Activity Improvement of Organophosphorus Hydrolase Enzyme by Error Prone PCR Method

Document Type: Original Article

Authors

1 Molecular Biology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Science, Tehran, Iran

2 Applied Biotechnology Research Center, Baqiyatallah University of Medical Science, Tehran, Iran

Abstract

Introduction: Organophosphorus compounds are frequently used as pesticides and insecticides in agriculture, livestock and home. Because of the high toxicity, it seems is very important its removal from the environment. An enzyme called organophosphorus hydrolase (OPH) is responsible for the decomposition of organophosphorus compounds in most of the strains. Production of enzymes and strains with more efficiency is frequently performed by genetic engineering techniques.
Materials and Methods: In this study, we used PCR-based method for quick and easy improvement in activity of OPH enzyme. We selected 5.5 mM Mg2+ and 0.2 mM and Mn2+ concentrations for high PCR product.
Results: After one round of error prone PCR (epPCR), The 5 number of screened strains (29%) were shown more ability than the native strains to degrade of diazinon, with more than 25% raising ratio. The E6 strain was found to have highest improvement degradation, with 29.3% improvement. At 48-hour time point, the E6 strains were able to completely remove of diazinon.
Conclusions: The epPCR method has the low complexity than other methods and can provide a diverse library include efficient mutants.

Keywords


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