Structural insight into the substrate- and dioxygen-binding manner in the catalytic cycle of rieske nonheme iron oxygenase system, carbazole 1,9a-dioxygenase

In the present study, we determined the crystal structures of the reduced carbazole (CAR)-bound, dioxygen-bound, and both CAR- and dioxygen-bound CARDO-O: CARDO-F binary complex structures at 1.95, 1.85, and 2.00？？ resolution. These structures revealed the conformational changes that occur in the catalytic cycle. Structural comparison between complex structures in each step of the catalytic mechanism provides several implications, such as the order of substrate and dioxygen bindings, the iron-dioxygen species likely being Fe(III)-(hydro)peroxo, and the creation of room for dioxygen binding and the promotion of dioxygen binding in desirable fashion by preceding substrate binding.The RO catalytic mechanism is proposed as follows: When the Rieske cluster is reduced, substrate binding induces several conformational changes (e.g., movements of the nonheme iron and the ligand residue) that create room for oxygen binding. Dioxygen bound in a side-on fashion onto nonheme iron is activated by reduction to the peroxo state [Fe(III)-(hydro)peroxo]. This state may react directly with the bound substrate, or O–O bond cleavage may occur to generate Fe(V)-oxo-hydroxo species prior to the reaction. After producing a cis-dihydrodiol, the product is released by reducing the nonheme iron. This proposed scheme describes the catalytic cycle of ROs and provides important information for a better understanding of the mechanism.Many studies have demonstrated that aromatic ring dihydroxylation plays a primary role in the initial step of aerobic bacterial degradation pathways for various natural and synthetic aromatic compounds, including dioxins, polychlorinated biphenyls, and crude oil components such as polycyclic aromatic hydrocarbons and heterocyclic aromatic compounds [1-7]. Such ring dihydroxylation is catalyzed by multicomponent oxygenase systems known as Rieske nonheme iron oxygenase systems (ROs). A member of ROs, called as aromatic ring-hydroxylating dioxygenase, catalyze the inco