ABSTRACT:   Reduction of nitrite anions (NO₂^–) to nitric oxide (NO), nitrous oxide (N₂O) and ultimately dinitrogen (N₂) takes place in a variety of environments, including in the soil as part of the biogeochemical nitrogen cycle and in acidified nuclear waste. Nitrite reduction typically takes place within the coordination sphere of a redox-active transition metal. Here we show that Lewis acid coordination can substantially modify the reduction potential of this polyoxoanion to allow for its reduction under non-aqueous conditions (–0.74 V versus NHE). Detailed characterization confirms the formation of the borane-capped radical nitrite dianion (NO₂^2–), which features a N(II) oxidation state. Protonation of the nitrite dianion results in the facile loss of nitric oxide (NO), whereas its reaction with NO results in disproportionation to nitrous oxide (N₂O) and nitrite (NO₂^–). This system connects three redox levels in the global nitrogen cycle and provides fundamental insights into the conversion of NO₂^– to NO. 

ABSTRACT:   Ammonia (NH₃)-oxidizing bacteria (AOB) derive total energy for life from the multi-electron oxidation of NH₃ to nitrite (NO₂⁻). One obligate intermediate of this metabolism is hydroxylamine (NH₂OH), which can be oxidized to the potent greenhouse agent nitrous oxide (N₂O) by the AOB enzyme cytochrome (cyt) P460. We have now spectroscopically characterized a 6-coordinate (6c) {FeNO}⁷ intermediate on the NH₂OH oxidation pathway of cyt P460. This species has two fates: it can either be oxidized to the {FeNO}⁶ that then undergoes attack by NH₂OH to ultimately generate N₂O, or it can lose its axial His ligand, thus generating a stable, off-pathway 5-coordinate (5c) {FeNO}⁷ species. We show that the wild type (WT) cyt P460 exhibits a slow nitric oxide (NO)-independent conversion (k_His-off = 2.90 × 10⁻³ s⁻¹), whereas a cross-link-deficient Lys70Tyr cyt P460 mutant protein underwent His dissociation via both a NO-independent (k_His-off = 3.8 × 10⁻⁴ s⁻¹) and a NO-dependent pathway [k_His-off(NO) = 790 M⁻¹ s⁻¹]. Eyring analyses of the NO-independent pathways for these two proteins revealed a significantly larger (ca. 27 cal mol⁻¹ K⁻¹) activation entropy (ΔS^‡) in the cross-link-deficient mutant. Our results suggest that the Lys–heme cross-link confers rigidity to the positioning of the heme P460 cofactor to avoid the fast NO-dependent His dissociation pathway and subsequent formation of the off-pathway 5c {FeNO}⁷ species. The relevance of these findings to NO signaling proteins such as heme-nitric oxide/oxygen binding (H-NOX) is also discussed.