EAGER: Leveraging Chaperones to Escape the Plant RuBisCO Catalytic Catch-22

EAGER：利用分子伴侣逃离植物 RubisCO 催化 Catch-22

• 批准号: 2244770
• 负责人: Matthew Shoulders
• 金额: $ 30万
• 依托单位: Massachusetts Institute of Technology
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-15 至 2025-03-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2244770&HistoricalAwards=false
• 关键词: EAGER; Leveraging; Chaperones; Escape; Plant

Photosynthetic organisms use carbon dioxide from the atmosphere to produce sugar and biomass that support all other life. The enzyme RuBisCO is responsible for this chemical capture of carbon dioxide. Unfortunately, it carries out this process only very slowly – to the extent that carbon dioxide capture often limits plant growth. Moreover, RuBisCO regularly and mistakenly reacts with oxygen instead of carbon dioxide, wasting energy and further limiting plant growth. An improved Rubisco, if translated into agriculture, would lead to a significant increase in crop yields, improved food security, and better climate resilience, and improving Rubisco function has long been viewed as an agricultural technology holy grail. With the potential value of such an enzyme in mind, researchers have asked whether it is possible to substantially improve these areas of RuBisCO function, since natural evolution has failed to do so thus far. One widely touted possibility is that RuBisCO is caught in an evolutionary trap and cannot be improved. An alternative view is that subjecting the enzyme to new-to-nature types of selection pressures and environments would enable evolutionary innovations that actually can unlock a better RuBisCO. The goal of this project is to test the latter idea, by integrating fundamental biophysical concepts with state-of-the-art techniques for laboratory evolution. The proposed research will also help to train the next generation of scientists and engineers by involving high school interns each summer in the work. Molecular chaperones interact with and assist other proteins (clients) in the cell and, in doing so, can profoundly influence the evolutionary trajectories accessible to their clients. In plants, RuBisCO requires nearly a dozen of these molecular chaperones to assist its production, regulation, and maintenance. How do the sequences and activities of these chaperones impact the evolution of plant RuBisCO, and are there ways to optimize the chaperones to open new regions of sequence space for RuBisCO? By applying laboratory evolution to RuBisCO and its chaperones in a microbial host, this project will seek to answer these questions. Next-generation mutagenesis technologies and novel screening systems will be used to achieve two key goals: (1) Demonstrating high-throughput in vivo directed evolution of plant RuBisCO and its chaperones and (2) Testing the idea that co-evolution of RuBisCO and its chaperones can lead to better evolutionary outcomes and enhanced enzymatic properties relative to evolving just RuBisCO alone.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.

光合生物利用大气中的二氧化碳来生产糖和生物质，以支持所有其他生命。酶RuBisCO负责这种二氧化碳的化学捕获。不幸的是，这一过程进行得非常缓慢，以至于二氧化碳的捕获往往限制了植物的生长。此外，RuBisCO经常错误地与氧气而不是二氧化碳反应，浪费能源并进一步限制植物生长。如果将Rubisco的改良转化为农业，将导致作物产量显著增加，粮食安全得到改善，气候适应能力得到改善，而改善Rubisco功能长期以来一直被视为农业技术的圣杯。考虑到这种酶的潜在价值，研究人员询问是否有可能大幅改善RuBisCO功能的这些领域，因为自然进化迄今未能做到这一点。一个被广泛吹捧的可能性是，RuBisCO陷入了进化陷阱，无法改进。另一种观点是，将酶置于新的自然选择压力和环境中，可以实现进化创新，实际上可以解锁更好的RuBisCO。该项目的目标是通过将基本生物物理概念与最先进的实验室进化技术相结合来测试后一种想法。这项拟议中的研究还将有助于培养下一代科学家和工程师，每年夏天让高中实习生参与这项工作。分子伴侣与细胞中的其他蛋白质（客户端）相互作用并提供帮助，这样做可以深刻影响客户端的进化轨迹。在植物中，RuBisCO需要近12个这样的分子伴侣来帮助其生产，调节和维护。这些分子伴侣的序列和活性如何影响植物RuBisCO的进化，以及是否有方法优化分子伴侣以打开RuBisCO序列空间的新区域？通过将实验室进化应用于微生物宿主中的RuBisCO及其分子伴侣，该项目将寻求回答这些问题。下一代诱变技术和新型筛选系统将用于实现两个关键目标：（1）证明植物RuBisCO及其伴侣蛋白的高通量体内定向进化，以及（2）测试共-相对于仅进化RuBisCO，RuBisCO及其伴侣的进化可以导致更好的进化结果和增强的酶特性。该奖项反映了NSF的法定使命，并已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，认为值得支持。