Mechanism of Radical Initiation and Transfer in Class Id Ribonucleotide Reductase Based on Density Functional Theory.
Mechanism of Radical Initiation and Transfer in Class Id Ribonucleotide Reductase Based on Density Functional Theory.
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DOI：
10.1021/acs.inorgchem.2c02926
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发表时间：
2023-01
影响因子：
4.6
通讯作者：
Jinxin Zou;Lu Q. Yang;Wei Feng
中科院分区：
文献类型：
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作者：
Jinxin Zou;Lu Q. Yang;Wei Feng
类核糖核苷酸还原酶（Class Id ribonnucleotide reducase， RNR）是一种新发现的酶，它以处于超氧化状态的二锰辅因子（MnIII/MnIV）作为自由基引发剂。Id类RNR的二锰辅因子处于还原态（无活性），显然是基于Fj-β亚基的晶体结构。然而，ii类RNR的二锰辅因子在氧化状态（活性）下的状态尚不清楚。x波段EPR谱显示，活化的Fj-β亚基存在于两个不同的配合物1和2中。在这项工作中，量子力学/分子力学计算进行了研究类Id RNR。首先，我们确定配合物2含有一个MnIII-（μ-氧）2- mniv簇，配合物1含有一个MnIII-（μ-羟基/μ-氧）- mniv簇。然后，在确定二锰辅助因子的基础上，揭示了Id类RNR中自由基起始和转移的机理。配合物2中的MnIII-(μ-oxo)2- mniv簇没有足够的还原电位直接引发自由基转移。相反，它需要在自由基转移之前被单质子化成MnIII-（μ-羟基/μ-氧）- mniv（配合物1）。μ-oxo的质子化状态可以通过改变蛋白质微环境来调节，这种微环境是由蛋白质的β亚基与α亚基的聚集和分离所诱导的。Fj-β亚基自由基转移链中MnIII-（μ-羟基/μ-氧）-MnIV簇和Trp30簇之间的自由基转移（MnIII/MnIV↔His100↔Asp194↔Trp30↔Arg99）是一种水介导的三质子偶联电子转移，它通过水分子Wat551和桥接μ-羟基配体将质子从Lys71的ε-氨基转移到Glu97的羧基上，经过三步反应。这种新发现的质子耦合电子转移机制与已知的Ia-Ic RNR不同。Lys71的ε-氨基作为质子供体，在自由基转移中起重要作用。
Class Id ribonucleotide reductase (RNR) is a newly discovered enzyme, which employs the dimanganese cofactor in the superoxidized state (MnIII/MnIV) as the radical initiator. The dimanganese cofactor of class Id RNR in the reduced state (inactive) is clearly based on the crystal structure of the Fj-β subunit. However, the state of the dimanganese cofactor of class Id RNR in the oxidized state (active) is not known. The X-band EPR spectra have shown that the activated Fj-β subunit exists in two distinct complexes, 1 and 2. In this work, quantum mechanical/molecular mechanical calculations were carried out to study class Id RNR. First, we have determined that complex 2 contains a MnIII-(μ-oxo)2-MnIV cluster, and complex 1 contains a MnIII-(μ-hydroxo/μ-oxo)-MnIV cluster. Then, based on the determined dimanganese cofactors, the mechanism of radical initiation and transfer in class Id RNR is revealed. The MnIII-(μ-oxo)2-MnIV cluster in complex 2 has not enough reduction potential to initiate radical transfer directly. Instead, it needs to be monoprotonated into MnIII-(μ-hydroxo/μ-oxo)-MnIV (complex 1) before the radical transfer. The protonation state of μ-oxo can be regulated by changing the protein microenvironment, which is induced by the protein aggregation and separation of β subunits with α subunits. The radical transfer between the cluster of MnIII-(μ-hydroxo/μ-oxo)-MnIV and Trp30 in the radical-transfer chain of the Fj-β subunit (MnIII/MnIV ↔ His100 ↔ Asp194 ↔ Trp30 ↔ Arg99) is a water-mediated tri-proton-coupled electron transfer, which transfers proton from the ε-amino group of Lys71 to the carboxyl group of Glu97 via the water molecule Wat551 and the bridging μ-hydroxo ligand through a three-step reaction. This newly discovered proton-coupled electron-transfer mechanism in class Id RNR is different from those reported in the known Ia-Ic RNRs. The ε-amino group of Lys71, which serves as a proton donor, plays an important role in the radical transfer.