An enzyme-fused phycobiliprotein synthesis system developed for visual whole-cell biosensors for the detection of cadmium during wastewater treatment

Transcription factor-based whole-cell biosensors are simple and inexpensive tools for monitoring heavy metals during environmental processes. However, most biosensors are depend on instrumentation or have low sensitivity. Here, all the enzymes in the phycobiliprotein synthesis system were fused with the phycocyanin apoprotein to create a system to synthesize enzyme-fused phycobiliproteins with comparable fluorescence. We replaced the biliprotein synthesis system T7 promoter with cadmium-sensing genetic elements to create two visual cadmium biosensors (Epcad-cbsA and Epcad-cbsS). Cd2+ was detected at nanomolar concentrations by measuring the fluorescence intensity and evaluating the degree of color change of the cell pellets. Moreover, good linear correlations were observed with Cd2+ concentrations up to 250 nM. Compared with cellular fluorescence, our cadmium biosensor is more sensitive when cell color is used as the output signal, with detection limits of 7.6 nM for Epcad-cbsA and 3.2 nM for Epcad-cbsS. This visual platform was validated by assessing bioavailable cadmium during wastewater treatment. By immobilizing large amounts of phycobilin with the proteins in cells, our designed biosensors do not require pigment extraction or specialized light sources or detection devices. This work demonstrates that a biliprotein synthesis system can be a novel platform for the development of simple, low-cost, and highly sensitive visual biosensors for real-world applications.