PacMn - Phytochelatin-actuated complexation of manganese
Due to the industrial revolution, for most people the standard of living drastically increased in the
last 150 years. On the one hand this resulted in an increase of life expectancy, economic prosperity and
improved access to medical care. On the other hand, the development was enabled through the carbon
dioxide emitting exploitation of fossil resources eliciting a gradually increasing burden on our
environment.
As a consequence, next to greenhouse gas (like carbon dioxide) emissions and the accumulation of
microplastic in the oceans, the advancing destruction of ecological and economical valuable soil is a
major problem of the 21st century. The catastrophic global impact of these developments is broadly
demonstrated by the drastic increase of environmental disasters such as droughts, which is also
reflected by the rising numbers of iGEM projects from all around the world, addressing environmental
issues.
An important issue which has re-emerged in scientists’ and environmental organizations’ focus is heavy
metal pollution of the environment. The anthropogenic release of heavy metals leads to their
accumulation in the soil. Here, they are taken up by organisms, including plants, entering the food
chain and finally ending up on our plates, thus posing a threat not only to animals but also us humans.
Detecting heavy metals using chemical testing is often very laborious and expensive, being conducted in
specialized laboratories. Therefore, especially structurally weak regions may not have the respective
infrastructure for preventive testing. Accordingly, the development of more versatile and economic
options, such as biosensors that use biological compounds and organisms for substance detection, has
become a major research focus of synthetic biology.
The rapidly evolving field of synthetic biology and the opportunities proposed by an increasing genetic
toolbox as well as the possibility of providing a product which may improve the standard of living for
many people has inspired us to develop PacMn, a biosensor for the detection and chelation of manganese.
Why manganese?
Despite the successful reduction of acute manganese poisoning at the workplace with the instatement of
workplace security measures, the recent development towards rising chronic manganese poisoning caused by
continuous exposure is very alarming (1). The consequences of manganese poisoning severely attack the
nervous system, causing Manganism, a Parkinson-like disease. Thus, it is of great importance to reduce
chronic manganese exposure (1).
With 0.05mg/l the World Health Organisation (WHO) has determined a limit for manganese in drinking
water. However, especially developing countries like Bangladesh, this threshold is often exceeded in
more than 50 % of the area (2). Via bioaccumulation, manganese accumulates in crops like rice, wheat and
tea and life-stock and fish, respectively (3). As these are basic foods for the majority of the world’s
population, chronic poisoning with manganese is a serious threat which should be eliminated.
Next to the demand for inexpensive alternatives for chemical manganese testing and the global threat
posed by the heavy metal, we were further inspired by the discovery that we are also locally affected by
manganese bioaccumulation. Due to the climate-change triggered increase in groundwater temperatures,
many regions in Germany have shown a measurable increase in manganese in groundwater resources, posing a
threat to the environment and the German public (4). Although the underlying mechanisms have not been
fully elucidated, this further supports the importance of research for heavy metal detection and
clearance systems.
PacMn - Phytochelatin-actuated complexation of manganese
Although there are highly specific heavy metal detection systems available, there is still a demand for
more economic but equally efficient detection methods. Thus, we want to utilize the vast possibilities
proposed by synthetic biology applications for the design of a simple but robust bifunctional biosensor
which not only detects manganese but also clears the environment of the pollutant.
In accordance with this, in this year’s iGEM project, to detect manganese we introduced a manganese
sensing riboswitch and a fluorescent FAST-2-tagged reporter protein into E. coli using classical methods
of synthetic biology. Under the control of either a constitutive anderson promoter the FAST-tagged
protein is continuously transcribed, however only translated upon the interaction of a manganese with
the manganese riboswitch located upstream in the untranslated region (UTR) of the protein. Via the
binding of manganese to this riboswitch, a conformational change is triggered and the ribosomal binding
site (RBS) is released, inducing translation of the FAST-2-tagged reporter. We chose FAST-2 due to
various reasons, namely its versatility and, unlike many established fluorescent proteins such as GFP,
putative application under anaerobic conditions. Furthermore, fluorescence is only detected upon the
addition of a fluorophore, which may be exchanged, allowing for measurements of different wavelengths.
As our bifunctional biosensor should not only sense but also clear manganese, we have additionally
cloned a gene for FAST-2-tagged synthetic phytochelatin into our biosensor sequence. Phytochelatins are
heavy metal detoxifying oligopeptides originating from plants. Via their chelation of manganese, they
can presumably be exploited for the clearance of manganese from the environment.
To conclude, a smart combination of the above mentioned biological modules will be an innovative
contribution to recent research in synthetic biology and metabolic engineering. Furthermore, following
the iGEM tradition, we propose an explicit application with immediate improvement on the quality of life
of many people, regionally and world-wide.
