CAREER: Engineering Non-Growth Metabolism for High-Yield Biochemical Production

职业：工程非生长代谢以实现高产生化生产

• 批准号: 1452549
• 负责人: Keith Tyo
• 金额: $ 50万
• 依托单位: Northwestern University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-03-01 至 2021-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1452549&HistoricalAwards=false
• 关键词: CAREER; Engineering; Non; Growth; Metabolism

1452549 Tyo, Keith E. Microbes are a promising catalyst to convert renewable resources such as sugars and non-food biomass into fuels and chemicals that are essential to our society. This proposal addresses a key challenge to realizing microbial catalysts, namely increasing the rate and efficiency that a microbial catalyst can produce the fuel or chemical. This proposal will substantially improve microbial catalysts by investigating the underlying enzyme regulation that limits catalyst productivity. While the regulation of many enzymes has been studied in isolation, the systems-level, condition-dependent regulation of enzymes has proved difficult to elucidate, but this understanding will be essential to engineering high productivity microbial catalysts. If successful, this proposal would impact the biomanufacturing competitiveness of the United States by reducing production costs of a wide range of drop-in replacements for diesel, jet fuel, and gasoline, as well as chemicals used to make plastics, preservatives, flavors and fragrances, and many other consumer products. The proposed work will also train students at the undergrad, master and doctoral levels through research, a new course in Global Health and Biotechnology and a certificate program in Sustainability and Global Health. The certificate will provide future biological engineers with an integrated understanding of biotechnology, challenging societal problems, and tools for market analysis and risk. The certificate will be piloted in the current proposal as a master?s program. This program will result in greater STEM educational infrastructure and promote interaction of under-represented minorities in low-income countries with STEM trainees. This will benefit our global partners through increased scientific activity and collaboration, technoeconomic analysis of country-specific societal challenges, as well as our society by training globally minded engineers.The rationale for the proposed work is to enable high flux metabolism in the absence of cell growth for highly productive non-growth-associated biomanufacturing processes. Developing cells with fast, non-growth product metabolism would remove a major barrier to a thriving biomanufacturing economy, by optimizing substrate conversion to product without sacrificing substrate consumption for cell growth. However, many biochemical products are growth-coupled, and in general non-growing cells have low metabolic rates. The overall objective of this proposal is to identify allosteric regulation and post-translational modification (enzyme-level regulation) that represses glycolytic metabolism in non-growth conditions. The central hypothesis is that enzyme-level regulation dominates metabolic downregulation in stationary phase. The proposed work will launch two new methods for engineering and characterizing metabolic regulation and developing a systems-level understanding of non-growth central carbon metabolic regulation. The proposal will generate minimal cells at the proteomic-level, circumventing problems with genetic deletions. The proposal will also develop a method for rapid, targeted degradation of proteins to engineer bioconversions with near optimal yields by knocking down byproduct enzymes. The method will enable new biological studies, as it will be useful for making conditional mutants to perturb biological systems in new ways. The second method will identify rate-limiting enzymes, based on thermodynamics, and thus focus engineering efforts on specific enzymes. The developed workflow will determine if reactions are near equilibrium (non-rate-limiting) or away from equilibrium (rate limiting) for a broad range of industrially relevant conditions. The novel workflow will be deployed to study central carbon metabolic regulation to garner a systems-level perspective on regulation of organic acid and terpene production.This CAREER award by the Biotechnology and Biochemical Engineering Program of the CBET Division is co-funded by the Systems and Synthetic Biology Program of the Division of Molecular and Cellular Biology.

1452549 Tyo，Keith E.微生物是一种很有前途的催化剂，可以将糖和非食品生物质等可再生资源转化为我们社会必不可少的燃料和化学品。这一建议解决了实现微生物催化剂的一个关键挑战，即提高微生物催化剂生产燃料或化学品的速度和效率。这项提议将通过研究限制催化剂生产率的潜在酶调节来显著改进微生物催化剂。虽然许多酶的调控已经被孤立地研究，但系统水平的、依赖于条件的酶调控被证明是很难阐明的，但这种理解将是设计高效微生物催化剂所必需的。如果成功，这项提议将通过降低柴油、喷气燃料和汽油的各种替代产品以及用于制造塑料、防腐剂、香料和香料和许多其他消费品的化学品的生产成本，影响美国的生物制造业竞争力。拟议的工作还将通过研究、一门新的全球卫生和生物技术课程以及一项可持续发展和全球卫生证书计划来培训本科生、硕士和博士生。该证书将为未来的生物工程师提供对生物技术的综合理解，具有挑战性的社会问题，以及市场分析和风险的工具。该证书将作为S大师项目在目前的提案中进行试点。该方案将加强STEM教育基础设施，并促进低收入国家中代表性不足的少数民族与STEM学员的互动。这将通过增加科学活动和合作，通过对特定国家社会挑战的技术经济分析，以及通过培训具有全球意识的工程师，使我们的全球合作伙伴受益。拟议工作的基本原理是，在没有细胞生长的情况下，为高效的非生长相关的生物制造过程实现高通量新陈代谢。开发具有快速、非生长产物代谢的细胞，将通过优化底物转化为产品而不牺牲底物消耗来实现细胞生长，从而消除生物制造经济蓬勃发展的主要障碍。然而，许多生化产品是生长偶联的，通常不生长的细胞代谢率较低。这项建议的总体目标是确定在非生长条件下抑制糖酵解代谢的变构调节和翻译后修饰(酶水平调节)。中心假说是，在稳定期，酶水平调节主导代谢下调。这项拟议的工作将推出两种新的方法来设计和表征代谢调节，并发展对非生长型中心碳代谢调节的系统水平的理解。该提案将在蛋白质组水平上产生最小的细胞，绕过基因缺失的问题。该提案还将开发一种快速、有针对性地降解蛋白质的方法，通过击倒副产物酶来实现接近最佳产量的生物转化。这种方法将使新的生物学研究成为可能，因为它将有助于制造条件突变，以新的方式扰乱生物系统。第二种方法将根据热力学确定限速酶，从而将工程努力集中在特定的酶上。开发的工作流程将确定在广泛的工业相关条件下，反应是接近平衡(非限速)还是偏离平衡(限速)。这一新的工作流程将被用于研究中央碳代谢调控，以获得对有机酸和萜类生产调控的系统水平的观点。这一职业奖由CBET部门的生物技术和生化工程计划获得，由分子和细胞生物学部门的系统和合成生物学计划共同资助。