Genetically Encoded Biosensors for Constrained Biological Functions in Probiotic Escherichia coli Nissle

Abstract

The probiotic Escherichia coli Nissle (EcN) is an exceptional strain that has attracted significant attention not only for its clinical efficacy in the treatment and prevention of gastrointestinal disorders but also as a burgeoning microbial chassis for living therapeutic applications. However, there is an immediate necessity to develop conditional expression systems that confine the activity of EcN specifically in the gastrointestinal tract, to avoid influencing the environment. Here, we constructed two genetically encoded interchangeable sensors responsive to body temperature at 37 °C, and small molecules such as protocatechuic acid (PCA), a metabolite found in green tea. By employing dCpf1 targeted deactivation of the LacI gene, we thereby coupled the above sensing modules with the P_trc-lacO system and achieved improved signal outputs and relatively high ON/OFF ratios. Subsequently, we validated the biological function of engineering EcN using the enhanced green fluorescent protein (eGFP) in an animal model of mice. Taken together, the construction of genetically encoded sensors to restrict the biological functions of EcN would be applicable for the real-world implementation of living therapeutics or drug delivery.