Versatile artificial mer operons in Escherichia coli towards whole cell biosensing and adsorption of mercury

Abstract

Mercury exists naturally and mainly as a man-made pollutant in the environment, where it exerts adverse effects on local ecosystems and living organisms. It is important to develop an appropriate synthetic biological device that recognizes, detects and removes the bioavailable fraction of environmental mercury. Both single-signal and double-signal output mercury biosensors were assembled using a natural mer operon as a template. Selectivity and sensitivity of whole-cell biosensors based on artificial mer operons were determined. Three whole-cell biosensors were highly stable at very high concentrations of mercuric chloride, and could detect bioavailable Hg(II) in the concentration range of 6.25–200 μM HgCl2. A novel Hg(II) bioadsorption coupled with biosensing artificial mer operon was assembled. This would allow Hg(II)-induced Hg(II) binding protein cell surface display and green fluorescence emission to be achieved simultaneously while retaining the linear relationship between fluorescent signal and Hg(II) exposure concentration. The present study provides an innovative way to simultaneously detect, quantify, and remove bioavailable heavy metal ions using an artificially reconstructed heavy metal resistance operon.

Detection of Bioavailable Cadmium by Double-Color Fluorescence Based on a Dual-Sensing Bioreporter System

“…No continuous increase in the biosensing signals was observed in previously developed CadR-based biosensors within non-cytotoxic concentration of cadmium exposure ( Tao et al, 2013 ; Guo et al, 2021 ). The saturation of metal-binding sites of low-level-background-expressed CadR may be the underlying cause, and it is very common in other developed biosensors using MerR-like regulators ( Joe et al, 2012 ; Guo et al, 2019a ; Hui et al, 2020a ; Zhang et al, 2021 ). Furthermore, the accumulation of red fluorescent protein mCherry responsive to above 50 μM Cd(II) ( Figure 4D ) can be obviously distinguished by the naked eye when compared with the accumulation of green fluorescent protein eGFP ( Figure 4B ).…”

“…A number of studies have demonstrated that metalloregulator MerR responded selectively to Hg(II). MerR-based whole-cell biosensors all showed extraordinary selectivity toward Hg(II) ( Hansen and Sorensen, 2000 ; Wang et al, 2020 ; Zhang et al, 2021 ). If CadR-based biosensing system and MerR-based biosensing system are integrated into one genetic construct, the resultant dual-sensing biosensor will be able to distinguish Cd(II) and Hg(II) simultaneously.…”

“…Finally, the induced groups with 400, 200, 100, 50, 25, 12.5, 6.25, 3.125, 1.56, 0.78, 0.39, 0.2, 0.1, 0.05, 0.025, and 0 μM Cd(II) exposure were obtained. This double dilution method has been described previously in other papers ( Guo et al, 2021 ; Zhang et al, 2021 ). The resultant cultures were grown at 37°C for 12 h before measurement of fluorescent signals and bacterial cell density.…”

Indigoidine biosynthesis triggered by the heavy metal-responsive transcription regulator: a visual whole-cell biosensor

“…The response strength of whole-cell biosensor was determined by both the response rate of the sensing element and the maturation time of the reporter product (Bereza-Malcolm et al 2015;Hansen and Sorensen 2000;Zhang et al 2021). To find the suitable induction time for the two kinds of indigoidinebased biosensors, exponential-phase TOP10/pPmer-ind and TOP10/pPpbr-ind were exposed to 1 μM Hg(II) and 30 μM Pb(II), respectively.…”

“…The extraordinary fidelity, sensory, and regulatory capacities of MerR family regulators have made them as a source of powerful biological parts for the assembly of whole-cell biosensors (Jung and Lee 2019). Among MerR family proteins, well-characterized MerR, PbrR, and CadR have been used as sensing modules to develop many classical heavy metal biosensors, and the output modules containing luciferase, fluorescent protein, and β-galactosidase were usually inserted downstream of the promoter/operator region to replace the detoxification genes (Bereza-Malcolm et al 2016;Cai et al 2018;Guo et al 2019a;Guo et al 2021;Hansen and Sorensen 2000;Kumar et al 2017;Zhang et al 2021). The resultant output signals including luminescence, fluorescence, and enzymatic activity can be detected quantitatively by various types of spectrophotometers.…”