Nitric oxide reductases (NORs) are a family of oxidoreductases that catalyze reduction of nitric oxide to nitrous oxide (2NO + 2H++ 2e- ? N2O + H2O), a critical step of the biological denitrification process. Bacterial NORs are membrane proteins that contain a binuclear active site consisting of a heme b3 site and an iron-containing non-heme site (FeB) where the reduction of nitric oxide takes place. Since the NORs are large membrane proteins, structural features and mechanism responsible for how the N-O bond is cleaved and the N-N bond is formed during the catalytic process is still not clear. To this end, we have engineered biosynthetic models of NORs using myoglobin, which is much simpler than the native NORs, can be easily purified and crystallized. Using this approach we have successfully engineered a non-heme iron (FeB) site in the distal pocket of myoglobin by introducing both histidines and glutamates. The resulting biosynthetic models mimic native NORs both structurally, as confirmed by the overlay of crystal structures of the models with native NORs, and functionally, as revealed by the new NOR activity.1 More importantly, we have used these NOR models to show that the presence of first glutamate is important for binding of non-heme iron and for conferring NOR activity, while the presence of the second glutamate and its associated hydrogen bonding network is important for tuning the NOR activity.1 Using various spectroscopic techniques, such as UV-Vis, EPR, Resonance Raman, Nuclear Resonance Vibrational Spectroscopy and M�ssbauer spectroscopy, combined with replacing the heme with zinc porphyrin, we have elucidated the mechanism of NO reduction.2 These and other recent results will be presented.a) Nature, 462, 1079 (2009); b) PNAS, 107, 8581 (2010).a) Angew. Chem., Int. Ed. 53, (9) 2417 (2014); b) J. Am. Chem. Soc. 136, (6) 2420 (2014).

Using biosynthetic models of nitric oxide reductases in myoglobin to understand structural features and mechanism responsible for NO reduction

248th National Meeting (2014)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.