Robust and tunable performance of a cell-free biosensor encapsulated in lipid vesicles

Cell-free systems have enabled the development of genetically encoded biosensors to detect a range of environmental and biological targets. By encapsulating these systems in synthetic membranes, we can reintroduce features of the cell membrane, including molecular containment and selective permeability, which together could significantly enhance sensing capabilities. Here, we demonstrate robust and tunable performance of an encapsulated fluoride riboswitch inside of lipid vesicles. The riboswitch remains active upon encapsulation in lipid vesicles and responds to fluoride added to the surrounding solution. We find the sensitivity of the encapsulated sensor can be tuned by varying membrane composition. We then show that encapsulation protects the sensor from degradation by the sample and use two types of genetically encoded outputs to detect fluoride in real-world samples. This work establishes the feasibility of vesicle-encapsulated cell-free systems to detect environmentally relevant small molecules.