Leveraging the unique metabolism of Megasphaera elsdenii for metabolic engineering to medium and long chain organic acids for use in jet fuels and biomaterials

包括航空燃料在内的可持续燃料和材料的生物生产是一个紧迫而紧迫的问题。该提案中开发的知识和技术通过创造碳中性燃料来缓解气候变化。这些生物技术通过创造对可持续种植的植物生物量的需求来促进农村经济发展，减少对化石碳的依赖，并提高美国的经济竞争力。该项目确定了长链有机酸生物合成途径，评估了其在不同生长条件下的活性，开发了先进的遗传工具，并利用这些工具改善己酸的生产，这是一种可持续的航空燃料前体。该项目与克拉克县公立学校系统五年级学生的“女性”科学项目有关。作为一项新举措，该项目与佐治亚大学教育学院合作，为职前实习教师提供STEM教育联合培训。微生物的代谢多样性才刚刚开始被理解，并且代表了一个未开发的途径来源，以产生在现有模式微生物中难以或不可能设计的化合物。Megasphaera elsdenii具有凝聚乙酰辅酶a以高通量和高产率高效生成C4到C8化合物的天然能力，使其成为研究这些途径并设计用于生产燃料，化学品和生物材料的生物体的引人注目的平台。该项目确定了长链有机酸生物合成途径，评估了其在不同生长参数下的活性，开发了先进的遗传工具，并利用这些工具提高了己酸的通量，这是一种可持续的航空燃料前体。允许基因删除和异源表达的基本遗传工具集用于删除预测参与链延伸的每个非必需基因，并分析对发酵产物的影响，以便了解负责每个步骤的基因。M. elsdenii具有罕见的以乳酸为碳源厌氧生长的能力，以及共同利用乳酸和乙酸的能力。该项目包括野生型的生理研究和突变分析，以剖析厌氧乙酸利用和乳酸生物转化的途径。该项目为将植物糖转化为长链分子和生物处理的替代方法奠定了基础，例如共培养或顺序发酵，其中一种生物将糖或生物质转化为乳酸，而经过工程处理的M. elsdenii将乳酸转化为更高价值的产品，从而实现广泛的厌氧微生物发酵策略。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。

Bioproduction of sustainable fuels and materials, including aviation fuel, is a pressing and urgent issue. The knowledge and technologies developed in this proposal, mitigates climate change by creating carbon-neutral fuel. These biotechnologies increase rural economic development by creating a demand for sustainably grown plant biomass, decrease reliance on fossil carbon, and increase U.S. economic competitiveness. This project identifies long chain organic acid biosynthetic pathways in Megasphaera elsdenii, evaluates their activity under varying growth conditions, develops advanced genetic tools, and leverages those tools to improve the production of hexanoic acid, which is a sustainable aviation fuel precursor. The project is linked to a “women” in science program for 5th graders in the Clarke County public school system. As a new initiative, the project institutes joint training in STEM education for preservice student teachers, in conjunction with the University of Georgia College of Education. The metabolic diversity of microorganisms is only beginning to be understood and represents an untapped source of pathways to produce compounds that are difficult or impossible to engineer in existing model microbes. Megasphaera elsdenii, has the native ability to condense acetyl-CoA to efficiently generate C4 to C8 compounds at high flux and high yield, making it a compelling platform for the study of these pathways and to engineer the organism for production of fuels, chemicals and biomaterials. This project identifies long chain organic acid biosynthetic pathways in Megasphaera elsdenii, evaluates their activity under varying growth parameters, develops advanced genetic tools, and leverages those tools to improve flux to hexanoic acid, which is a sustainable aviation fuel precursor. A basic genetic toolset that allows gene deletion and heterologous expression is used to delete every nonessential gene predicted to be involved in chain elongation and analyze the impact on fermentation products to enable understanding of the genes responsible for each step. M. elsdenii has the rare ability to grow anaerobically on lactic acid as a carbon source as well as the ability to co-utilize lactic and acetic acids. The project involves physiological studies on wild type as well as mutational analysis to dissect the pathway(s) for anaerobic acetic acid utilization and lactic acid bioconversion. This project lays the foundation for engineering plant sugars to longer chain molecules and for alternate approaches to bioprocessing, such as the co-culture or sequential fermentation in which one organism converts sugars or biomass to lactic acid and an engineered M. elsdenii converts the lactic acid to a higher value product enabling strategies for a broad range of anaerobic microbial fermentations.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.