FACTORS INFLUENCING PARATHION DEGRADATION BY RECOMBINANT ESCHERICHIA-COLI WITH SURFACE-EXPRESSED ORGANOPHOSPHORUS HYDROLASE

A unique approach for organophosphorus pesticides detoxification was d eveloped 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 in tracellularly. However, the precise conditions for surface targeting o r pesticide degradation were not fully understood. In this paper, seve ral factors influencing parathion degradation were investigated. Produ ction 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 fa vorably maintained in strain XL1-Blue. OPH activity was highly depende nt 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 culture s during the late stationary phase. The resulting cultures grown under the optimized conditions had an eight-fold increase in parathion degr adation.