Mn(II) Oxidation by the Multicopper Oxidase Complex Mnx: A Binuclear Activation Mechanism
Mn(II) Oxidation by the Multicopper Oxidase Complex Mnx: A Binuclear Activation Mechanism
复制标题
多铜氧化酶复合物 Mnx 对 Mn(II) 的氧化：双核激活机制
DOI：
10.1021/jacs.7b02771
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发表时间：
2017
影响因子：
15
通讯作者：
Spiro, Thomas G.
中科院分区：
文献类型：
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作者：
Soldatova, Alexandra V.;Tao, Lizhi;Romano, Christine A.;Stich, Troy A.;Casey, William H.;Britt, R. David;Tebo, Bradley M.;Spiro, Thomas G.
细菌蛋白质复合物Mnx包含多铜氧化酶（MCO）MnxG，其不寻常地通过Mn（III）中间体催化Mn（II）到MnO 2生物矿物的双电子氧化。虽然Mn（III）/Mn（II）和Mn（IV）/Mn（III）还原电位预计较高，但我们发现MnxG 1型Cu 2+（电子受体）的还原电位较低，为0.38 V（相对于正常氢电极，pH 7.8）。事实上，1型Cu 2+在不存在分子氧的情况下不被Mn（II）还原，表明底物氧化需要活化步骤。我们通过电子吸收光谱研究了酶的作用机制，并利用化学计量学分析将酶催化的MnO 2形成与MnO 2纳米颗粒老化分开。纳米粒子老化的时间过程是成核和颗粒生长的特征;这些过程的速率遵循预期的Mn（II）浓度和温度的依赖关系，但表现出不同的pH最佳值。酶促时间过程是S形的，在周转之前发出激活步骤的信号。Mn（II）浓度和pH值的依赖性的前一个滞后期表明弱Mn（II）的结合。活化步骤通过pKa> 8.6的去质子化实现，其被分配给Mn（II）结合的H2O;其诱导增加Mn（II）亲和力的构象变化（与高活化能106 kJ/mol一致）。Mnx的活化被认为是通过在底物位点形成氢氧化物桥连的双核配合物Mn（II）（μ-OH）Mn（II）来降低Mn（III/II）的还原电位，使其低于1型Cu（II/I）的还原电位。营业额被认为是合作依赖于两个锰（II），并通过PKA7.6双去质子化。有人提出，周转产生Mn（III）（μ-OH）2 Mn（III）中间体，该中间体继续进行酶产物，可能是Mn（IV）（μ-O）2 Mn（IV）或低聚物，随后使MnO 2纳米颗粒成核。我们的结论是，锰利用锰多核化学，以促进否则困难的氧化反应，以及生物矿化。Mn（III/IV）转化步骤的机理在随附的论文中阐明。
The bacterial protein complex Mnx contains a multicopper oxidase (MCO) MnxG that, unusually, catalyzes the two-electron oxidation of Mn(II) to MnO2biomineral, via a Mn(III) intermediate. Although Mn(III)/Mn(II) and Mn(IV)/Mn(III) reduction potentials are expected to be high, we find a low reduction potential, 0.38 V (vs Normal Hydrogen Electrode, pH 7.8), for the MnxG type 1 Cu2+, the electron acceptor. Indeed the type 1 Cu2+is not reduced by Mn(II) in the absence of molecular oxygen, indicating that substrate oxidation requires an activation step. We have investigated the enzyme mechanism via electronic absorption spectroscopy, using chemometric analysis to separate enzyme-catalyzed MnO2formation from MnO2nanoparticle aging. The nanoparticle aging time course is characteristic of nucleation and particle growth; rates for these processes followed expected dependencies on Mn(II) concentration and temperature, but exhibited different pH optima. The enzymatic time course is sigmoidal, signaling an activation step, prior to turnover. The Mn(II) concentration and pH dependence of a preceding lag phase indicates weak Mn(II) binding. The activation step is enabled by a pKa> 8.6 deprotonation, which is assigned to Mn(II)-bound H2O; it induces a conformation change (consistent with a high activation energy, 106 kJ/mol) that increases Mn(II) affinity. Mnx activation is proposed to decrease the Mn(III/II) reduction potential below that of type 1 Cu(II/I) by formation of a hydroxide-bridged binuclear complex, Mn(II)(μ-OH)Mn(II), at the substrate site. Turnover is found to depend cooperatively on two Mn(II) and is enabled by a pKa7.6 double deprotonation. It is proposed that turnover produces a Mn(III)(μ-OH)2Mn(III) intermediate that proceeds to the enzyme product, likely Mn(IV)(μ-O)2Mn(IV) or an oligomer, which subsequently nucleates MnO2nanoparticles. We conclude that Mnx exploits manganese polynuclear chemistry in order to facilitate an otherwise difficult oxidation reaction, as well as biomineralization. The mechanism of the Mn(III/IV) conversion step is elucidated in an accompanying paper.