Sensitive electrochemical microbial biosensor for p-nitrophenylorganophosphates based on electrode modified with cell surface-displayed organophosphorus hydrolase and ordered mesopore carbons

• OPH displayed on the surface of Escherichia coli with excellent enzyme activity and stability.
• Sensitive electrochemical microbial biosensor for p-nitrophenyl OPs was developed with ordered mesoporous carbons and OPH-bacteria.
• The amperometric biosensor was capable of detecting trace p-nitrophenyl OPs with high specificity.

A novel electrochemical microbial biosensor for the rapid monitoring of p-nitrophenyl-substituted organophosphates (OPs) compounds based on glass carbon electrode (GCE) modified with both ordered mesopore carbons (OMCs) and cell surface-expressed organophosphorus hydrolase (OPH) (OPH-bacteria/OMCs/GCE) was described in this paper. The genetically engineered Escherichia coli strain surface displayed mutant OPH (S5) with improved enzyme activity and favorable stability was constructed using a newly identified N-terminal of ice nucleation protein as an anchoring motif, which can be used directly without further time-consuming enzyme-extraction and purification, thereafter greatly improved the stability of the enzyme. Compared to OPH-bacteria modified GCE (OPH-bacteria/GCE), the OPH-bacteria/OMCs/GCE not only significantly enhanced the current response but also reduced the oxidation overpotential towards oxidizable p-nitrophenol (p-NP), which was the hydrolysate of p-nitrophenyl-substituted OPs. Under the optimized experimental conditions, at +0.84 V (vs. SCE), the current–time curve was performed with varying OPs concentration. The current response was linear with paraoxon concentration within 0.05–25 μM. Similarly, linear range of 0.05–25 μM was found for parathion, and 0.08–30 μM for methyl parathion. The low limits of detection were evaluated to be 9.0 nM for paraoxon, 10 nM for parathion and 15 nM for methyl parathion (S/N=3). Thus, a highly specific, sensitive and rapid microbial biosensor was established, which holds great promise for on-site detection of trace p-nitrophenyl-substituted OPs.