Heavy metal(loid) biosensor based on split-enhanced green fluorescent protein: development and characterization.

Heavy metal(loid)s such as Cd and Hg adversely affect human health and are therefore strictly regulated and monitored; however, their quantitation in the environment is usually performed by expensive and time-consuming instrumental analysis techniques, which necessitates the search for more practical alternatives. Herein, we prepare enhanced green fluorescent protein (eGFP)-based biomolecules for metal(loid) sensing by insertion of metal-binding loops (MBLs) into a loop region of eGFP to render this protein inactive and show that the binding of metal ions to MBLs induces a conformational change and restores the original activity. Specifically, eGFP with an MBL sequenced as CTTCGCG regains fluorescence upon exposure to Cd and Hg, which allows the above metals to be quantified in the concentration range of 0-5 μM. For practical applicability verification, the developed sensing platform is used to quantify Cd in artificially amended soil and water samples. Although the obtained results imply that sensor performance needs to be significantly improved, the presented design concept is believed to be of high value to researchers in the field of heavy metal sensing and facilitate the development of new biosensors.