NSF/BIO-DFG: Biological Fe-S intermediates in the synthesis of nitrogenase metalloclusters

NSF/BIO-DFG：固氮酶金属簇合成中的生物 Fe-S 中间体

• 批准号: 536145634
• 负责人: Professor Dr. Oliver Einsle
• 金额: --
• 依托单位: Institut für Biochemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/536145634?language=en
• 关键词: NSF; BIO; DFG; Biological; Fe

Nitrogenase catalyzes the biological conversion of N2 gas into NH4+, a critical step in the biogeochemical nitrogen cycle. Despite its importance for life on Earth, nitrogen fixation is a trait limited to a subset of microbes, and the capacity to fix N2 is complex in mechanism, genetics, and regulation. The activity of nitrogenase depends on factors involved in the synthesis of its associated metallocofactors and on metabolic conditions that support this highly endergonic reaction. Therefore, defining the reactions performed by these factors has intrinsic intellectual merit and helps to understand their evolution. It is significant for developing sustainable, energy-saving strategies to produce nitrogen-based food crop supplements. This proposal aims to elucidate initial reaction steps involve the synthesis of Fe-S clusters destined for nitrogenase that are promoted by NifU and NifS. The presence of NifU and NifS is dominant in other aerobic nitrogen-fixing microbes, suggesting the requirement of a dedicated Fe-S cluster biosynthesis system for sustaining nitrogen fixation exactly then. In the model bacterium Azotobacter vinelandii, NifUS is required for nitrogen fixation by any of the three nitrogenase isoforms (Nif, Anf, and Vnf). Our preliminary data now show a previously uncharacterized cluster species on NifU when expressed in its native organism, A. vinelandii. We also show that Fpr2 is a suitable physiological reductant enabling for the first time the detection of novel, EPR-active species associated with NifU. We hypothesize that this transient cluster species represents a functional intermediate in the synthesis of nitrogenase metalloclusters. The research plan is aimed at investigating the mechanistic and chemical steps of Fe-S cluster synthesis by establishing 1) the location and spectroscopic features of novel cluster species associated with NifU, 2) the involvement additional factors in this process, and 3) the reactivity of the new cluster species as building blocks for the synthesis of nitrogenase metalloclusters. Defining the molecular and evolutionary details that enable nitrogen fixation has a prospect for a major agronomic, economic, and environmental impact. These principles can also guide foundational studies on Fe-S clusters on more complex systems not limited to nitrogen fixation. The interdisciplinary, collaborative international project will engage undergraduate and graduate students through research experiences and professional development opportunities. A German and an American PI partnership will support two synergistic research and professional development efforts: (a) collaborative research experiences abroad for graduate students; and (b) mentored research fellowships for local undergraduate students.

固氮酶催化氮气生物转化为NH4+，这是生物地球化学氮循环的关键步骤。尽管固氮对于地球上的生命很重要，但它是一种仅限于微生物子集的特性，并且固氮的能力在机制、遗传学和调控方面很复杂。固氮酶的活性取决于参与其相关金属辅因子合成的因素以及支持这种高吸能反应的代谢条件。因此，定义这些因素所进行的反应具有内在的智力价值，并有助于理解它们的演变。这对于制定可持续、节能的战略来生产氮基粮食作物补充剂具有重要意义。该提案旨在阐明初始反应步骤，涉及合成由 NifU 和 NifS 促进的固氮酶的 Fe-S 簇。 NifU 和 NifS 在其他需氧固氮微生物中占主导地位，这表明当时需要专用的 Fe-S 簇生物合成系统来维持固氮作用。在模型细菌 Azotobacter vinelandii 中，NifUS 是三种固氮酶亚型（Nif、Anf 和 Vnf）中任何一种固氮所必需的。我们的初步数据现在显示，当 NifU 在其本地生物 A. vinelandii 中表达时，存在以前未表征的簇物种。我们还表明，Fpr2 是一种合适的生理还原剂，首次能够检测与 NifU 相关的新型 EPR 活性物质。我们假设这种瞬时簇物种代表固氮酶金属簇合成中的功能中间体。该研究计划旨在通过确定 1) 与 NifU 相关的新簇物种的位置和光谱特征，2) 该过程中涉及的其他因素，以及 3) 反应性来研究 Fe-S 簇合成的机理和化学步骤。新簇物种作为固氮酶金属簇合成的构建模块。定义固氮的分子和进化细节有望对农艺、经济和环境产生重大影响。这些原理还可以指导在更复杂的系统（不限于固氮）上对 Fe-S 簇的基础研究。这个跨学科的国际合作项目将通过研究经验和专业发展机会吸引本科生和研究生。德国和美国的 PI 合作伙伴关系将支持两项协同研究和专业发展工作：(a) 为研究生提供海外合作研究经验； (b) 为当地本科生提供指导研究奖学金。