Collaborative Research: Investigating and Improving the Production of Butanol by C. Pasteurianum for the Value-Added Conversion of Biodiesel-Derived Crude Glycerol

随着原油价格不断创新高，生物精炼成为可再生燃料和化工生产的必要选择。然而，目前缺乏能够利用多种原料进行生物转化的健壮和通用的微生物。为了显著降低石油的利用率，开发一种基于生物质的多样化工艺组合，从而为各种石油产品产生可比较的替代品，需要设计出更好的工程微生物，使其在综合生物精炼系统中发挥作用。利用生物柴油衍生的粗甘油生产丁醇是提高生物燃料工业总体可持续性的一种切实可行的方法，尤其对生物柴油工业的增长和稳定至关重要。此外，利用粗甘油作为玉米衍生原料的替代品，将为受玉米价格和可用性限制的新兴生物精炼行业提供急需的多样性和灵活性。丁醇是一种重要的生物炼制平台化学品，作为一种可再生的运输燃料具有显著的优势。使用基本无成本的原料产生更高产量的潜力加强了工业规模丁醇发酵的技术和经济可行性。本研究的目的是量化巴氏梭菌的代谢和耐受性，并设计这种细菌，以提高生物柴油衍生的粗甘油的厌氧发酵的增值转化。该项目将侧重于基础研究，以了解和改善细胞外和细胞内途径控制甘油利用和底物形成的代谢途径。拟议的研究有三个目标。第一个目的是阐明巴氏杆菌对底物和产物毒性的细胞反应。第二个目的是评估细胞膜结构和稳定性对增加底物杂质和丁醇浓度的反应。第三个目标是对巴氏杆菌进行代谢工程，以增加丁醇的产量。这项研究具有创新性，因为它提出了甘油发酵的另一种途径，其中代谢工程方法用于敲除还原途径中的基因，而不是上调氧化途径中的基因。这项研究具有潜在的变革性，因为它为微生物代谢工程开辟了新的途径，将生物炼制过程中的粗甘油副产品转化为液体运输燃料。更广泛的影响本研究项目为教育学生和其他教育工作者提供了关于替代能源、可再生资源和生物精炼的机会。该教育计划将培养研究生和本科生的技术和实验室专业知识，为他们在生物燃料行业的职业生涯做好准备，该行业目前正经历着大幅增长。计划开展各种外展活动，包括举办研讨会和示范活动，向K-12学生、教师和社区成员传授生物燃料的生产和利用知识。教育工作还将侧重于通过旨在将这些学生纳入的活动，从代表性不足的群体中招收学生。这些教育和推广活动将激发人们对科学和工程的广泛兴趣，并激励K-12学生在工程和能源行业接受高等教育和职业。拟议的外展活动还包括一个基于网络的维基网站和夏季交流计划，这将促进两个研究小组的研究生和本科生之间的合作。

0966846/0966818Taconi/BothunAs the price of crude oil continues to reach record highs, biorefining becomes a necessary option for renewable fuels and chemical production. However, robust and versatile microorganisms for bioconversion that can utilize multiple feedstocks are currently lacking. Better engineered microorganisms designed to function within an integrated biorefining system are required to significantly reduce petroleum utilization and develop a diverse portfolio of biomass-based processes that can generate comparable alternatives for a wide range of petroleum products.Utilization of biodiesel-derived crude glycerol for the production of butanol represents a tangible approach to improving the sustainability of the biofuels industry in general and is critical for the growth and stability of the biodiesel industry in particular. Furthermore, utilizing crude glycerol as an alternative to corn-derived feedstocks will provide much-needed diversity and flexibility to an emerging biorefining industry limited by the price and availability of corn. Butanol is a key biorefinery platform chemical and has significant advantages as a renewable transportation fuel. The potential to generate higher yields using an essentially no-cost feedstock strengthens the technical and economic feasibility for industrial-scale butanol fermentation.Intellectual MeritThe goal of this research is to quantify metabolism and tolerance in Clostridium pasteurianum and to engineer this bacterium to improve the value-added conversion of biodiesel-derived crude glycerol using anaerobic fermentation. This project will focus on fundamental research to understand and improve the metabolic pathways that control glycerol utilization and substrate formation from both extracellular and intracellular approaches. The proposed research has three objectives. The first objective is toelucidate the cellular response of C. pasteurianum to both substrate and product toxicity. The second objective is to evaluate cell membrane structure and stability in response to increasing concentrations of substrate impurities and butanol. And the third objective is to metabolically engineer C. pasteurianum to increase butanol production.The research is innovative because it proposes an alternative pathway to glycerol fermentation, where metabolic engineering approaches are used to knock down the genes in the reductive pathway as opposed to up-regulation of genes in the oxidative pathway. The research is potentially transformative because it opens up new approaches for metabolic engineering of microorganisms to convert crude glycerol byproducts from biorefinery operations to liquid transportation fuels.Broader ImpactsThis research project provides opportunities to educate students and other educators about alternative energy, renewable resources, and biorefining. The education plan will develop the technical and laboratory expertise for graduate and undergraduate students to prepare them for careers in the biofuels industry, which is currently experiencing substantial growth. Various outreach activities are planned, including the development of a seminars and demonstrations to educate K-12 students, teachers, and community members about biofuels production and utilization. Education efforts will also focus on the recruitment of students from underrepresented groups through activities that will target the inclusion of these students. These educational and outreach activities will stimulate broad interest in science and engineering, and inspire K-12 students to pursue higher education and careers in engineering and energy industries. The proposed outreach activities also include a web based wiki site and summer exchange program that will facilitate collaboration among graduate and undergraduate students in the two research groups.