Development of a pigment-based whole-cell biosensor for the analysis of environmental copper

Abstract

Using engineered microorganisms to detect heavy metals in the environment has proven to be highly effective and robust. This paper reports on the development of a novel microbial sensor for the detection of copper ions. To develop this microbial sensor, we screened and characterized various biological parts, including promoters, output signals, and hosts. In addition, we used the plant pigment betaxanthin to output fluorescent signals in order to reduce the detection time. The resulting whole-cell biosensor presented a good sensitivity when detecting copper ions in environmental samples including freshwater pond and tap water.Metal ions including iron, copper, zinc, and manganese play important roles as cofactors for enzymes involved in the catalysis of metabolic processes, and the maintenance of cell integrity, which is achieved through regulation by osmotic pressure.1,2 These ions are ubiquitous in all organisms; however, heavy metals can be toxic at high intracellular concentrations.3,4 For example, an excess of copper ions can cause protein dysfunction by interacting strongly with thiol groups and interfere with assembled ironsulfur cofactors.5 This has led many microorganisms to develop the ability to sense small variations in metal concentrations as well as the means to control the inux and efflux of metal ions.

Sensors Applied for the Detection of Pesticides and Heavy Metals in Freshwaters

“…The most sensitive sensor was a bioelectrochemical sensor which was developed by Zhang et al [251] for the detection of silver in tap and lake water, and the LOD was as low as 5:0 × 10 −8 ng/ml. On the contrary, a pigment-based whole-cell biosensor developed for the analysis of copper in pond and tap water showed the highest LOD (5547.6 ng/ml) [262]. The average LOD of heavy metals based on electrochemical methods was 12:187 ± 5:446 ng/ml (n = 65, mean ± SE).…”

Light Emission in Betalains: From Fluorescent Flowers to Biotechnological Applications

“…This was done studying the effect of different promoters and linking the receptor to the expression of the enzyme 4,5-DODA from M. jalapa. This way the bacteria were able to synthesize betalamic acid, which then condensed with amines to form the corresponding fluorescent betaxanthins [90]. This fluorescence was the signal measured, which turned out to be dependent on the concentration of copper.…”

“…Increased production of betalains in plant cell bioreactors can be specifically and easily followed through fluorescence (F) [77]. Fluorescence of betaxanthins is a suitable output signal for virtually any biosensor provided that betalamic acid production is engineered (G) [90]. The success of transgenesis in plants can be ascertained by betaxanthin fluorescence when using genes for betalamic acid-producing dioxygenases (H) [84].…”

“…Fluorescence of betalains provides a reliable and robust signal, which is easily detected by conventional fluorescence apparatus. For this reason, it is a suitable candidate signal for use in biosensors, as demonstrated by the development of whole-cell sensors for the analysis of environmental copper [90]. By using the bacteria Cupriavidus metallidurans and Ralstonia eutropha, it was possible to transform a copper-sensing signal mediated by a kinase into protein expression and light emission mediated by betaxanthins.…”

Integrated optical and electrochemical detection of Cu2+ ions in water using a sandwich amino acid–gold nanoparticle-based nano-biosensor consisting of a transparent-conductive platform

“…The linear range and the limit of detection of the opticalelectrochemical biosensor for detection of copper ions were compared with that of reported values by other researchers. [40][41][42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57][58][59] This comparison is summarized in Table 4. According to the table, the developed LSPR-SWV biosensor was superior to many other methods in terms of linear range and LOD.…”