Enhancing Microbial Flavonoid Production with Synthetic Biology

Enhancing Microbial Flavonoid Production with Synthetic Biology Enhancing Microbial Flavonoid Production with Synthetic Biology Flavonoids represent a diverse class of naturally occurring phytochemicals with numerous biochemical activities of benefit to human health. Several epidemiologic studies have suggested that diets rich in flavonoid intake are correlated with reduced occurrences of cardiovascular disease the current leading cause of death in the United States (accounting for over 25% of all deaths). However, as realizing such potential benefits has been suggested to rely upon relatively high doses of daily flavonoid intake, there remains a need to develop economical and scalable methods for flavonoid production. Current flavonoid production routes primarily rely upon their isolation from plants, however, the approach is saddled by low production rates/efficiencies and the need for costly extraction/purification steps. Chemical synthesis of flavonoids, meanwhile, is challenged by their complex molecular structures and the need for toxic precursors and harsh reaction conditions. Accordingly, flavonoids have emerged as attractive targets for microbial biosynthesis through metabolic engineering. Currently, however, a key bottleneck limiting microbial flavonoid biosynthesis is the low availability of precursor malonyl-CoA, due largely to the competing and growth essential role that it plays in fatty acid biosynthesis. The overall goal of the proposed research is to apply synthetic biology tools to create a novel and modular genetic system capable of enhancing malonyl-CoA availability and thus flavonoid biosynthesis. We hypothesize that flavonoid production will be enhanced through the engineering and use of a dynamically actuated metabolite valve capable of efficiently directing malonyl-CoA flux between fatty acid and flavonoid biosynthesis pathways, thereby effectively decoupling the processes biomass growth and flavonoid production. Accordingly, constructed using synthetic biology tools including recombination, CRISPRi, and targeted proteolysis both individually and in parallel the proposed work first seeks to develop and then rigorously evaluate the dynamic behavior of a malonyl-CoA valve. Following its characterization and optimal tuning, said valve will be applied to improve the de novo biosynthesis of pinocembrin and naringenin, model flavanones with important heart health benefits. This project will furthermore support the PIs continued development as an investigator in the fields of metabolic engineering and synthetic biology. This project will serve as the ideal platform for broadening the focus of the PIs research to include the biosynthesis of specialty products of greater significance to human health. This project will support the PIs ultimate career goals of understanding and demonstrating how metabolic complexity of microbial systems can be harnessed and engineered to solve key challenges of the utmost societal importance.