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

一氧化二氮（N2 O）是一种强有力的大气污染物，可通过一种称为原核一氧化二氮还原酶（N2 OR）的细菌酶进行生物去除。在高产农业土壤中，N2 OR通常不充分活跃。因此，N2 O在后工业化大气中一直在持续增加，因此它占目前温室气体辐射强迫的12%左右。N2 O也是一种重要的臭氧消耗化学品。一个新的和潜在的变革性的解决方案，农业排放的N2 O将在这个项目中进行调查。N2 OR将在植物线粒体中表达，使其持续活跃，从土壤和大气中去除N2 O。据报道，N2 OR在体外缺氧条件下在用N2 OR的结构基因nosZ常规转化的植物提取物中是有活性的。利用研究人员在实验室中开发的用于构建植物修复的新型合成遗传载体的专业知识，他们将表达用氨基酸序列修饰的nosZ，这些氨基酸序列将蛋白质靶向线粒体基质和线粒体的内周质膜。呼吸子是细胞中呼吸减少的位点，即，氧消耗的位点，类似于反硝化细菌中N2 O还原的位点。这是一个高风险项目，但具有全球重要性的巨大潜在利益。在一年的资金，该项目旨在确定：1。利用目前的nosZ序列和线粒体靶向序列是否可能在整个植物中减少N2 O，2. nos操纵子中假定的辅助基因nosXDFYL是否是最佳N2 OR表达所必需的;如果N2 OR在植物中具有活性，并且如果这项技术大规模应用于玉米和水稻等作物，预计谷物种植中的N2 O排放量可能会显著减少，从而导致N2 O增加量的突然减少。潜在地，这种减少可以可持续地实现，几乎没有额外的能量消耗。这将是温室气体和臭氧消耗排放量的重大且受欢迎的削减，这是IPCC目前的预测所没有预料到的。该奖项反映了NSF的法定使命，并且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.