Team:NRP-UEA-Norwich/Description
THE BACKGROUND
Colon cancer is the second most common cause of cancer death in England and Wales, with 30,000 new cases diagnosed every year and a registered cause of death in half that number.
Recent studies have suggested that high dietary intake of resistant starch may reduce colon cancer and inflammatory bowel disease. Resistant starches escape digestion in the small intestine and are fermented by microbiota in the colon. A small proportion of these colonic bacteria produce short chain fatty acids such as butyrate, which can activate apoptosis in colon cancer cells.
Our project is focused on increasing the amount of butyrate in the colon.
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OUR SOLUTION
We are taking two approaches to increase butyrate levels in the colon.
The first approach is to produce modified starches that might be consumed as a prebiotic. We will test the capability of various putative acyltransferases to transfer acyl groups such as butyryl to the alpha-1,4 carbohydrates produced in bacteria and plants. This will involve modelling carbohydrate molecules to find optimal branching and solubility states.
The second approach is to transfer the butyrate biosynthetic pathway into culturable bacterial species with the long-term aim of producing a probiotic.
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MODELLING AND SOFTWARE
We used glycogen as a model system to gain a better understanding of the parameters involved in carbohydrate structure, branching and modification. We worked with an existing mathematical model to create structural predictions of carbohydrates.
We developed software to model the putative changes in glycogen structure depending on the position of the butyrate group in the glucose molecule, as growth or branching might be disrupted.
We also built 3D models of carbohydrates to show the importance of the branching degree and number of tiers in the final molecule conformation, and we 3D printed them!
Finally, we followed a deterministic approach to model the enzyme kinetics of glycogen branching and debranching.
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RESULTS
As of 18th September 2015, we have assembled parts and developed methods for testing enzymes that might make enzymatic modifications to glycogen in bacteria and to starch in plants. We are able to detect changes in branching but have not yet detected addition of new groups. This might be below the detection limit of our current assays – we are currently considering different methods for measuring acylation.
In plants we have been able to direct our putative modification enzymes to the chloroplast, the site of starch synthesis, and also to show that the quantity of starch produced is not affected. We can now progress to stable transformation of plants to allow us to obtain sufficient quantities of modified starch for analysis. We have also cloned the butyrate synthesis pathway, which requires eight coding sequences. We are currently troubleshooting a method to monitor the production of the butyryl-CoA and butyrate metbolites.
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PARTS
Our project required us to make several groups of parts including parts for engineering bacterial glycogen, parts for engineering plant starch, and parts to make a butyrate biosynthesis pathway.
As well as making new parts, we used several parts from the Registry of Biological Parts, especially those parts for plants developed by the 2014 NRP-UEA iGEM team. We have contributed to the further characterisation of those parts in the Registry. These include BBa_K1467400, BBa_K1467101 and BBa_K1467204.
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PRACTICES
There are significant ethical considerations with the production of bioengineered food supplements. By talking to Cancer charities, people who run trials to test the effects of diet on health, and the public we found out that while the genetic engineering of food is still controversial in Europe, opinions may differ when the reasons for engineering are for the prevention of cancer. We went on to design a theoretical feeding trial that would enable us to test the efficacy of our products in humans.
Additionally, we wanted to find a way to work with experienced creative writers to open the narrative on bioengineering to a new audience. We held a Science and Writing workshop, at which scientists and poets worked together to produce poems inspired by synthetic biology and our project.
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