Metabolic Engineering of Escherichia coli for the Synthesis of Defined Polyhydroxyalkanoates from Unrelated Feedstocks.

摘要

The global challenges of sustainability and environmental preservation demand solutions that tap into renewable resources and avoid the need for petroleum derived fuels and chemicals. This thesis addresses the above challenges by applying synthetic biology techniques to the synthesis of a naturally occurring class of polyesters known as polyhydroxyalkanoates (PHA). PHA are synthesized by microorganisms as a form of carbon and energy storage. Today, PHA are produced at small-scale industrial facilities around the world. However, PHA face a number of obstacles to commercial adoption. First, PHA are sold at a significant premium to traditional plastics. Second, while PHA can be incorporated directly into various plastic products, the physical properties of the current generation of PHA limit usage to a narrow marketplace. To reduce the cost of production and expand the applications of PHA, strategies were developed for synthesizing novel PHA from inexpensive, renewable feedstocks.;This thesis explores how synthetic biology can be applied to renewable plastic production. Chapter 2 reviews existing biosynthetic routes to PHA from unrelated carbon sources and updates the diverse list of monomers incorporated into PHA since 1995. Next, two alternative strategies for producing PHA monomers are proposed – involving either fatty acid metabolism or polyketide biosynthesis. Chapter 3 describes a thioesterase-based strategy for producing mcl-PHA with a defined composition that circumvents the iterative nature of fatty acid metabolism. Chapter 4 describes PHA production involving type I polyketide synthases, a class of multi-functional enzymes that synthesize their products in a predictable, assembly-line fashion. Previous research has demonstrated the possibility of modifying polyketide synthases to produce novel products. Therefore, a strategy was explored for the polyketide-based production of novel PHA precursors. Finally, Chapter 5 discusses the potential for further improvements in PHA biosynthesis. In summary, this thesis explores routes to tailor-made PHA that begin with inexpensive, renewable substrates in hopes of both reducing the cost of industrial PHA biosynthesis and expanding the applications of this class of biodegradable polyesters.