Difference between revisions of "Team:Manchester-Graz"

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<h1 style="margin-bottom: 60px;"> Project Outlines</h1>
 
<h1 style="margin-bottom: 60px;"> Project Outlines</h1>
 
 
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<p style="text-align:justify">This year Manchester-Graz team is trying to develop a whole new system for L-DOPA/dopamine treatment by regulating the pathway in an innovative way.</p><div id="pictureright"> <img src="https://static.igem.org/mediawiki/2015/b/b3/Manchester-Graz_Human-Gutbacteria.jpg" alt="human gut bacteria" width="400"> <br> <b> Fig 1 </b>Administration of dopamine via gut bacteria</div>
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<p style="text-align:justify">iGEM Manchester-Graz’s objective as a team is to find a better way to treat and alleviate the symptoms of Parkinson’s disease (PD) through the use of synthetic biology. Degradation of dopaminergic neurons and therefore low levels of dopamine is the main cause of Parkinson’s disease (PD), for which the current treatment involves oral doses of L-DOPA (or levodopa), which unlike dopamine itself is able to cross the blood-brain-barrier. Within the brain L-DOPA is enzymatically converted into dopamine and therefore able to relieve many of the motor symptoms of PD (Figure 1).</p><div id="pictureright"> <img src="https://static.igem.org/mediawiki/2015/b/b3/Manchester-Graz_Human-Gutbacteria.jpg" alt="human gut bacteria" width="400"> <br> <b> Fig 1 </b>Administration of dopamine via gut bacteria</div>
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<p style="text-align:justify">Low dopamine levels are linked to Parkinson’s disease, Alzheimer’s disease and several other health problems. The current treatment involves L-DOPA which crosses the blood-brain barrier, and is then enzymatically converted into dopamine. Studies show that Escherichia coli, Bacillus subtilis, Bacillus cereus, Bacillus mycoides, Proteus vulgaris, Serratia marcescens, and Staphylococcus aureus synthesize dopamine and that varying levels of dopamine in the gut can have neurological effects. Having bacteria in the gut producing dopamine would abolish the need for traditional medication, and only a single dose of culture in probiotics containing strains of self-regulating DOPA-producing bacteria will be needed. This is especially convenient for elderly people suffering from these diseases as there will be no such problem as an overdose or missing a medicine intake. This bacteria-based controlled drug delivery system incorporated could potentially improve the lifestyle of patients (Fig 1).</p>
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<p style="text-align:justify">Our aim is to take the first steps in the development of a novel technology of drug delivery by developing self-regulating drug-producing bacterial strains that, in the future, could be incorporated into patients’ gut microflora to secrete medicines directly inside the body. To control the bacterial L-DOPA production, we plan to develop a multidimensional cell density dependent auto-regulation system that could also be used to control other multistep enzyme pathways.</p>
 
<div id="pictureleft"><img src="https://static.igem.org/mediawiki/2015/e/ef/Manchester-Graz_Pathway.jpg" alt="Pathway" width="300"> <br><b> Fig 2 </b>Novel dopamine expressing pathways </div>
 
<div id="pictureleft"><img src="https://static.igem.org/mediawiki/2015/e/ef/Manchester-Graz_Pathway.jpg" alt="Pathway" width="300"> <br><b> Fig 2 </b>Novel dopamine expressing pathways </div>
  
