Enhancing the copper-sensing capability of Escherichia coli-based whole-cell bioreporters by genetic engineering.

Abstract

Metals are essential to all organisms; accordingly, cells employ numerous genes to maintain metal homeostasis as high levels can be toxic. In the present study, the gene operons responsive to metal(loid)s were employed to generate bacterial cell-based biosensors to detect target metal(loid)s. The cluster of genes related to copper transport known as the cop-operon is regulated by the interaction between the copA promoter region (copAp) and CueR, turning on and off gene expression upon copper ion binding. Therefore, the detection of copper ions could be achieved by inserting a plasmid harboring the fusion of copAp and reporter genes, such as enzymes and fluorescent genes. However, copAp is not as strong a promoter as other metal-inducible promoters, such as znt-, mer-, and ars-operons; thereby, its sensitivity toward copper ions was not sufficient for quantification. To overcome this problem, we engineered Escherichia coli with a deletion of copA to interfere with copper export from cells. The engineered E. coli whole-cell bioreporter was able to detect copper ions at 0 to 10μM in an aqueous solution. Most importantly, it was specific to copper among several tested heavy metal(loid)s. Therefore, it will likely be useful to detect copper in diverse environmental systems. Although additional improvements are still required to optimize the E. coli-based copper-sensing whole-cell bioreporters presented in this study, our results suggest that there is huge potential to generate whole-cell bioreporters for additional targets by molecular engineering.