EAGER: NSF/ABF Dynamic Control of Metabolism Using Redesigned Proteins

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Microbes can create fuels and chemicals from biomass. Bio-based fuels help contribute to energy security in the US. One current limitation to using microbes for production is that the microbes also use the same biomass to grow. Rapid growth reduces fuel and chemical production. However, new strategies for controlling the microbial metabolism could separate growth from product formation. The objective of this project is to develop a modular control system to be applied across many different organisms. Success of this project will enhance microbial production of fuels and chemicals, thus advance the bioeconomy. The objective of this project is to evaluate a novel strategy for dynamic metabolic control. We will use advanced computational protein design and engineering to remodel T7 RNA polymerase (T7RNAP), which transcribes DNA into mRNA, such that it requires a user-defined small molecule to function. This remodeled polymerase would be constitutively expressed. This induction molecule activates the polymerase, resulting in expression of T7-specific genes. We will (i) identify inexpensive chemicals that are non-toxic and non-metabolizable for diverse non-model host organisms; (ii) deploy dynamically controlled circuits in Pseudomonas putida for production of muconate to show improved metrics of titer/yield/productivity; and (iii) demonstrate the utility of T7 RNAP circuits for other biomanufacturing-relevant organisms.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.