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Team:UC Davis/Project/Strain
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The strain section of our project focuses on the degradation of ethylene glycol, a chemical that is metabolized to oxalic acid further downstream. Oxalic acid is toxic to the kidney and fatal to the organism. We found an E. coli mutant from the University of Barcelona in Barcelona, Spain, that is able to grow solely on ethylene glycol, one of the two products created during PET degradation [1]. The scientists in Barcelona created these mutants through directed evolution – a process that selects for the most fit in a group under increasing conditions of ethylene glycol. From this, they learned that the main contributors in the degradation were propanediol oxidoreductase and glycolaldehyde dehydrogenase. These two enzymes are expressed at low levels in MG1655 but not at all in DH5α. We want to overexpress these enzymes in MG1655 through directed strain engineering. Rather than take the enzymes from the Barcelona strain, we want to clone the enzymes from the MG1655 and be able to control them ourselves, also known as rational strain engineering. | The strain section of our project focuses on the degradation of ethylene glycol, a chemical that is metabolized to oxalic acid further downstream. Oxalic acid is toxic to the kidney and fatal to the organism. We found an E. coli mutant from the University of Barcelona in Barcelona, Spain, that is able to grow solely on ethylene glycol, one of the two products created during PET degradation [1]. The scientists in Barcelona created these mutants through directed evolution – a process that selects for the most fit in a group under increasing conditions of ethylene glycol. From this, they learned that the main contributors in the degradation were propanediol oxidoreductase and glycolaldehyde dehydrogenase. These two enzymes are expressed at low levels in MG1655 but not at all in DH5α. We want to overexpress these enzymes in MG1655 through directed strain engineering. Rather than take the enzymes from the Barcelona strain, we want to clone the enzymes from the MG1655 and be able to control them ourselves, also known as rational strain engineering. | ||
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Revision as of 05:41, 29 September 2012
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Background
Rationale
Our main goal is for the E. coli to be able to live off PET as the sole carbon source. In order to do this, it must be able to sequester the carbon into its metabolism. In the diagram below, the ethylene glycol binds to the glycolaldehyde reductase to form glycolaldehyde. After, the glycolaldehyde attaches to the glycolaldehyde dehydrogenase to form glycolate. The glycolate goes in to the metabolism via further reactions with glycolate dehydrogenase and malate synthase. The (S)-malate is the final product that is incorporated in to the citric acid (TCA) cycle. As the citric acid cycle propagates, more energy is made for the cell, allowing growth and self-sufficient development on PET.
Our Strain
References
2. Bsc, S. N. and Gp Savage Bsc(hons), PhD, Nz Reg NutR. (1999), Oxalate content of foods and its effect on humans. Asia Pacific Journal of Clinical Nutrition, 8: 64–74. doi: 10.1046/j.1440-6047.1999.00038.x
