Nature of the intermediate formed in the reduction of O2 to H2O at the trinuclear copper cluster active site in native laccase
Nature of the intermediate formed in the reduction of O2 to H2O at the trinuclear copper cluster active site in native laccase
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DOI：
10.1021/ja0114052
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发表时间：
2002-05-29
影响因子：
15
通讯作者：
Solomon, EI
中科院分区：
文献类型：
--
作者：
Lee, SK;George, SD;Solomon, EI
多铜氧化酶含有至少四个铜原子，并在三核铜簇上催化O-2到H2O的四电子还原。一种中间体，称为天然中间体，已被捕获的快速冷冻淬灭技术从漆树漆酶时，完全还原的形式与分子氧反应。该中间体被描述为与三核铜簇合物结合的氧自由基，其中一个Cu位点被还原，然而，XAS表明，在该中间体中所有的铜原子都被氧化。EXAFS，多频EPR，和VTVH MCD的组合已被用来了解这种完全氧化的三核铜簇与完全氧化的静息形式的酶。确定在天然中间体中，簇的所有铜原子通过完全O2还原的产物桥接。相反，静止形式具有与其他磁隔离的簇的一个铜原子（T2 Cu）。天然中间体衰变为静止的氧化形式，其速率太慢而不能处于催化循环中。因此，天然中间体似乎是催化相关的完全氧化形式的酶，其在催化中的作用被认为是。
The multicopper oxidases contain at least four copper atoms and catalyze the four-electron reduction of O-2 to H2O at a trinuclear copper cluster. An intermediate, termed native intermediate, has been trapped by a rapid freeze-quench technique from Rhus vernicifera laccase when the fully reduced form reacts with dioxygen. This intermediate had been described as an oxygen-radical bound to the trinuclear copper cluster with one Cu site reduced, XAS, however, shows that all copper atoms are oxidized in this intermediate. A combination of EXAFS, multifrequency EPR, and VTVH MCD has been used to understand how this fully oxidized trinuclear Cu cluster relates to the fully oxidized resting form of the enzyme. It is determined that in the native intermediate all copper atoms of the cluster are bridged by the product of full 02 reduction. In contrast, the resting form has one copper atom of the cluster (the T2 Cu) magnetically isolated from the others. The native intermediate decays to the resting oxidized form with a rate that is too slow to be in the catalytic cycle. Thus, the native intermediate appears to be the catalytically relevant fully oxidized form of the enzyme, and its role in catalysis is considered.