A curcumin direct protein biosensor for cell-free prototyping.

In synthetic biology, biosensors are routinely coupled with a gene expression system for detecting small molecules and physical signals. We reveal a fluorescent complex, based on the interaction of an Escherichia coli double bond reductase ( Ec CurA), as a detection unit with its substrate curcumin-we call this a direct protein (DiPro) biosensor. Using a cell-free synthetic biology approach, we use the Ec CurA DiPro biosensor to fine tune 10 reaction parameters (cofactor, substrate, and enzyme levels) for cell-free curcumin biosynthesis, assisted through acoustic liquid handling robotics. Overall, we increase Ec CurA-curcumin DiPro fluorescence within cell-free reactions by 78-fold. This finding adds to the growing family of protein-ligand complexes that are naturally fluorescent and potentially exploitable for a range of applications, including medical imaging to engineering high-value chemicals.