厦门大学王斌举团队揭示了膜结合颗粒甲烷单加氧酶中铜中心的价态和反应性。相关研究成果于2023年11月13日发表于国际一流学术期刊《美国化学会杂志》。

颗粒甲烷单加氧酶（pMMO）在催化甲烷转化为甲醇方面发挥着关键作用，构成了甲烷营养细菌C1代谢途径的初始步骤。然而，膜结合的pMMO的结构和催化机制，特别是铜的价态和甲烷氧化的真正活性位点，仍然难以捉摸。

基于最近表征的膜结合pMMO的结构，研究人员进行了广泛的计算研究来解决这些长期存在的问题。研究人员将量子力学/分子力学（QM/MM）分子动力学（MD）模拟结构与低温EM数据进行比较的综合分析表明，在膜环境中，CuC和CuD位点都倾向于保持在Cu（I）价态。此外，Cu（I）同时存在于CuC和CuD位点，导致位于它们之间的配体结合腔显著减少，使其不太可能容纳还原剂分子，如二氢醌（DQH₂）。随后的QM/MM计算表明，在形成H₂O₂和Cu（II）-O物种的过程中，CuD（I）位点在氧活化中比CuC（I）更具反应性。

最后，模拟表明，天然还原剂泛醌醇（CoQH₂）在CuD（I）位点呈现生产性结合构象，但在CuC（I）位置没有。表明膜结合pMMOs的真正活性位点可能是CuD而不是CuC。这些发现阐明了pMMO的催化机制，并强调了膜环境在调节pMMO内铜中心的配位结构和活性中的关键作用。

附：英文原文

Title: Unraveling the Valence State and Reactivity of Copper Centers in Membrane-Bound Particulate Methane Monooxygenase

Author: Wei Peng, Zikuan Wang, Qiaoyu Zhang, Shengheng Yan, Binju Wang

Issue&Volume: November 13, 2023

Abstract: Particulate methane monooxygenase (pMMO) plays a critical role in catalyzing the conversion of methane to methanol, constituting the initial step in the C1 metabolic pathway within methanotrophic bacteria. However, the membrane-bound pMMO’s structure and catalytic mechanism, notably the copper’s valence state and genuine active site for methane oxidation, have remained elusive. Based on the recently characterized structure of membrane-bound pMMO, extensive computational studies were conducted to address these long-standing issues. A comprehensive analysis comparing the quantum mechanics/molecular mechanics (QM/MM) molecular dynamics (MD) simulated structures with cryo-EM data indicates that both the CuC and CuD sites tend to stay in the Cu(I) valence state within the membrane environment. Additionally, the concurrent presence of Cu(I) at both CuC and CuD sites leads to the significant reduction of the ligand-binding cavity situated between them, making it less likely to accommodate a reductant molecule such as durohydroquinone (DQH2). Subsequent QM/MM calculations reveal that the CuD(I) site is more reactive than the CuC(I) site in oxygen activation, en route to H2O2 formation and the generation of Cu(II)–O– species. Finally, our simulations demonstrate that the natural reductant ubiquinol (CoQH2) assumes a productive binding conformation at the CuD(I) site but not at the CuC(I) site. This provides evidence that the true active site of membrane-bound pMMOs may be CuD rather than CuC. These findings clarify pMMO’s catalytic mechanism and emphasize the membrane environment’s pivotal role in modulating the coordination structure and the activity of copper centers within pMMO.

DOI: 10.1021/jacs.3c08834

Source: https://pubs.acs.org/doi/abs/10.1021/jacs.3c08834