Efficient Surface Display of Diisopropylfluorophosphatase (DFPase) in E. coli for Biodegradation of Toxic Organophosphorus Compounds (DFP and Cp).

Abstract

Compounds including organophosphorus pesticides (OPs) and chemical nerve agents are toxic compounds synthesized recently which disrupt the mechanisms of neural transmission. Therefore, a critical requirement is the development of a bio-refining technology to facilitate the biodegradation of organophosphorus pollutants. The diisopropylfluorophosphatase (DFPase, EC 3.1.8.2) from the ganglion and brain of Loligo vulgaris acts on P-F bonds present in some OPs. Intracellular production of OPs-degrading enzymes or the use of native bacteria and fungi leads to a low degradation rate of OPs due to a mass transfer issue which reduces the overall catalytic efficiency. To overcome this challenge, we expressed DFPase on the surface of E. coli for the first time by employing the N-terminal domain of the ice nucleation protein (InaV-N) as an anchoring motif. Tracking the recombinant protein confirmed that DFPase is successfully located on the outer membrane. Further studies on its activity to degrade diisopropylfluorophosphate (DFP) showed its significant ability for the biodegradation of diisopropylfluorophosphate (DFP) with a specific activity of 500 U/mg of wet cell weight. Recombinant cells could also degrade chlorpyrifos (Cp) with an activity equivalent to a maximum value of 381.44 U/ml with a specific activity of 476.75 U/mg of cell, analyzed using HPLC technique. The optimum activity of purified DFPase was found at 30 °C. A more increased activity was also obtained in the presence of glucose-mineral-salt (GMS) supplemented with tryptone and 100 mg/L Co(2+) ion. These results highlight the high potential of the InaV-N anchoring domain to produce an engineered bacterium that can be used in the bioremediation of pesticide-contaminated environments.

• Authors: Ali Mohammad Latifi, Ali Karami, Samaneh Khodi
• Journal: Applied biochemistry and biotechnology (Appl Biochem Biotechnol) Vol. 177 Issue 3 Pg. 624-36 (Oct 2015) ISSN: 1559-0291 [Electronic] United States
• PMID: 26239441 (Publication Type: Journal Article)

Chemical References

• Environmental Pollutants
• Isoflurophate
• Phosphoric Triester Hydrolases
• diisopropyl-fluorophosphatase
• Chlorpyrifos

Topics

• Biodegradation, Environmental
• Chlorpyrifos (isolation & purification, metabolism)
• Environmental Pollutants (isolation & purification, metabolism)
• Escherichia coli (genetics)
• Isoflurophate (isolation & purification, metabolism)
• Phosphoric Triester Hydrolases (chemistry, genetics, metabolism)
• Protein Structure, Tertiary
• Pseudomonas syringae (enzymology, genetics)