Monitoring N2O Production Using a cNOR Modeled Active Site

Nitrous oxide (N2O) is a potent greenhouse gas with a 100-year global warming potential 265-296 times greater than carbon dioxide (CO2). It is the leading contributor to ozone depletion and can persist in the stratosphere for approximately 114 years. Hence, understanding the sources of atmospheric N2O emissions is critical to remediating the effects of climate change. Agricultural activities are the largest contributor to N2O emissions in the U.S. with microbial nitrification and denitrification as the dominating soil processes. The enzyme cytochrome c nitric oxide reductase (cNOR) is involved in bacterial denitrification. It is often difficult to study the enzymes involved in biotic N2O production, hence, model enzymes are a useful tool. The enzyme I107EFeBMb, a sperm whale myoglobin derivative, models the active site of cNOR and was used to simulate the anaerobic reduction of NO to N2O by cNOR. Dithionite was used to induce the catalytic activity of I107EFeBMb by reducing the enzyme. However, dithionite is a strong reductant that is capable of reducing NO to N2O directly. Therefore, the dithionite-enzyme mixture was passed through a size-exclusion column to isolate the reduced enzyme. This reduced and purified enzyme was then utilized to investigate N2O production from NO. This project will provide both an enzymatic and abiotic model to study N2O production.