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

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.