Crystal structures of the soluble methane monooxygenase hydroxylase from Methylococcus capsulatus (Bath) demonstrating geometrical variability at the dinuclear iron active site.
Crystal structures of the soluble methane monooxygenase hydroxylase from Methylococcus capsulatus (Bath) demonstrating geometrical variability at the dinuclear iron active site.
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DOI：
10.1021/ja003240n
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发表时间：
2001-01
影响因子：
15
通讯作者：
D. Whittington;S. Lippard
中科院分区：
文献类型：
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作者：
D. Whittington;S. Lippard
甲烷氧化成甲醇是在可溶性甲烷单加氧酶羟基酶(MMOH)中的羧酸桥联的双核铁中心上进行的。以前对MMOH和相关的核糖核苷酸还原酶R2亚基的结构研究已经证明，连接催化铁原子的谷氨酸残基发生了羧基转移。这些转变被认为具有重要的机制含义。最近的动力学和理论研究也强调了活性中心的氢键和pH效应的重要性。本文报道了在pH为6.2、7.0和8.5的条件下，二铁(II)、二铁(III)和混价Fe(II)Fe(III)氧化态的MMOH的晶体结构。研究这些结构是为了描绘MMOH活性部位可能的运动范围，并确定氢键相互作用，这可能对理解酶的催化作用很重要。我们的结果首次提出了混合价状态下双铁中心的观点，并表明酶中亚铁离子的不稳定程度增加。根据氧化还原状态和在双铁(II)状态下邻近金属中心的一个α螺旋的扭曲，还鉴定了Asn214在活性中心附近的交替构象。这些变化改变了催化核心附近蛋白质的表面，并可能影响到活性部位的小分子可及性，以及甲烷单加氧酶系统中蛋白质组分的相互作用。总的来说，这些结果有助于解释之前的光谱观察，并为酶的催化提供了新的见解。
The oxidation of methane to methanol is performed at carboxylate-bridged dinuclear iron centers in the soluble methane monooxygenase hydroxylase (MMOH). Previous structural studies of MMOH, and the related R2 subunit of ribonucleotide reductase, have demonstrated the occurrence of carboxylate shifts involving glutamate residues that ligate the catalytic iron atoms. These shifts are thought to have important mechanistic implications. Recent kinetic and theoretical studies have also emphasized the importance of hydrogen bonding and pH effects at the active site. We report here crystal structures of MMOH from Methylococcus capsulatus (Bath) in the diiron(II), diiron(III), and mixed-valent Fe(II)Fe(III) oxidation states, and at pH values of 6.2, 7.0, and 8.5. These structures were investigated in an effort to delineate the range of possible motions at the MMOH active site and to identify hydrogen-bonding interactions that may be important in understanding catalysis by the enzyme. Our results present the first view of the diiron center in the mixed-valent state, and they indicate an increased lability for ferrous ions in the enzyme. Alternate conformations of Asn214 near the active site according to redox state and a distortion in one of the alpha-helices adjacent to the metal center in the diiron(II) state have also been identified. These changes alter the surface of the protein in the vicinity of the catalytic core and may have implications for small-molecule accessibility to the active site and for protein component interactions in the methane monooxygenase system. Collectively, these results help to explain previous spectroscopic observations and provide new insight into catalysis by the enzyme.