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<p style="text-align:justify">We plan to develop an innovative multidimensional cell density dependent auto-regulation system that could be used to control multistep enzyme pathways. L-DOPA is a good example as it is nowadays produced in bacterial cultures to cover the demand needed all over the world. The aim is to make the first steps in the development of a novel technology of drug delivery by developing self-regulating drug producing bacterial strains that, in the future, could be incorporated into patients’ intestinal and gut microflora to secrete medicines directly inside patients’ bodies in the response to any physiological changes, in particular low levels of dopamine. </p>
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<p style="text-align:justify">The Manchester section of the team are working on L-DOPA and dopamine biosynthesis in E. coli BL21 and Nissle 1917 via various enzyme pathways. The focus is on three enzyme pathways, the main one being the conversion of L-tyrosine to L-DOPA via tyrosine hydroxylase and tyrosinase. In addition we are also synthesising dopamine in two different ways using aromatic amino acid decarboxylase, cytochrome P450 and transaminase. Although the primary goal would be to implement the L-DOPA synthesis within patients, we also aim to create a greener, more efficient way of the industrial synthesis of each of the above products using our modified bacteria. (Figure 2) </p>
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<p style="text-align:justify">Team Manchester is focusing on dopamine and L-DOPA biosynthesis. Two biochemical pathways will be worked on (Fig 2).</p>
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<p style="text-align:justify">The team will identify the nature occurring enzymes for the biosynthetic pathways of L-DOPA and dopamine. Once biochemical pathways are proved to be credible, the genes of interest will be cloned into E.coli and correct synthesis will be further analyzed. <br><br></p>
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<p style="text-align:justify">Graz-located team will use two quorum sensing based mechanisms for an autoregulated and time shifted consecutive induction of protein expression. This is first demonstrated and visualized on the example of fluorescent protein synthesis (Fig3). The fluorescent protein could then be exchanged with genes for the L- DOPA production in a way that at low cell density levels phenylalanine synthesis will get channeled.</p>
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<p style="text-align:justify">The Graz section are using two quorum sensing based mechanisms for an auto-regulated and time shifted consecutive induction of protein expression, first demonstrated using fluorescent protein synthesis (Figure 3). The fluorescent protein could then be exchanged with genes for the L- DOPA production such that at low cell density levels tyrosine synthesis will be channeled. After a certain biomass is reached actual L-DOPA production will be induced. </p>
 
<div id="pictureright"><img src="https://static.igem.org/mediawiki/2015/4/4f/Manchester-Graz_Cell_density.jpg" alt="cell%20density" width="400"><br><b>Fig 3 </b> Fluorescent protein synthesis dependent on different levels of cell density</div>
 
<div id="pictureright"><img src="https://static.igem.org/mediawiki/2015/4/4f/Manchester-Graz_Cell_density.jpg" alt="cell%20density" width="400"><br><b>Fig 3 </b> Fluorescent protein synthesis dependent on different levels of cell density</div>
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<p style="text-align:justify">After a certain biomass is reached actual L-DOPA production will be induced. As a future outlook this system could be further utilized to activate suicide genes in E.coli to avoid possible overgrowth of the native intestinal flora. We want to show proof of concept not only in E. coli strains, typically used in academic research, but also in strains that naturally occur in the guts. Therefore we plan to implement our system in E.coli Nissle 1917. Even though we cannot regulate the proliferation of our engineered strain yet, it allows us to provide an outlook of a possible application as a self-regulating drug mill in the GI- tract, that might be used in medicine in a couple of years. </p>
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<p style="text-align:justify">Looking to the future this system could be further utilized to activate suicide genes in E.coli to avoid possible overgrowth of the native intestinal flora, which we aim to show proof of concept in both strains common to academic research as well as probiotics, specifically BL21 and Nissle 1917 respectively. Even though we cannot regulate the proliferation of our engineered strain yet, it allows us to provide an outlook of a possible application as a self-regulating drug dispensing system in the GI tract, which may have clinical applications in the future.</p>
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<p style="text-align:justify">Other potential applications for this project include in situ cell density measurement directly inside the bioreactor. Furthermore, this regulatory system allows human independent induction of protein expression by exchanging the fluorescent protein genes with the corresponding gene of interest. From an economical point of view this system circumvents the issue of expensive inducers like IPTG. </p>
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<p style="text-align:justify">A main part of our project is also thinking about ethical problems, sociological aspects and human practice. These topics will be discussed in detail with high school students after talking about Synthetic Biology. In surveys, questions about public opinion concerning Synthetic Biology and genetic engineering will be asked. Survey data constructed by these discussions will be compared between Austria and the UK.
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<p style="text-align:justify">Throughout the course of the project we also aim to shape our actions in accordance to the opinions of academics, charities, industry leaders and the public. This includes what implications our project will have for patients both economically and in terms of quality of life and whether the real world implementation of this technology is feasible. We will also compare ethical opinions throughout outreach about our project and synthetic biology in general across the two countries our team spans and assess this in a wider sociological context. We feel as a team that the issues and human practices surrounding our project are just as important as the project itself and as such we endeavour to tackle a wide selection of concerns.</p>
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Our project will make the first step to provide patients with drug-independent, novel- delivery and alternative-pathway technology to tackle neurological diseases. Additionally it is designed to allow a vast public outreach to show people advantages and benefits of Synthetic Biology. </p>
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Revision as of 17:17, 14 July 2015

