Electron donation from ascorbic acid to nitrite reductase via ferredoxin in a reconstituted system

In this study, addition of ascorbic acid (AsA) to a reconstituted system composed of nitrite reductase (NiR) and ferredoxin (Fd) proteins could reduce nitrite at a rate nearly equal to that is obtainable with the NADPH-FNR-Fd-NiR system. The addition of spinach (Spinacia oleracea L. 'Birohurei') leaf extract (LES) enabled acceleration of nitrite reduction in the former system but not in the latter system. In the AsA-Fd-NiR-LES system, switching the incubation atmosphere from air to N₂ gas also accelerated nitrite reduction. We added ∨15N-labeled nitrite to the system and examined the formation of ammonia by using a ∨15N-tracing technique. At pH 6.5, we observed quantitative recovery of ∨15N-labeled ammonia in reference with nitrite reduction, but ammonia loss was apparent in the assay at pH 7.5.