Engineering microbial metabolic pathways allows the production of chemicals, biofuels, materials, and pharmaceuticals from renewable and low-cost feedstocks. For this technology to be scalable and economically viable, engineered microbial hosts must be able to continuously balance the pathway metabolism to cope with high titer and high yield production. Dynamic metabolic regulation systems (DMRSs) are synthetic biological components that provide dynamic control to the concentration of pathway enzymes and metabolites (the chemicals on which enzymes act). This study will provide a systematic and quantitative understanding of how DMRSs change the dynamic behavior of metabolic pathways and how DMRSs improve pathway productivity. The knowledge will enable researchers to design metabolic control systems more easily, effectively, and reliably, improving productivities, yields and robustness for a variety of metabolic pathways and making the microbial production of chemicals, pharmaceuticals, biofuels, and material more economically viable. This project will provide tools that are useful for studying the effect of dynamics on natural metabolic pathways, revealing the evolutionary significance of natural regulatory systems. These tools can be further extended to regulate a variety of complex synthetic systems, such as chemical-controlled, contaminant-cleaning microbes. This project will also provide educational and training opportunities for K-12 students and teachers from local public schools with a high percentage of underrepresented students through a series of activities including "Hot Topic Workshops" and "Lab Research Experience" These educational and training programs will promote students' interests in cutting-edge science and to develop basic skills for research in systems and synthetic biology.
Synthetic regulatory systems that provide dynamic control on metabolic pathways (referred to as DMRSs) are very powerful tools in microbial engineering for the production of chemicals from complex metabolic pathways. Previous proof-of-concept studies have demonstrated that DMRSs were able to significantly enhance both pathway productivities and yields. However, due to the complex protein-metabolite/protein-DNA interactions, construction of effective DMRSs is challenging and often needs to test a large number of parameters and regulatory topologies (e.g. upregulation v.s. downregulation, long-range feedback vs. short-range feedback). For DMRSs to work robustly and to be applicable to a broad range of pathways, this project aims to develop a systematic understanding of metabolic dynamics and of how metabolic dynamics affects pathway productivities. Using a combination of experimental and theoretical strategies, including biosensor-enabled ensemble-fluorescence measurement, single cell imaging, chromatographic metabolite quantifications, and kinetic models, this project will systematically characterize metabolic dynamics, metabolic fluctuation, and cell-to-cell metabolic variation under various regulatory topologies.
