Dioxetane and lactone pathways in dioxygenolytic ring cleavage catalyzed by 2,5-dihydroxypyridine dioxygenase

2,5-Dihydroxypridine dioxygenase (NicX) catalyzes an aromatic N-heterocylic ring opening. We renew reaction mechanisms of NicX and discover a competitive dioxetane pathway. Although the initial steps involve single-electron transfer from 2,5-hydroxypyridine (DHP) to dioxygen in both the lactone and dioxetane pathways, the ring-opening potential energy surface of the dioxetane pathway was calculated to be more favored. Energetic span (δE) of the lactone pathway was calculated to be 16.2 kcal/mol for the aromatic ring-opening step, higher by 9.4 kcal/mol than that of the dioxetane pathway. Moreover, the reactivity and pre-reaction state analyses suggest that DHP prefers a monoanionic form, or a neutral form upon proton shuttling of H105, rather than a dianionic form. Our results indicate that the 2-His-1-carboxylate-1-serine iron enzyme catalyzes the N-heterocylic aromatic ring opening via the preferential dioxetane pathway, affording a missing piece in the understanding of non-heme iron-enzyme chemistry.