Engineering immobilized microbes with surface-displayed cold-adapted nitroreductase: An efficient strategy for nitrobenzene degradation at low temperature

Abstract

Nitrobenzene (NB), a typical example of aromatic nitro compounds, has become a significant pollutant because its wastewater is difficult to degrade in low-temperature environments. This study constructed an engineered strain displaying a cold-adapted nitroreductase (PsNTR) for enhanced low-temperature degradation efficiency, employing a cell surface display system in which the N-terminal domain of the ice nucleation protein (INPN) served as the anchoring carrier. SDS-PAGE, Western blot analysis, and protease accessibility assays confirmed that the INPN-PsNTR fusion protein (55.40 kDa) was stably expressed and displayed on the surface of Escherichia coli. The engineered strain BL21/INPN-PsNTR maintained outer membrane integrity and exhibited robust growth during the display process. To further enhance the storage stability and reusability of the engineered strain for practical applications, it was immobilized using modified diatomite (DA-PEI/GA). Notably, PsNTR@DA-PEI/GA demonstrated high efficiency in degrading NB at low temperatures, with a degradation rate of 92.75% for 500 mg/L NB within 120 h at 15 °C. In summary, this study employed PsNTR surface display technology as the core strategy, combined with a cell immobilization approach, to establish a novel and promising technological pathway for efficient and green degradation of NB at low temperatures, thereby offering a new strategy for the remediation of NB contamination.