Denitrification under feast and famine – the role of the bc1 complex in securing electrons for N2O reduction

While CO2 emissions are predicted to decrease during the next decades, N2O emissions will continue to increase unless we find novel mitigation options. A major part of anthropogenic N2O is produced by denitrifying bacteria in agricultural systems. These are facultative anaerobes that use nitrogen oxides as terminal electron acceptors when O2 becomes scarce. Organisms with a complete denitrification pathway are either net sources or sinks for N2O, depending on their regulation. Current knowledge about this regulation is biased because experiments were conducted under optimal conditions, while bacteria in soils starve most of the time. Based on few studies and little evidence, the general notion is that N2O reduction will “loose” under electron donor limitation. Our recent study1 showed that Bradyrhizobium isolates carrying N2O reductase (Nos) and the periplasmic nitrate reductase Nap, but lacking the membrane-bound Nar, had a strong preference for N2O over NO3- when grown under well-fed conditions. This was not due to different abundancies of the reductases, suggesting a metabolic-level phenomenon where the bc1 complex, which channels electrons to Nos via cytochrome c, wins the competition for electrons over the Nap pathway. Here we compared the denitrification phenotypes of some other denitrifying bacteria, which supported our hypothesis that Nap+NosZ strains are strong sinks for N2O while Nar+NosZ strains show similar preference for nitrate and N2O, suggesting that the Nar and Nos pathways compete equally for electrons from the quinol pool. Moreover, we exposed cultures of Bradyrhizobium, Thauera and Pseudomonas strains to starvation using a bioassay protocol in which the respiration rate was reduced to 1-18% compared to well-fed cells. As for well-fed cultures, strains with Nap+Nos retained a strong preference for N2O over NO3- under electron donor limitation, contrasting the general notion that Nos activity diminishes under substrate limitation. The results suggest that denitrifiers with Nap+Nos but lacking Nar can be strong N2O sinks in soil. Such organisms should be chosen when developing methods to engineer soil microbial systems for N2O mitigation.