Isobutene is a short chain gaseous hydrocarbon that is used in fuel additives, antioxidants, detergents, and butyl-rubbers with a global market value of approximately 22 Billion USD/year. Isobutene is currently produced through steam cracking of crude oil. This process is energetically intensive and emits large quantities of greenhouse gasses. To reduce dependence on extraction and refinement of crude oil, enhancing biological production of petrochemicals is important. Biological production of isobutene has been demonstrated previously, but economically viable production faces many hurdles. Seeking to improve this viability, a new isobutene producing microbe was engineered. E. coli MG1655 was genetically engineered to produce isobutene through expression of two enzymes, M3K (P. torridus) & MVD (S. cerevisiae). This engineered E. coli was able to produce isobutene during growth on glucose and wastewater, but production rates remain low and further optimization is needed. To identify potential genes to target for genetic engineering that will increase production rates, the transcriptomic response of the engineered E. coli to several stressful conditions was analyzed. The stress responses to solvent exposure showed conserved responses of increasing membrane stability in response to solvent interactions and protein repair. Furthermore, due to interest in the potential for biofuel production in space, the effects of spaceflight on the gene expression of cell cultures aboard the International Space Station (ISS) were assessed and compared to controls grown on Earth. Initially, cell cultures aboard the ISS had a strong expression of stress response systems related to freezing. However, this response faded and there was increased expression of systems that would increase mutation rates. In both solvent and spaceflight stress experiments there was upregulation of chaperones that repair damaged proteins and increased saturation of membrane lipids to maintain membrane integrity. These experiments provide potential targets for future engineering efforts to engineer a more robust and stress tolerant microbe for enhanced production of isobutene.
|Advisor:||Briggs, Brandon R.|
|Commitee:||Duddleston, Khrystyne N., Milligan-Myhre, Kathryn|
|School:||University of Alaska Anchorage|
|School Location:||United States -- Alaska|
|Source:||MAI 81/2(E), Masters Abstracts International|
|Keywords:||Biofuels, CRISPR, Microbiology|
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