Systems metabolic engineering of Escherichia coli for the production of platform chemicals used as monomers for polymer synthesis

Abstract

The establishment of a sustainable chemical industry through systems metabolic engineering is becoming essential to overcome serious environmental issues that are now being faced. Among various chemicals and materials that can be produced by microbial cell factories, bio-based polymers has recently received considerable attention to replace current petroleum-driven plastics. To make this bioprocess to be more industrially competitive, the development of microbial strains capable of efficiently producing plastic precursors must be accompanied. In this dissertation, Escherichia coli was metabolically engineered to produce platform chemicals used for polymer synthesis: acrylic acid, propionic acid, and putrescine. For the production of acrylic acid, the novel $\beta$-alanine route was employed. After the validation and construction of the designed pathway, E. coli was metabolically engineered to produce acrylic acid from glucose by introducing the constructed pathway and optimizing metabolic pathways. Microbial production of propionic acid was also attempted by employing the β-alanine route. In specific, E. coli was engineered to produce propionic acid by applying metabolic engineering strategies, including screening of efficient enzymes, deletion of competing pathways, optimization of the ppc gene expression level, and application of propionyl-CoA biosensor for identifying gene knockdown targets. Last, for the enhanced production of putrescine, the previously constructed XQ52 strain was subjected to adaptive laboratory evolution to increase its tolerance on putrescine. After 100 days of serial transfers, evolved strains tolerating up to high-level of putrescine were obtained, followed by the screening of best-performing variants. As a result, the evolved XQ52 strain capable of producing putrescine much above its native tolerance was developed. Taken together, systems metabolic engineering tools and strategies were successfully applied to demonstrate the construction of E. coli producing various plastic precursors. It is expected that the strategies described here will be helpful for the development of other microbial strains producing platform chemicals.