Factors influencing parathion degradation by recombinant Escherichia coli with surface-expressed organophosphorus hydrolase.
A unique approach for organophosphorus pesticides detoxification was developed previously by anchoring organophosphorus hydrolase (OPH) onto the surface of Escherichia coli with a tightly regulated tac promoter. The resulting recombinant cells degraded parathion very effectively without the diffusional limitation observed in cells expressing OPH intracellularly. However, the precise conditions for surface targeting or pesticide degradation were not fully understood. In this paper, several factors influencing parathion degradation were investigated. Production of active OPH onto the cell surface was highly host-specific; a high rate of parathion degradation was observed from strains JM105 and XL1-Blue, which regulated production of the OPH fusion very tightly. However, in the absence of ampicillin selection, plasmids were only favorably maintained in strain XL1-Blue. OPH activity was highly dependent on growth conditions. Optimal OPH activity was observed when cells were grown in Luria-Bertani (LB)-buffered medium at 37 degrees C. OPH activity was further improved by supplementing the growth medium with cobalt chloride, which favors the formation of the metal active center. The timing of cobalt addition also influenced parathion degradation. Maximum OPH activity was obtained by adding cobalt to induced cultures during the late stationary phase. The resulting cultures grown under the optimized conditions had an eight-fold increase in parathion degradation.
Kaneva
I
I
Department of Chemical and Environmental Engineering, University of California, Riverside, Riverside, California 92521, USA.
Mulchandani
A
A
Chen
W
W
D043303:Aryldialkylphosphatase; D001673:Biodegradation, Environmental; D002462:Cell Membrane / Q000378:metabolism; D003035:Cobalt / Q000494:pharmacology; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D004950:Esterases / Q000096:biosynthesis*; D007306:Insecticides / Q000378:metabolism*; D007544:Isopropyl Thiogalactoside / Q000096:biosynthesis; D010278:Parathion / Q000378:metabolism*; D011995:Recombination, Genetic; D013045:Species Specificity; D013696:Temperature
D003470:Culture Media; D007306:Insecticides; D007544:Isopropyl Thiogalactoside; D003035:Cobalt; D010278:Parathion; D004950:Esterases; D043303:Aryldialkylphosphatase