Purified particulate methane monooxygenase from Methylococcus capsulatus (Bath) is a dimer with both mononuclear copper and a copper-containing cluster
Purified particulate methane monooxygenase from Methylococcus capsulatus (Bath) is a dimer with both mononuclear copper and a copper-containing cluster
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DOI：
10.1073/pnas.0536703100
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发表时间：
2003-04-01
影响因子：
11.1
通讯作者：
Rosenzweig, AC
中科院分区：
文献类型：
--
作者：
Lieberman, RL;Shrestha, DB;Rosenzweig, AC
颗粒甲烷单加氧酶（AMMO）是一种膜结合酶，在嗜甲烷细菌中催化甲烷氧化为甲醇。了解这种酶如何在环境温度和压力下使甲烷羟基化具有基本的化学和潜在的商业重要性。然而，溶解和纯化活性pMMO的困难导致了关于其生物化学和生物物理性质的相互矛盾的报道。我们已经从荚膜甲基球菌（Bath）中纯化了pMMO并检测了活性。纯化的酶分子量约为200 kDa，可能对应于α（2）/β（2）γ（2）多肽排列。每个200-kDa pMMO复合物含有4.8 +/- 0.8个铜离子和1.5 +/- 0.7个铁离子。对应于40-60%的总铜的电子顺磁共振光谱参数是一致的单核2型铜网站的存在。X射线吸收近边光谱表明，纯化的pMMO是Cu（I）和Cu（II）氧化态的混合物。最后，扩展的X射线吸收精细结构数据与氧/氮配体和2.57埃的Cu-Cu相互作用最佳拟合，为pMMO中的含铜簇提供了直接证据。
Particulate methane monooxygenase (AMMO) is a membrane-bound enzyme that catalyzes the oxidation of methane to methanol in methanotropic bacteria. Understanding how this enzyme hydroxylates methane at ambient temperature and pressure is of fundamental chemical and potential commercial importance. Difficulties in solubilizing and purifying active pMMO have led to conflicting reports regarding its biochemical and biophysical properties, however. We have purified pMMO from Methylococcus capsulatus (Bath) and detected activity. The purified enzyme has a molecular mass of approximate to200 kDa, probably corresponding to an alpha(2)/beta(2)gamma(2) polypeptide arrangement. Each 200-kDa pMMO complex contains 4.8 +/- 0.8 copper ions and 1.5 +/- 0.7 iron ions. Electron paramagnetic resonance spectroscopic parameters corresponding to 40-60% of the total copper are consistent with the presence of a mononuclear type 2 copper site. X-ray absorption near edge spectra indicate that purified pMMO is a mixture of Cu(I) and Cu(II) oxidation states. Finally, extended x-ray absorption fine structure data are best fit with oxygen/nitrogen ligands and a 2.57-Angstrom Cu-Cu interaction, providing direct evidence for a copper-containing cluster in pMMO.