CAREER: Connecting eukaryotic electron transfer components to nitrogenase using a bacterial chassis

职业：使用细菌底盘将真核电子传递组件连接到固氮酶

• 批准号: 2338085
• 负责人: Kathryn Fixen
• 金额: $ 103.62万
• 依托单位: University of Minnesota-Twin Cities
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2024
• 资助国家: 美国
• 起止时间: 2024-02-01 至 2029-01-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2338085&HistoricalAwards=false
• 关键词: CAREER; Connecting; eukaryotic; electron; transfer

Important advancements in our understanding of biological nitrogen fixation, bolstered by emerging synthetic biology tools, suggest we are closer than ever to engineering plants to fix nitrogen. Engineering plants to fix nitrogen could improve the sustainability of the bioeconomy. However, we lack the knowledge to predict the behavior and specificity of electron carriers such as ferredoxin, which are needed to power nitrogenase. There is a critical need to test eukaryotic electron transfer components for their ability to interact with nitrogenase and measure how changes in the cellular redox environment sustain electron flow. The overall objective of the research proposed here is to use a bacterial chassis to rapidly define how eukaryotic electron transfer components can participate in electron delivery to nitrogenase and invent a powerful platform for evolution of synthetic electron flow pathways. The research aims synergize with educational goals by incorporating a semester-long project that focuses on bioengineering nitrogen fixation into existing courses using a novel culturally responsive pedagogical framework.The central hypothesis for the project is that electron transfer to nitrogenase is one of the primary constraints preventing introduction of this enzyme into eukaryotic systems, but it is possible to select for variants in eukaryotic electron transfer components to overcome this bottleneck. To test this hypothesis, the investigator proposes to develop a new tool to analyze electron flow to nitrogenase. This tool will use a bacterial chassis to test eukaryotic electron transfer components at physiological levels and evolve these electron transfer components for enhanced electron flow to nitrogenase. Such a contribution would be significant because it would enable more accurate predictions regarding nitrogenase functionality within plant organelles and would establish a robust platform for optimizing electron transfer to nitrogenase. This would not only make the goal of engineering plants to fix nitrogen more attainable, but it would also further our understanding of the determinants of electron flow and how electron transfer pathways can be optimized for biotechnological purposes. This project is supported by the Systems and Synthetic Biology Cluster of the Division of Molecular and Cellular Biosciences.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.

在新兴的合成生物学工具的支持下，我们对生物固氮的理解取得了重要进展，这表明我们比以往任何时候都更接近工程化植物来固氮。改造植物固氮可以提高生物经济的可持续性。然而，我们缺乏预测电子载体（例如为固氮酶提供动力所需的铁氧还蛋白）的行为和特异性的知识。迫切需要测试真核电子转移成分与固氮酶相互作用的能力，并测量细胞氧化还原环境的变化如何维持电子流。这里提出的研究的总体目标是使用细菌底盘快速定义真核电子传递组件如何参与电子传递到固氮酶，并为合成电子流路径的进化发明一个强大的平台。该研究旨在利用一种新颖的文化响应式教学框架，将一个为期一个学期的专注于生物工程固氮的项目纳入现有课程，从而与教育目标产生协同作用。该项目的中心假设是，电子转移到固氮酶是阻止固氮酶的主要限制之一。将这种酶引入真核系统，但可以选择真核电子转移成分的变体来克服这一瓶颈。为了检验这一假设，研究人员建议开发一种新工具来分析流向固氮酶的电子流。该工具将使用细菌底盘在生理水平上测试真核电子转移成分，并进化这些电子转移成分以增强电子流向固氮酶。这样的贡献将是意义重大的，因为它将能够更准确地预测植物细胞器内固氮酶的功能，并将建立一个强大的平台来优化固氮酶的电子转移。这不仅使工程植物固氮的目标更容易实现，而且还将进一步加深我们对电子流决定因素以及如何针对生物技术目的优化电子转移途径的理解。该项目得到了分子和细胞生物科学部系统和合成生物学集群的支持。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。