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At present, few genetically encoded fluorescent probes are currently available for the analysis of toxic heavy metal ions, and most have poor performance that cannot meet the requirements of sensitive and dynamic detection in living cells. In this study, we designed a single fluorescent protein-based probe sfGFP-MerBD, which can specifically response to Hg2+ with high binding affinity and wide dynamic range. More importantly, the developing probe can timely and reversibly monitor changes of Hg2+ concentration in living mammalian cells. The excellent performance of this probe is largely due to the recognition element of the probe, MerBD, which adopts an unusual planar trigonometric coordination configuration with Hg2+, and the coordination can cause enough conformational change to influence the fluorescence of skeleton protein sfGFP coupled with it. The small peptide MerBD was delicately designed based on the three-dimensional structure of metalloprotein MerR. This novel design strategy solves the challenging problems that there are few natural functional proteins in the process of constructing fluorescent probes for toxic metal ions and some functional proteins cannot be directly used as recognition elements. Based on the new strategy, more genetically encoded fluorescent probes of toxic heavy metal ions can be efficiently constructed and applied in the future. [Display omitted] • Genetically encoded probes are ideal tools in living cellular imaging. • Few genetically encoded probes related for toxic heavy metal are currently available. • Novel strategy is adopted in constructing the probe sfGFP-MerBD for sensing Hg2+. • The probe maintains the unusual planar trigonometric coordination with Hg2+. • The probe responses to Hg2+ with high binding affinity and wide dynamic range. [ABSTRACT FROM AUTHOR]

Hg(II) is well-known for quenching fluorescence in a distance dependent manner. Nevertheless, when we exposed the fluorophore of a green fluorescent protein (GFP) toward Hg(II), through H148C mutation, the GFP fluorescence could be “lighted up” by Hg(II) down to sub-nM level. The detection linear range is 0.5–3.0 nM for protein solutions at 8.0 nM. The GFPH148C protein displayed a promising selectivity toward Hg(II) and also the cellular imaging capacity. Spectra measurements suggested that the ground-state redistribution of protein contributed to the fluorescence enhancement, which was found not limited to Hg(II), and thus presented an opening for building a pool of GFP-based chemosensors toward other heavy metal ions. [ABSTRACT FROM AUTHOR]