Bn archetypical extradiol-cleaving catecholic dioxygenase:: the crystal structure of catechol 2,3-dioxygenase (metapyrocatechase) from Pseudomonas putida mt-2
Bn archetypical extradiol-cleaving catecholic dioxygenase:: the crystal structure of catechol 2,3-dioxygenase (metapyrocatechase) from Pseudomonas putida mt-2
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DOI：
10.1016/s0969-2126(99)80006-9
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发表时间：
1999-01-15
期刊：
影响因子：
5.7
通讯作者：
Miki, K
中科院分区：
文献类型：
--
作者：
Kita, A;Kita, S;Miki, K
背景：儿茶酚双加氧酶以传统或外部方式催化邻苯二酚及其衍生物的裂环反应，这些酶在土壤细菌降解环境中的芳香族分子中起着关键作用。邻苯二酚2，3-双加氧酶催化氧与邻苯二酚结合，并通过外环裂解生成2-羟基邻甲酸半醛。恶臭假单胞菌Mt-2产生的邻苯二酚2，3-双加氧酶(MPC)是第一个以纯种形式获得的外源性双加氧酶，并得到了广泛的研究，缺乏MPC结构阻碍了对外源性双加氧酶一般机理的理解。结果：通过多重同象置换法在2.8埃分辨率下确定了MPC的三维结构。这种酶是一个同源四聚体，每个亚基折叠成两个相似的结构域。MPC亚基的结构类似于2，3-二羟基联苯-1，2-双加氧酶，尽管这些酶之间的氨基酸序列同源性很低。结论：MPG目前的结构与两个2，3-二羟基联苯1，2-双加氧酶的结构相结合，揭示了活性中心的保守核心区，包括三个Fe(II)配体(His153，His214和Glu265)，一个酪氨酸(Tyr255)和两个组氨酸(His199和His246)残基。结果表明，外源性双加氧酶在识别不同底物的儿茶酚环状结构和激活氧气方面采用了共同的机制。其中一个保守的组氨酸残基(His199)似乎在催化循环中扮演着重要的角色。
Background: Catechol dioxygenases catalyze the ring cleavage of catechol and its derivatives in either an intradiol or extradiol manner, These enzymes have a key role in the degradation of aromatic molecules in the environment by soil bacteria. Catechol 2,3-dioxygenase catalyzes the incorporation of dioxygen into catechol and the extradiol ring cleavage to form 2-hydroxymuconate semialdehyde. Catechol 2,3-dioxygenase (metapyrocatechase, MPC) from Pseudomonas putida mt-2 was the first extradiol dioxygenase to be obtained in a pure form and has been studied extensively, The lack of an MPC structure has hampered the understanding of the general mechanism of extradiol dioxygenases.Results: The three-dimensional structure of MPC has been determined at 2.8 Angstrom resolution by the multiple isomorphous replacement method. The enzyme is a homotetramer with each subunit folded into two similar domains. The structure of the MPC subunit resembles that of 2,3-dihydroxybiphenyl 1,2-dioxygenase, although there is low amino acid sequence identity between these enzymes. The active-site structure reveals a distorted tetrahedral Fe(II) site with three endogenous ligands (His153, His214 and Glu265), and an additional molecule that is most probably acetone.Conclusions: The present structure of MPG, combined with those of two 2,3-dihydroxybiphenyl 1,2-dioxygenases, reveals a conserved core region of the active site comprising three Fe(II) ligands (His153, His214 and Glu265), one tyrosine (Tyr255) and two histidine (His199 and His246) residues. The results suggest that extradiol dioxygenases employ a common mechanism to recognize the catechol ring moiety of various substrates and to activate dioxygen. One of the conserved histidine residues (His199) seems to have important roles in the catalytic cycle.