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

植物中的干物质被称为木质纤维生物质。它是世界上最丰富的可再生原材料来源。将其转化为燃料和产品将极大地促进循环生物经济。不幸的是，它的一部分很难解构，这增加了成本，降低了收益率。某些微生物能够分解木质纤维素。这个项目的目标是阐明一种这样的微生物的新陈代谢。一旦了解，将努力扩大有机体可以利用的底物的范围。在研究活动的同时，两位初级调查人员将通过既定渠道，让K-12学生参加科学博览会和暑期研究体验。天然的纤维素分解生物(如高温假单胞菌)的底物范围有限，因此很难理解它们的新陈代谢，也很难将它们用于工程应用。本研究的目的是扩大嗜热假单胞菌的底物范围，以提高我们对其代谢的了解，从而诊断产物形成的局限性。为了实现这一目标，将实现以下一系列协调一致的目标。(1)表征底物转运和中枢代谢的可逆性，以确定限制底物范围的因素。(2)使用固定相进化和基于组合多底物恒化器的方法相结合的方法，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。这项建议中的工作将提高我们对非典型糖酵解形式的理解，这将扩大我们对这一普遍存在的高度保守的代谢途径的可能变异的理解。它将提高我们对生物体中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造成的限制。它还将使我们了解限制产品效价的因素，这是代谢工程领域的核心挑战。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势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.