Novel nitric oxide dismutase drives effective NO degradation in oxygenic denitrification

Nitrogenous wastewater is a major environmental problem. Denitrification refers to the sequential reduction of nitrate (NO 3 - ) and nitrite (NO 2 - ) to nitric oxide (NO) and nitrous oxide (N 2 O) to form N 2 . Oxygenic denitrification could transform NO into N 2 and O 2 via NO dismutase (Nod), an unusual O 2 -forming process to reduce N 2 O emission during nitrogen removal. After confirming the presence of the nod gene in three samples including anaerobically-activated sludge, farmland soil, and paddy soil, the targeted enrichment experiment was conducted. The abundance of key denitrification genes rose by 4.6–13 fold, including NO 2 - reductase ( nir ), N 2 O reductase ( nos ), NO reductase ( nor ), and nod . Principal coordinate and α -diversity analyses revealed the progressive enrichment of oxygenic denitrifiers by eliminating irrelevant microorganisms. Focusing on paddy soil with optimal O 2 -generating capacity, novel nod genes were identified. Specifically, recombinant-engineered bacteria containing the nod genes were constructed to assess their NO dismutation activity. Results demonstrated that these recombinant strains exhibited excellent NO degradation efficiency at 72.6–85.4%, indicating superior dismutation performance. Apparent enzymatic activity assays using the crude Nod enzyme extract from the representative recombinant strain SD52149 showed Michaelis-Menten-type saturation behavior. These findings provide critical insights into the application of oxygenic denitrification for reducing N 2 O emissions in nitrogenous wastewater treatment and enhancing microbial remediation strategies.