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The ability of bacteria to sense specific molecules within their environment and trigger metabolic responses in accordance is an invaluable biotechnological resource. While many transcription factors (TFs) mediating such processes have been studied, only a handful have been leveraged for molecular biology applications. To expand the repertoire of biotechnologically relevant sensors we present a strategy for the construction and testing of chimeric TF libraries, based on the fusion of highly soluble periplasmic binding proteins (PBPs) with DNA-binding domains (DBDs). We validate this concept by constructing and functionally testing two unique sense-and-respond regulators for benzoate, an environmentally and industrially relevant metabolite. This work will enable the development of tailored biosensors for novel synthetic regulatory circuits.

Biosensor libraries harness large classes of binding domains for construction of allosteric transcriptional regulators