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

微生物可以从生物质中产生燃料和化学物质。生物燃料有助于美国的能源安全。目前使用微生物进行生产的一个限制是微生物也使用相同的生物量来生长。经济的快速增长减少了燃料和化学品的生产。然而，控制微生物代谢的新策略可以将生长与产物形成分开。这个项目的目标是开发一个模块化的控制系统，适用于许多不同的生物体。该项目的成功将促进燃料和化学品的微生物生产，从而推动生物经济。本项目的目的是评估一种新的动态代谢控制策略。我们将使用先进的计算蛋白设计和工程来改造T7RNA聚合酶（T7RNAP），它将DNA转录成mRNA，因此它需要一个用户定义的小分子来发挥作用。这种重组的聚合酶将被组成性表达。这种诱导分子激活聚合酶，导致t7特异性基因的表达。我们将(i)确定对各种非模式宿主生物无毒且不可代谢的廉价化学品；（ii）在恶臭假单胞菌中部署动态控制电路，用于生产muconate，以改善滴度/产量/生产率指标；（iii）证明T7 RNAP电路在其他生物制造相关生物中的实用性。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

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.