Tang Hongzhi's research group and Zhao Yilei's research group of Shanghai Jiaotong University cooperated to elucidate the catalytic mechanism of non-heme iron dioxygenase NicX

Recently, Professor Tang Hongzhi's group and Professor Zhao Yilei's group from the State Key Laboratory of Microbial Metabolism, School of Life Science and Technology, Shanghai Jiao Tong University, published an online publication entitled "Dioxetane and Lactone Pathways in Dioxygenolytic Ring Cleavage Catalyzed by 2,5- Dihydroxypyridine Dioxygenase", elucidating the reaction mechanism
of a novel non-heme ferrase NicX that uses its unique iron-catalyzed center to activate oxygen molecules and degrade substrate aromatic rings with rare retro-[2+2] ring opening after addition to 2,5-dihydroxypyridine (DHP).
。 By numerically comparing the oxygen-oxygen bond breaking radical Criegee lactone rearrangement of the dioxaduct intermediate with the novel dioxocyclic reaction pathway, the authors found that the enzyme has the selectivity of dioxacyclo-opening, thereby explaining the experimental phenomenon
of H105F mutant maintaining activity.
Ouyang Xingyu, a doctoral student in the School of Life Science and Technology, is the first author, and Professor Tang Hongzhi and Professor Zhao Yilei are co-corresponding authors
.

DHP is a refractory pyridine heterocyclic aromatic contaminant
in nicotine degradation.
NicX enzyme has the ability to specifically catalyze the degradation of DHP, converting it to N-formylmaleamide acid (NFM) and entering the main metabolic tricarboxylic acid cycle
of microorganisms 。 In the analysis of the crystal structure of NicX in cooperation between Tang Hongzhi's research group and Zhao Yilei's group, it was found that the enzyme was different from the iron-catalyzed center of traditional non-heme iron oxygenase (Liu et al.
, Nature Communication, 2021, 12: 1301), in addition to the three common ligands 2-His-1-carboxylate (H265-H318-D320), there was also a para-serine residue S302.
This structure is similar to the structure of Heme-Fe-Ser, and the authors proposed several possible catalytic reaction mechanisms related to this special iron coordination structure in Nature Comm.
, which quickly attracted widespread interest among peers and published quantum chemical calculation results
of related mechanisms.

This work proposes a breakthrough update to the early molecular mechanism and discovers the degradation mechanism of the dioxane aromatic ring: the activated energy span (δE) of the Criegee rearranged lactone pathway is as high as 16 kcal/mol, which is much higher than the dioxane nucleophilic addition-retro[2+2] pathway, and has no competitive advantage
.
In addition, the pre-reaction state analysis of Michael's complex also showed that the substrate molecules were strictly constrained by H189-E177 residues and formed positive and negative ion pairs with H105, which avoided invalid secondary electron transport and improved the electrophilic addition ability
of the aromatic ring.
The proton shuttle and competitive loop-opening pathway between DHP-H105 indicate that this residue is important but not necessary in catalysis, while iron ions play a key catalytic role
in oxygen activation, nucleophilic addition, and ring opening.
The research results demonstrate NicX's novel "2-His-1-carboxylate-1-Ser" dioxotetracyclic catalytic ring-opening pyridine nitrogen heterocyclic aromatic pollutants, which fills an extremely important link
in the catalytic degradation mechanism of NicX.

This work was supported
by the National Key Research and Development Program of the Ministry of Science and Technology (2018YFA0901200) and the National Natural Science Foundation of China (31970041).

Link to paper: https://doi.
org/10.
1016/j.
checat.
2022.
11.
019