Design of a Transcriptional Biosensor for the Portable, On-Demand Detection of Cyanuric Acid

Abstract

Rapid molecular biosensing is an emerging application area for synthetic biology. Here, we engineer a portable biosensor for cyanuric acid (CYA), an analyte of interest for human and environmental health, using a LysR-type transcription regulator (LTTR) from <i>Pseudomonas</i> within the context of <i>Escherichia coli</i> gene expression machinery. To overcome cross-host portability challenges of LTTRs, we rationally engineered hybrid <i>Pseudomonas-E. coli</i> promoters by integrating DNA elements required for transcriptional activity and ligand-dependent regulation from both hosts, which enabled <i>E. coli</i> to function as a whole-cell biosensor for CYA. To alleviate challenges of whole-cell biosensing, we adapted these promoter designs to function within a freeze-dried <i>E. coli</i> cell-free system to sense CYA. This portable, on-demand system robustly detects CYA within an hour from laboratory and real-world samples and works with both fluorescent and colorimetric reporters. This work elucidates general principles to facilitate the engineering of a wider array of LTTR-based environmental sensors.