Bioluminescence Biosensor for the Detection of Organomercury Contamination

Abstract

To selectively detect organomercurial compounds in the environment, in this study a bioluminescence biosensor for organomercurials was developed using a bacterial gene expression system for the mercury resistance determinant. merB 3 -Luciferase (mer-lux) transcriptional fusion plasmids pHYΒ3Lux and pHY∆Β3Lux were constructed to evaluate the gene expression system with a new organomercury lyase gene merB 3 from Bacillus megaterium strain MB1, which is resistant to a broad spectrum of mercury compounds, and with its 3'end-deleted defective merB 3 , respectively. Another plasmid, pGR 1 A, encoding an operator/promoter sequence, merR 1 , merE, merT, merP and merA from the same bacterial strain was constructed and used as a transacting gene expression vector which combines the gene expression vector of mer-lux transcriptional fusion plasmids in the same Escherichia coli cells. The transformants that carried a set of the two plasmids were used as biological sensors for the detection of organomercurials. Transformant (E. coli DH5α/pHYB3Lux, pGR1A) is available to distinguish the organomercury from inorganic mercury, since inorganic mercurials can induce the bioluminescence of both the bacterial strain lines with pHY∆B3Lux and pHYB3Lux, whereas organomercurials can only induce the bioluminescence of the lines with pHYB3Lux. These experimental results showed that the transformant with a merB 3 -defective fusion plasmid, and the gene expression vector responded to only mercury chloride. On the other hand, the transformant with an intact merB 3 fusion plasmid and the gene expression vector responded to mercury chloride and all organomercurials tested in the study. The sensor system responded to the existence of the phenyl mercury acetate of 50 nanomolar and was more sensitive than that of inorganic mercury (100 nanomolar). The result also indicated the capability of the system to detect bio-affecting inorganic mercury from several hundred nanomolar to several ten micromolar.

Effect of Inorganic and Organic Ligands on the Bioavailability of Methylmercury as Determined by Using a mer-lux Bioreporter

“…This is consistent with the mechanism of passive diffusion. The plasmid pDES1 is a slight improvement on the design of Endo et al (8), who reported a detection limit of 50 nM for phenylmercury acetate, an organomercurial compound. A 1-to 2-order-of-magnitude improvement in the detection limit allows the examination of bioavailability at concentrations closer to those found in the environment.…”

“…In order to effectively determine organomercurial compounds using a mer-lux plasmid bioreporter, it is necessary to convert organomercurials to Hg 2ϩ . Endo et al (8) incorporated the gene merB into a similar design of a mer-lux plasmid bioreporter (pHYB3Lux) and showed that the detection limit of this new bioreporter for phenylmercury acetate ranged from 50 nM to 5 M. MerB catalyzes the breakdown of phenylmercury and CH 3 Hg(II) to Hg 2ϩ (2,26), which binds to MerR, thereby switching on the lux system. In this study, we attempted to develop a similar biosensor/bioreporter that can quantify the amount of CH 3 Hg(II) that has entered a cell from the medium and to use the biosensor to investigate the impacts of chloride and different forms of NDOM, such as thiols and humic acids, on the bioavailability of CH 3 Hg(II) in bacterial cells.…”

Supramolecular Control of Single‐Crystal‐to‐Single‐Crystal Transformation through Selective Guest Exchange

“…The Nobel Prize in Chemistry of 2008 was awarded to Osamu Shimomura, Martin Chalfie and Roger Tsien as a recognition of their research on the green fluorescent protein (GFP). Bioluminescence has already found successful applications in different fields, such as gene expression,1 biosensors for environmental pollutants,2 and cancer monitoring 3. Among the bioluminescence systems, the firefly luciferin–luciferase system is one of the most widely studied.…”

Application of internal standard method in recombinant luminescent bacteria test

“…The testing medium in this research was completely LB (primitive cell culture or LB diluted sample). Medium composition might affect mercury bioavailability for a bacterial strain, and reduce the sensitivity of the luminescence test, thus changing to an appropriate medium would be helpful to achieve a lower detection limit (Barkay et al, 1997;Endo et al, 2003;Rasmussen et al, 2000;Thouand et al, 2003).…”