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iGEM Manchester - Graz Project

Project Outlines

iGEM Manchester-Graz’s objective as a team is to find a better way to treat and alleviate the symptoms of Parkinson’s disease (PD) through the use of synthetic biology. Degradation of dopaminergic neurons and therefore low levels of dopamine is the main cause of Parkinson’s disease (PD), for which the current treatment involves oral doses of L-DOPA (or levodopa), which unlike dopamine itself is able to cross the blood-brain-barrier. Within the brain L-DOPA is enzymatically converted into dopamine and therefore able to relieve many of the motor symptoms of PD (Figure 1).

human gut bacteria
Fig 1 Administration of dopamine via gut bacteria

Our aim is to take the first steps in the development of a novel technology of drug delivery by developing self-regulating drug-producing bacterial strains that, in the future, could be incorporated into patients’ gut microflora to secrete medicines directly inside the body. To control the bacterial L-DOPA production, we plan to develop a multidimensional cell density dependent auto-regulation system that could also be used to control other multistep enzyme pathways.

Pathway
Fig 2 Novel dopamine expressing pathways

The Manchester section of the team are working on L-DOPA and dopamine biosynthesis in E. coli BL21 and Nissle 1917 via various enzyme pathways. The focus is on three enzyme pathways, the main one being the conversion of L-tyrosine to L-DOPA via tyrosine hydroxylase and tyrosinase. In addition we are also synthesising dopamine in two different ways using aromatic amino acid decarboxylase, cytochrome P450 and transaminase. Although the primary goal would be to implement the L-DOPA synthesis within patients, we also aim to create a greener, more efficient way of the industrial synthesis of each of the above products using our modified bacteria. (Figure 2)

The Graz section are using two quorum sensing based mechanisms for an auto-regulated and time shifted consecutive induction of protein expression, first demonstrated using fluorescent protein synthesis (Figure 3). The fluorescent protein could then be exchanged with genes for the L- DOPA production such that at low cell density levels tyrosine synthesis will be channeled. After a certain biomass is reached actual L-DOPA production will be induced.

cell%20density
Fig 3 Fluorescent protein synthesis dependent on different levels of cell density

Looking to the future this system could be further utilized to activate suicide genes in E.coli to avoid possible overgrowth of the native intestinal flora, which we aim to show proof of concept in both strains common to academic research as well as probiotics, specifically BL21 and Nissle 1917 respectively. Even though we cannot regulate the proliferation of our engineered strain yet, it allows us to provide an outlook of a possible application as a self-regulating drug dispensing system in the GI tract, which may have clinical applications in the future.

Throughout the course of the project we also aim to shape our actions in accordance to the opinions of academics, charities, industry leaders and the public. This includes what implications our project will have for patients both economically and in terms of quality of life and whether the real world implementation of this technology is feasible. We will also compare ethical opinions throughout outreach about our project and synthetic biology in general across the two countries our team spans and assess this in a wider sociological context. We feel as a team that the issues and human practices surrounding our project are just as important as the project itself and as such we endeavour to tackle a wide selection of concerns.