Collaborative Research: Expanding the substrate range of Clostridium thermocellum to improve understanding of its metabolism

植物中的干物质称为木质纤维素生物质。它是世界上最丰富的可再生原料来源。将其转化为燃料和产品将对循环生物经济作出重大贡献。不幸的是，它的某些部分很难解构，这增加了成本，降低了产量。某些微生物能够分解木质纤维素。这个项目的目的是阐明一种这样的微生物的代谢。一旦了解，将努力扩大生物可以利用的底物范围。在开展研究活动的同时，两位主要研究人员将通过已建立的渠道，让K-12学生参与科学展览和暑期研究体验。天然纤维素降解生物（如C. thermocellum）的底物范围有限，这使得人们很难理解它们的代谢并设计它们用于应用目的。本研究的目的是扩大C. thermocellum的底物范围，以提高我们对其代谢的理解，从而诊断产品形成的局限性。为实现这一目标，将实现下列一系列协调一致的目标。(1)表征底物运输和中枢代谢的可逆性，以确定限制底物范围的因素。(2)结合使用固定阶段进化和基于组合多底物化学控制因子的方法，识别允许在替代底物上生长的突变。(3)使用替代生长基质来确定限制乙醇生产的因素。本提案中的工作将提高我们对非典型糖酵解形式的理解，这将扩大我们对这种普遍存在且高度保守的代谢途径的可能变化的理解。它将提高我们对生物体内控制底物范围的因素的理解，包括由于运输和代谢反应可逆性的限制。这也将使我们了解限制产物滴度的因素，这是代谢工程领域的一个核心挑战。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。

The dry matter in plants is called lignocellulosic biomass. It is the most abundant source of renewable raw material in the world. Converting it to fuels and products would contribute substantially to a circular bioeconomy. Unfortunately, parts of it are difficult to deconstruct, which increases costs and lowers yields. Certain microbes are capable of deconstructing lignocellulose. The objective of this project is to elucidate the metabolism of one such microbe. Once understood, efforts will be made to expand the range of substrates the organism can utilize. In parallel with the research activities, both primary investigators will work through established channels to engage K-12 students in science fairs and summer research experiences. The limited substrate range of natively cellulolytic organisms (such as C. thermocellum) makes it difficult to understand their metabolism and engineer them for applied purposes. The goal of this study is to expand the substrate range of C. thermocellum to improve our understanding of its metabolism to diagnose limitations to product formation. To accomplish this, the following set of coordinated aims will be pursued. (1) Characterize substrate transport and reversibility of central metabolism to determine factors that limit substrate range. (2) Identify mutations that allow growth on alternative substrates, using a combination of stationary-phase evolution and a combinatorial multi-substrate chemostat-based approach. (3) Use alternative growth substrates to determine factors that limit ethanol production. The work in this proposal will improve our understanding of atypical forms of glycolysis, which will expand our understanding of the possible variations of this ubiquitous and highly conserved metabolic pathway. It will improve our understanding of the factors that control substrate range in organisms, including limitations due to transport and metabolic reaction reversibility. It will also allow us to understand factors that limit product titer, a central challenge in the field of metabolic engineering.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.