EAGER: Preventing N2O Emissions by Expressing N2O Reductase in Plant Mitochondria

一氧化二氮（N2O）是一种强效的大气污染物，可以通过一种称为原核一氧化二氮还原酶（N2OR）的细菌酶进行生物去除。在高产农业土壤中，N2OR的活性往往不足。因此，后工业时代大气中N2O持续增加，约占目前温室气体辐射强迫的12%。一氧化二氮也是一种重要的消耗臭氧的化学物质。在这个项目中，将研究一种新的和潜在的变革性解决方案，以解决农业排放N2O的问题。N2OR会在植物线粒体中表达，使其持续活跃，从土壤和大气中去除N2O。据报道，在体外缺氧条件下，N2OR在用N2OR结构基因nosZ转化的植物提取物中具有活性。利用研究人员在实验室开发的专业知识，构建用于植物修复的新型合成遗传载体，他们将表达用氨基酸序列修饰的nosZ，将蛋白质靶向线粒体基质和线粒体内质周膜。线粒体是细胞中呼吸还原的部位，即耗氧部位，类似于反硝化细菌中N2O还原的部位。这是一个高风险的项目，但具有全球重要性的巨大潜在效益。在一年的资金支持下，该项目力求确定：目前nosZ序列和线粒体靶向序列是否可能降低整个植物的N2O；2 . nos操纵子中假定的辅助基因nosxdyl是否对最佳的N2OR表达是必需的；表达N2OR的植物是否能阻止肥料中N2O的排放。如果能够使N2OR在植物中发挥活性，并将该技术大规模应用于玉米、水稻等作物，预计可以显著减少粮食种植过程中N2O的排放，从而使N2O的增加突然减少。这种减少有可能以可持续的方式实现，几乎不需要额外的能源消耗。这将大大减少温室气体和消耗臭氧层的排放，这是IPCC目前的预测所没有预料到的。该奖项反映了美国国家科学基金会的法定使命，并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，被认为值得支持。

Nitrous oxide (N2O) is a potent atmospheric pollutant that is removed biologically by a bacterial enzyme called prokaryotic nitrous oxide reductase (N2OR). In high yield agricultural soils N2OR is often inadequately active. Thus, N2O has been continuously increasing in post-industrial atmosphere, so that it accounts for about 12% of present radiative forcing by greenhouse gases. N2O is also a significant ozone depleting chemical. A novel and potentially transformative solution to agricultural emissions of N2O will be investigated in this project. N2OR will be expressed in plant mitochondria so that it is active continuously, removing N2O from soil and atmosphere. N2OR has been reported to be active under anoxic conditions in vitro in extracts of plants conventionally transformed with nosZ, the structural gene for N2OR. Using the expertise that the investigators have developed in their lab for the construction of novel synthetic genetic vectors for phytoremediation, they will express nosZ modified with the amino acid sequences that target the proteins to the mitochondria matrix and inner periplasmic membrane of the mitochondria. The mitochondrion is the site of respiratory reduction in the cell, i.e., the site of oxygen consumption, and is analogous to the site of N2O reduction in denitrifying bacteria. This is a high-risk project, but with great potential benefits of global importance. With one year of funding, the project seeks to determine:1. whether N2O reduction in whole plants is possible using current nosZ sequences and mitochondrial targeting sequences,2. whether the putative helper genes in the nos operon, nosXDFYL, are necessary for optimal N2OR expression, and3. whether N2OR expressing plants can prevent N2O emissions from fertilized soils.If N2OR can be made to be active in plants and if this technology is applied on a large scale to crops such as maize and rice, it is expected that N2O emissions from grain cultivation could be cut significantly, resulting in a sudden reduction in N2O increases. Potentially this reduction could be accomplished sustainably with little to no additional energy expenditure. This would be a major and welcome cut in greenhouse gas and ozone depleting emissions, unanticipated by present IPCC projections.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.