title: Surface display of organophosphorus hydrolase on E. coli using N-terminal domain of ice nucleation protein InaV.
abstract: Recombinant Escherichia coli displaying organophosphorus hydrolase (OPH) was used to overcome the diffusion barrier limitation of organophosphorus pesticides. A new anchor system derived from the N-terminal domain of ice-nucleation protein from Pseudomonas syringae InaV (InaV-N) was used to display OPH onto the surface. The designed sequence was cloned in the vector pET-28a(+) and then was expressed in E. coli. Tracing of the expression location of the recombinant protein using SDS-PAGE showed the presentation of OPH by InaV-N on the outer membrane, and the ability of recombinant E. coli to utilize diazinon as the sole source of energy, without growth inhibition, indicated its significant activity. The location of OPH was detected by comparing the activity of the outer membrane fraction with the inner membrane and cytoplasm fractions. Studies revealed that recombinant E. coli can degrade 50% of 2 mM chlorpyrifos in 2 min. It can be concluded that InaV-N can be used efficiently to display foreign functional protein, and these results highlight the high potential of an engineered bacterium to be used in bioremediation of pesticide-contaminated sources in the environment.
authors.affiliation: Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
authors.forename: Samaneh
authors.initials: S
authors.lastname: Khodi
authors.forename: Ali Mohammad
authors.initials: AM
authors.lastname: Latifi
authors.forename: Mojtaba
authors.initials: M
authors.lastname: Saadati
authors.forename: Morteza
authors.initials: M
authors.lastname: Mirzaei
authors.forename: Hossein
authors.initials: H
authors.lastname: Aghamollaei
mesh_terms: D043303:Aryldialkylphosphatase / Q000235:genetics / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D004390:Chlorpyrifos / Q000378:metabolism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D030781:Organisms, Genetically Modified; D010957:Plasmids; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D001425:Bacterial Outer Membrane Proteins; C108515:InaV protein, Pseudomonas syringae; D011993:Recombinant Fusion Proteins; D043303:Aryldialkylphosphatase; D004390:Chlorpyrifos