CAREER:The Role of ATP Hydrolysis in Biological Nitrogen Fixation

职业：ATP 水解在生物固氮中的作用

• 批准号: 0643777
• 负责人: Faik Tezcan
• 金额: $ 88.74万
• 依托单位: University of California-San Diego
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0643777&HistoricalAwards=false
• 关键词: CAREER; Role; ATP; Hydrolysis; Biological

Reduced forms of molecular nitrogen (dinitrogen) are essential for the production of fertilizers and countless industrial chemicals, as well as the biosynthesis of amino and nucleic acids. The reduction of dinitrogen to ammonia (nitrogen fixation) by the Haber-Bosch process requires high temperatures and pressures, consuming over 100 billion watts of power every year. Biological dinitrogen fixation does not require such extreme conditions; indeed, the enzyme nitrogenase catalyzes ammonia synthesis under ambient conditions. After decades of intense research efforts, however, it is still not known how nitrogenase activates dinitrogen. Nitrogenase catalysis is distinct from other multielectron/multiproton catalytic reactions in its requirement of 16 ATP molecules per turnover reaction despite a favorable thermodynamic driving force. This project aims to elucidate why and how ATP hydrolysis is required in biological nitrogen fixation. Recent structural studies show that the two constituents of nitrogenase, the Fe-protein (electron donor/ATPase) and the MoFe-protein (catalytic component), can assume at least three docking geometries, depending on the ATP-hydrolysis state. By developing and utilizing several powerful chemical and biophysical tools, this project will probe whether multiple Fe-protein:MoFe-protein docking modes are functionally important, and if they are involved in timing the successive electron and proton transfers into the catalytic metal cluster (FeMoco). In parallel, photochemical methods will be utilized to investigate the possibility of driving substrate reduction at FeMoco without requiring ATP hydrolysis. The demonstration of photoactivation of nitrogenase will open new avenues for studying its mechanism that in turn could lead to the development of biocatalytic systems for ammonia and hydrogen production.Broader Impacts: The complexity of biological nitrogen fixation requires a multidisciplinary plan of attack. This project combines a multitude of experimental approaches that will provide an expansive training ground for graduate and undergraduate students. Biological nitrogen fixation sustains a large fraction (40%) of the world's population, and the industrial Haber-Bosch process is responsible for considerable amounts of energy consumption and greenhouse gas emissions. A thorough understanding of nitrogenase mechanism could lead to the design of clean and efficient biocatalysts for ammonia production, which would have an immense economic and environmental impact. Nitrogen fixation also provides a conduit into the education goals of this project, which is to raise the awareness of students about the global energy problem, and to train scientists in energy biosciences. These goals will be addressed on several fronts, including a) the interdisciplinary training of graduate and undergraduate students in the laboratory, b) restructuring of an advanced course in Bioinorganic Chemistry to focus on redox-catalytic processes involved in global carbon, nitrogen, oxygen, and sulfur cycles, as well as the design of a multidisciplinary course on Global Energy Problem and Alternative Energy Research, and c) outreach to low-income students from underrepresented groups attending a local charter school, and their recruitment through seminars.

分子氮的还原形式（二氮）对于肥料和无数工业化学品的生产以及氨基和核酸的生物合成至关重要。通过哈伯-博世工艺将二氮还原为氨（固氮）需要高温和高压，每年消耗超过1000亿瓦的电力。生物固氮并不需要这样极端的条件;事实上，固氮酶在环境条件下催化氨的合成。然而，经过几十年的深入研究，仍然不知道固氮酶如何激活双氮。固氮酶催化与其他多电子/多质子催化反应的不同之处在于，尽管有有利的热力学驱动力，但每次周转反应需要16个ATP分子。本项目旨在阐明生物固氮中ATP水解的原因和方式。最近的结构研究表明，固氮酶的两个组成部分，铁蛋白（电子供体/ATP酶）和钼铁蛋白（催化组分），可以假设至少有三个对接的几何形状，这取决于ATP水解状态。通过开发和利用几种强大的化学和生物物理工具，该项目将探讨多种Fe-蛋白质：MoFe-蛋白质对接模式是否在功能上重要，以及它们是否参与了连续的电子和质子转移到催化金属簇（FeMoco）的时间。同时，将利用光化学方法研究在FeMoco驱动底物还原而无需ATP水解的可能性。固氮酶光活化的演示将为研究其机制开辟新的途径，从而可能导致氨和氢生产的生物催化系统的发展。更广泛的影响：生物固氮的复杂性需要一个多学科的攻击计划。该项目结合了大量的实验方法，将为研究生和本科生提供广阔的培训基地。生物固氮维持了世界人口的很大一部分（40%），而工业哈伯-博施过程是造成大量能源消耗和温室气体排放的原因。深入了解固氮酶的作用机理，可以设计出清洁高效的生物催化剂用于合成氨，这将产生巨大的经济和环境影响。固氮还为实现该项目的教育目标提供了一个渠道，该项目旨在提高学生对全球能源问题的认识，并培养能源生物科学方面的科学家。这些目标将在几个方面得到解决，包括a）研究生和本科生在实验室的跨学科培训，B）生物无机化学高级课程的重组，重点关注全球碳，氮，氧和硫循环中涉及的氧化还原催化过程，以及全球能源问题和替代能源研究的多学科课程的设计，（c）向在当地特许学校就读的低收入家庭学生（来自代表性不足的群体）进行外联，并通过研讨会招募他们。