RESUMO
Anammox bacteria performed the reaction of NH4+ and NO with hydrazine synthase to produce N2H4, followed by the decomposition of N2H4 with hydrazine dehydrogenase to generate N2. Ferroheme/ferriheme, which serves as the active center of both hydrazine synthase and hydrazine dehydrogenase, is thought to play a crucial role in the synthesis and decomposition of N2H4 during Anammox due to its high redox activity. However, this has yet to be proven and the exact mechanisms by which ferroheme/ferriheme is involved in the Anammox process remain unclear. In this study, abiotic and biological assays confirmed that ferroheme participated in NH4+ and NO reactions to generate N2H4 and ferriheme, and the produced N2H4 reacted with ferriheme to generate N2 and ferroheme. In other words, the ferroheme/ferriheme cycle drove the continuous reaction between NH4+ and NO. Raman, ultraviolet-visible spectroscopy, and X-ray absorption fine structure spectroscopy confirmed that ferroheme/ferriheme is involved in the synthesis and decomposition of N2H4 via the core FeII/FeIII cycle. The mechanism of ferroheme/ferriheme participation in the synthesis and decomposition of N2H4 was proposed by density functional theory calculations. These findings revealed for the first time the heme electron transfer mechanisms, which are of great significance for deepening the understanding of Anammox.