Understanding nitrous oxide emission from denitrifying bacteria: integrating chemostat and soil studies

我们人类完全依赖于我们呼吸的氧气来支持我们的代谢过程。值得注意的是，对于许多种类的细菌来说，情况并非如此。面对环境中缺氧的情况，许多细菌物种能够转向使用硝酸盐（NO3-），而不是氧气来支持呼吸。其中一种能量生成过程称为反硝化，将水溶性硝酸盐转化为气态产物，即一氧化氮（NO）、氧化亚氮（N2O）和二氮（N2）。这种反硝化过程可以在农业土壤中广泛发生，其中添加富氮肥料以刺激植物生长也可以刺激细菌氮循环。能够反硝化的土壤细菌需要保护自己免受自身代谢产生的NO的自毒作用。它们有一种酶，一氧化氮还原酶（NOR），通过将内源性一氧化氮转化为相对无害的一氧化二氮（N2O），从而将其释放到大气中，从而保持低水平。从细菌代谢的角度来看，细胞毒性NO的解毒工作是在它转化为N2O时完成的，但从环境的角度来看，一种环境毒素，一种温室气体，已经产生了。在讨论温室气体排放时，公众对二氧化碳和甲烷所带来的问题非常敏感。然而，一氧化二氮的排放也应该引起人们的关注，二氧化二氮可能是最著名的牙科麻醉“笑气”。1793年，英国化学家约瑟夫·普里斯特利（Joseph Priestley）首次发现了一氧化二氮，当时它的大气水平已经稳定了几千年。然而，在过去的100年里，大气中的一氧化二氮增加了十亿分之五十，并且每年以0.25%的速度进一步增加，大多数评论家将这种增加与20世纪为提高农田生产力而密集使用化肥联系起来。虽然它在大气中的含量只是二氧化碳的一小部分，但它的全球变暖潜力是二氧化碳的300倍。因此，当以二氧化碳当量表示时，它约占全球温室气体排放总量的10%。由于今天产生的一氧化二氮在大气中的寿命约为150年，它可能会影响我们的曾曾孙所经历的气候，因此，现在制定减少这些排放的策略是很重要的。反硝化细菌从硝酸盐中产生NO的途径是从分子水平上理解的，其中将硝酸盐转化为亚硝酸盐（硝酸盐还原酶）和将亚硝酸盐转化为一氧化氮（亚硝酸盐还原酶）的酶的结构是已知的。这些酶是金属蛋白，依赖于过渡金属如钼、铁和铜的活性。将N2O分解成惰性N2的酶是一种含铜酶（Nos）。它是地球上产生强效一氧化二氮温室气体的主要酶。如果没有它，大气中的一氧化二氮水平将比现在高得多。Nos的分子结构是已知的。它含有12个铜原子，因此它在环境中的活性取决于铜的生物利用度。它对pH值和氧气也很敏感，因此它在环境中的活性取决于许多不同的环境变量。由于在土壤中施用氮肥，人为排放的N2O的最大来源是农业土壤。自英国签署《京都议定书》以来，许多非生物源的一氧化二氮排放已经减少，但生物源的排放却不太容易管理。改善农业N2O排放预测和管理的努力将受益于更好地了解影响细菌N2O净产量的因素。这需要在受控的实验室环境中对模式生物的研究和在土壤中的原位研究相结合。这个项目将提供这样一个综合研究。

We humans we are entirely dependent on the oxygen we breath to support our metabolic processes. Significantly, this is not so for many species of bacteria. Faced with a shortage of oxygen in their environment many bacterial species are able to switch to using nitrate (NO3-), rather than oxygen to support respiration. One of these energy yielding processes, known as denitrification, converts water-soluble nitrates to gaseous products, nitric oxide (NO), nitrous oxide (N2O) and dinitrogen (N2). This denitrification process can take place extensively in agricultural soils where nitrogen rich fertilisers added to stimulate plant growth can also stimulate bacterial nitrogen cycling. Soil bacteria which can denitrify need to protect themselves from the autotoxic effects of NO produced through their own metabolism. They have an enzyme, nitric oxide reductase (NOR) that has evolved to keep endogenous NO levels low by converting it to the relatively benign nitrous oxide (N2O) which can be released into the atmosphere. From the perspective of bacterial metabolism the job of detoxifying the cytoxic NO is done when it is converted to N2O, but from an environmental perspective an envirotoxin, a greenhouse gas, has been produced. When discussing greenhouse gas emissions the public are acutely aware of the problems posed by carbon dioxide and methane. However, emissions of N2O, perhaps best known as the dental anaesthetic 'laughing gas', should also cause concern. N2O was first discovered by the British chemist Joseph Priestley in 1793 when its atmospheric levels had been steady for millennia. However, over the last 100 years N2O in the atmosphere has increased by 50 parts per billion and this atmospheric loading is increasing further by 0.25% each year, with most commentators linking this increase to intensive use of fertiliser to increase farmland productivity in the 20th Century. Although its atmospheric levels are only a fraction of that of CO2 it has a 300-fold greater global warming potential. Thus when expressed in terms of CO2 equivalents it represents around 10% of total global emissions of greenhouse gases. Since it has an atmospheric lifetime of some 150 years the N2O produced today will potentially influence the climate experienced by our great-great grandchildren thus it is important to devise strategies to mitigate these releases now. The pathways by which denitrifying bacteria produce NO from nitrate are undertstood from a molecular level with structures of enzymes that convert nitrate to nitrite (nitrate reductases) and nitrite to nitric oxide (nitrite reductases) being known. These enzymes are metalloproteins that depend on transition metals such as molybdenum, iron and copper for activity. The enzyme that breaks down N2O to inert N2 is a copper-containing enzyme (Nos). It is the major enzyme on the planet that is responsible for the potent N2O greenhouse gas. Without it the atmospheric levels of N2O would be much greater that they currently are. The molecular structure of Nos is known. It contains twelve atoms of copper and so its activity in the environment is dependent on the bioavailability of copper. It is also sensitive to pH and oxygen and so its activity in the environment is dependent on a number of different environmental variables. The largest source of anthropogenic N2O emissions is agricultural soils because of the application of nitrogenous fertilisers to soils. Since the UK signed up to the Kyoto Protocol, many non-biological sources of N2O emissions have been reduced, but emissions from biological sources are less easy to manage. Efforts to improve the prediction and management of agricultural N2O emissions will benefit from a better understanding of the factors that influence the net production of N2O by bacteria. This requires a combination of studies on model organisms in controlled laboratory environments and on studies in situ in soils. This project will provide such an a integrated study.