Abstract

Nowadays, the chemical compounds of organophosphate cause biological contamination due to their presence in the environment and production of metabolites, which endanger the health of humans and organisms. Thus, it is crucial to use a safe method to eliminate the contamination of these compounds. The objective of the present study was cloning and surface expression of the engineered enzyme of organophosphorus hydrolase in E. coli and comparing its function with the non-engineered enzyme. The present study aimed at improving the function of the organophosphorus hydrolase enzyme by expressing this enzyme along with Ice nucleation protein (INP) on the surface of bacterial cells. For this purpose, the OPH enzyme coding sequence was cloned in the pET28a-InaV-N plasmid and sent into E. coli, the strain of BL21 (DE3) and protein expression was evaluated at 37 degrees C after 16 hours of induction. To examine the site of expression of the recombinant protein, cell components (cytoplasm, inner membrane, and outer membrane) were separated and the results were analyzed by using SDS-PAGE. The results revealed that OPH enzyme protein was presented as a protein fusion with a molecular weight of 58 KDa next to the INP anchor on the surface of the outer membrane of the cell. The specific activity of the resulting protein was calculated at 3828.8 U/mg after incubation of this enzyme in the vicinity of the substrate at 37 degrees C. based on the results of the present study, it seems that the expression of engineered enzyme and its presentation on the cell surface is an achievement in using the cell as a degrading agent of organophosphate toxins and eliminating the costs of downstream processes such as purification and refolding of protein.