterologous proteins on the Escherichia coli cell surface for the detoxification and detection of organophosphates.
abstract: This paper reports the codisplay of organophosphorus hydrolase (OPH) and methyl parathion hydrolase (MPH)-green fluorescent protein (GFP) fusion on the cell surface of Escherichia coli using the truncated ice nucleation protein (INPNC) and Lpp-OmpA as the anchoring motifs. The surface localization of both OPH and MPH-GFP was demonstrated by cell fractionation, Western blot analysis, protease accessibility experiment, and immunofluorescence microscopy. Anchorage of the foreign proteins on the outer membrane neither inhibits cell growth nor affects cell viability. The recombinant strain can be used as a whole-cell biocatalyst and showed a broader substrate range than strains expressing either OPH or MPH. A mixture of six organophosphorus pesticides (OPs) (0.2 mM each) could be degraded completely within 5 h. The broader substrate specificity in combination with the rapid degradation rate makes the recombinant strain a promising candidate for detoxification of OPs. The fluorescence of surface-displayed GFP is very sensitive to environmental pH change. Because hydrolysis of OPs by OPH or MPH generates protons, the recombinant E. coli could be used as a whole-cell biosensor for the rapid detection of OPs by evaluating fluorescence changes as a function of OP concentrations.
authors.affiliation: College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University , Tianjin 300071, China.
authors.forename: Ruihua
authors.initials: R
authors.lastname: Liu
authors.forename: Chao
authors.initials: C
authors.lastname: Yang
authors.forename: Yingming
authors.initials: Y
authors.lastname: Xu
authors.forename: Ping
authors.initials: P
authors.lastname: Xu
authors.forename: Hong
authors.initials: H
authors.lastname: Jiang
authors.forename: Chuanling
authors.initials: C
authors.lastname: Qiao
mesh_terms: D043303:Aryldialkylphosphatase / Q000235:genetics / Q000378:metabolism; D055162:Biocatalysis; D001681:Biological Assay / Q000295:instrumentation / Q000379:methods*; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D002462:Cell Membrane / Q000201:enzymology / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005818:Genetic Engineering; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D010755:Organophosphates / Q000032:analysis* / Q000378:metabolism; D010575:Pesticides / Q000032:analysis* / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D010755:Organophosphates; D010575:Pesticides; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D010744:Phosphoric Monoester Hydrolases; D043303:Aryldialkylphosphatase