Gongquan Liu1, Yi-Lei Zhao1, Fangyuan He2, Peng Zhang2, Xingyu Ouyang1, Hongzhi Tang3, Ping Xu1. 1. State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic and Developmental Sciences, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, People's Republic of China. 2. National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People's Republic of China. 3. State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic and Developmental Sciences, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, People's Republic of China. tanghongzhi@sjtu.edu.cn.
Abstract
Biodegradation of aromatic and heterocyclic compounds requires an oxidative ring cleavage enzymatic step. Extensive biochemical research has yielded mechanistic insights about catabolism of aromatic substrates; yet much less is known about the reaction mechanisms underlying the cleavage of heterocyclic compounds such as pyridine-ring-containing ones like 2,5-hydroxy-pyridine (DHP). 2,5-Dihydroxypyridine dioxygenase (NicX) from Pseudomonas putida KT2440 uses a mononuclear nonheme Fe(II) to catalyze the oxidative pyridine ring cleavage reaction by transforming DHP into N-formylmaleamic acid (NFM). Herein, we report a crystal structure for the resting form of NicX, as well as a complex structure wherein DHP and NFM are trapped in different subunits. The resting state structure displays an octahedral coordination for Fe(II) with two histidine residues (His265 and His318), a serine residue (Ser302), a carboxylate ligand (Asp320), and two water molecules. DHP does not bind as a ligand to Fe(II), yet its interactions with Leu104 and His105 function to guide and stabilize the substrate to the appropriate position to initiate the reaction. Additionally, combined structural and computational analyses lend support to an apical dioxygen catalytic mechanism. Our study thus deepens understanding of non-heme Fe(II) dioxygenases.
Biodegradation of aromatic and heterocyclic compounds requires an oxidative ring cleavage enzymatic step. Extensive biochemical research has yielded mechanistic insights about catabolism of aromatic substrates; yet much less is known about the reaction mechanisms underlying the cleavage of heterocyclic compounds such as pyridine-ring-containing ones like n class="Chemical">2,5-hydroxy-pyridine (DHP). 2,5-Dihydroxypyridinedioxygenase (NicX) from Pseudomonas putida KT2440 uses a mononuclear nonhemeFe(II) to catalyze the oxidative pyridine ring cleavage reaction by transforming DHP into N-formylmaleamic acid (NFM). Herein, we report a crystal structure for the resting form of NicX, as well as a complex structure wherein DHP and NFM are trapped in different subunits. The resting state structure displays an octahedral coordination for Fe(II) with two histidine residues (His265 and His318), a serine residue (Ser302), a carboxylate ligand (Asp320), and two water molecules. DHP does not bind as a ligand to Fe(II), yet its interactions with Leu104 and His105 function to guide and stabilize the substrate to the appropriate position to initiate the reaction. Additionally, combined structural and computational analyses lend support to an apical dioxygen catalytic mechanism. Our study thus deepens understanding of non-hemeFe(II)dioxygenases.
Authors: Keri L Colabroy; Huili Zhai; Tingfeng Li; Ying Ge; Yang Zhang; Aimin Liu; Steven E Ealick; Fred W McLafferty; Tadhg P Begley Journal: Biochemistry Date: 2005-05-31 Impact factor: 3.162