Difference between revisions of "Team:Utrecht/Description"

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<p>The past decades have brought astonishing progress in the development and use of pharmaceuticals promoting public health. Unfortunately, increasing amounts of medical waste now threaten our ecosystems. We therefore developed Detaxion, a biosensor aimed to rapidly identify harmful water-based pharmaceuticals. Detaxion is synthetic biology-based, and is highly amendable to detect various pharmaceutical species. Specifically, we mutated the binding site of the chemotaxis receptor Tar in E. coli bacteria to recognize paired pharmaceutical compounds. Accordingly, upon ligand binding the fluorescence of the Tar-receptor associated BRET-pair decreases in a quantifiable manner. We additionally introduced customized methylation-sites to extend the detection range of the Detaxion biosensor. Our results thus far show successful measurement of the fluorescence energy transfer pairs used in Detaxion. Moreover, we developed a capillary-based assay to confirm BRET measurements. Taken together, Detaxion constitutes a synthetic biology-based approach to detect and quantify pharmaceutical waste in water, for safeguarding of vital water supplies.
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<p>Humans have been polluting water with natural products for hundreds of years. However, during the last decennia the amount of unnatural chemicals that are being produced, used and carelessly disposed of, has been increasing rapidly. A substantial part of these chemicals introduced in our water supplies are derived from pharmaceuticals.</p>
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<p>We focussed on the one pharmaceutical contaminant that is associated with current day lifestyle, Selective Serotonin Reuptake Inhibitors (SSRI), commonly known as antidepressants. After leaving the human body, the remainder of the medicine ends up in the sewage system and is not broken down readily by the sewage treatment plants, resulting in an increasing concentration in surface waters. Even in low concentrations, antidepressant waste products pose a threat to ecological systems, for example because they are still biologically active in vertebrates, changing their behaviour and fertility. Current techniques for the detection of pharmaceutical waste products are either expensive or inaccurate. An improved system is therefore needed.</p>
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<p>Consultation of experts and stakeholders provided us with five core requirements:</p>
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<li>Possibility for detecting diverse compounds</li>
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<li>Accurate detection at different concentrations</li>
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<li>Clear and easily measurable detection signal</li>
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<li>Rapid signaling</li>
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<li>Low-cost system</li>
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<p>These requirements were met by developing DeTaXion, a biodetection system that exploits the fast and highly customizable chemotaxis pathway of <i>Escherichia coli</i>, a bacterium that can be produced at low cost. This pathway was adapted using a three step approach to accomplish the desired biosensor. First, mutating the binding site of the <i>E. coli</i> chemotaxis receptor Tar will allow the bacteria to recognize diverse pharmaceutical compounds. We have already developed and implemented a validation assay to assess the functionality of the receptor. Next, customized methylation sites can be introduced to extend the detection range of the DeTaXion biosensor and allow accurate pollutant detection at different concentration ranges. We have mathematically modelled the effect of changes in the methylation sites to assess the effect on the detection range. Finally, a bioluminescence resonance energy transfer (BRET) pair was introduced to generate an easily detectable and quantifiable signal. We have successfully prepared fusion mutants of the relevant chemotaxis proteins as an important step to achieve this.</p>
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<p>The mutated <i>E. coli</i> was integrated into an easily usable hand-held application. Our design includes various features to solve safety, regulatory, and technical issues. Ultimately, the product will enable low-cost, rapid, and accurate detection of pharmaceutical waste products for diverse ligands and at a range of concentrations. It can be used as a stand-alone product to continuously monitor a water supply, or to complement traditional chemical analyses, thereby improving the detection of pharmaceutical waste products in aquatic systems and contribute to a cleaner environment.</p>
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Revision as of 13:25, 15 October 2018

Description Safety Model BRETAssay EndProduct Biobricks Receptor Methylation

Humans have been polluting water with natural products for hundreds of years. However, during the last decennia the amount of unnatural chemicals that are being produced, used and carelessly disposed of, has been increasing rapidly. A substantial part of these chemicals introduced in our water supplies are derived from pharmaceuticals.

We focussed on the one pharmaceutical contaminant that is associated with current day lifestyle, Selective Serotonin Reuptake Inhibitors (SSRI), commonly known as antidepressants. After leaving the human body, the remainder of the medicine ends up in the sewage system and is not broken down readily by the sewage treatment plants, resulting in an increasing concentration in surface waters. Even in low concentrations, antidepressant waste products pose a threat to ecological systems, for example because they are still biologically active in vertebrates, changing their behaviour and fertility. Current techniques for the detection of pharmaceutical waste products are either expensive or inaccurate. An improved system is therefore needed.

Consultation of experts and stakeholders provided us with five core requirements:

  • Possibility for detecting diverse compounds
  • Accurate detection at different concentrations
  • Clear and easily measurable detection signal
  • Rapid signaling
  • Low-cost system

These requirements were met by developing DeTaXion, a biodetection system that exploits the fast and highly customizable chemotaxis pathway of Escherichia coli, a bacterium that can be produced at low cost. This pathway was adapted using a three step approach to accomplish the desired biosensor. First, mutating the binding site of the E. coli chemotaxis receptor Tar will allow the bacteria to recognize diverse pharmaceutical compounds. We have already developed and implemented a validation assay to assess the functionality of the receptor. Next, customized methylation sites can be introduced to extend the detection range of the DeTaXion biosensor and allow accurate pollutant detection at different concentration ranges. We have mathematically modelled the effect of changes in the methylation sites to assess the effect on the detection range. Finally, a bioluminescence resonance energy transfer (BRET) pair was introduced to generate an easily detectable and quantifiable signal. We have successfully prepared fusion mutants of the relevant chemotaxis proteins as an important step to achieve this.

The mutated E. coli was integrated into an easily usable hand-held application. Our design includes various features to solve safety, regulatory, and technical issues. Ultimately, the product will enable low-cost, rapid, and accurate detection of pharmaceutical waste products for diverse ligands and at a range of concentrations. It can be used as a stand-alone product to continuously monitor a water supply, or to complement traditional chemical analyses, thereby improving the detection of pharmaceutical waste products in aquatic systems and contribute to a cleaner environment.

Team
Project Design
Integrated Human Practices