This research aims to produce new technologies that will enable the synthesis of valuable commodity chemicals and pharmaceuticals from microbes. These engineered microbes are commonly called ?microbial chemical factories,? and they are now enabling the biological production of valuable chemicals that have traditionally been obtained through oil refining. The emerging field of synthetic biology is enabling the construction of new metabolic pathways, and this is rapidly broadening the scope of chemicals that can be produced by microbial cell factories. In this research, a novel de novo pathway to 2-pyrrolidone will be produced and optimized. This valuable target chemical is used to produce over 700 differently substituted pyrrolidones that have uses ranging from ?green? solvents to pharmaceuticals. The new de novo pathway to 2-pyrrolidone will be constructed using a model-guided platform involving molecular docking and dynamics simulations in order to engineer existing enzymes to work with non-native substrates to ultimately produce 2-pyrrolidone. This research will also develop a model-guided platform for installing this de novo pathway in a microbial host and ensure the pathway is highly utilized. In this case, the bacterium Clostridium cellulolyticum H10 will be used as the host. This organism is of interest due to its ability to consume cellulosic plant material as a substrate, ensuring the resulting production of 2-pyrrolidone will be renewable and ultimately cost-efficient. To ensure high usage of the 2-pyrrolidone de novo pathway, metabolic engineering strategies will be developed through a new technique involving an established method of ?genome-scale metabolic flux modeling.? While this project will engineer a strain of C. cellulolyticum H10 to produce 2-pyrrolidone from cellulose, the tools developed in this project will apply broadly. Ultimately, this toolset will help enable the production of any targeted chemical from any host cell. These developments will have profound impacts by (i) transitioning the origin of many vital commodity chemicals and pharmaceuticals from non-renewable oil refining to renewable production from sources such as plant material and sunlight, (ii) dramatically increasing the role of the biotechnology industry in the commodity chemicals market, and (iii) stimulating new startup ventures related to commodity chemicals and pharmaceutical production through synthetic biology.
