Department of Chemistry and Biochemistry , The Ohio State University , 100 W. 18th Avenue , Columbus , Ohio 43210 , United States.
Department of Chemistry , University of Michigan , 930 N. University Avenue , Ann Arbor , Michigan 48109 , United States.
Inorg Chem. 2019 Jul 15;58(14):8969-8982. doi: 10.1021/acs.inorgchem.8b03546. Epub 2019 Feb 21.
Nickel-containing enzymes such as methyl coenzyme M reductase (MCR) and carbon monoxide dehydrogenase/acetyl coenzyme A synthase (CODH/ACS) play a critical role in global energy conversion reactions, with significant contributions to carbon-centered processes. These enzymes are implied to cycle through a series of nickel-based organometallic intermediates during catalysis, though identification of these intermediates remains challenging. In this work, we have developed and characterized a nickel-containing metalloprotein that models the methyl-bound organometallic intermediates proposed in the native enzymes. Using a nickel(I)-substituted azurin mutant, we demonstrate that alkyl binding occurs via nucleophilic addition of methyl iodide as a methyl donor. The paramagnetic Ni-CH species initially generated can be rapidly reduced to a high-spin Ni-CH species in the presence of exogenous reducing agent, following a reaction sequence analogous to that proposed for ACS. These two distinct bioorganometallic species have been characterized by optical, EPR, XAS, and MCD spectroscopy, and the overall mechanism describing methyl reactivity with nickel azurin has been quantitatively modeled using global kinetic simulations. A comparison between the nickel azurin protein system and existing ACS model compounds is presented. Ni-CH Az is only the second example of two-electron addition of methyl iodide to a Ni center to give an isolable species and the first to be formed in a biologically relevant system. These results highlight the divergent reactivity of nickel across the two intermediates, with implications for likely reaction mechanisms and catalytically relevant states in the native ACS enzyme.
含镍酶，如甲基辅酶 M 还原酶 (MCR) 和一氧化碳脱氢酶/乙酰辅酶 A 合酶 (CODH/ACS)，在全球能量转换反应中发挥着关键作用，对碳中心过程有重要贡献。这些酶在催化过程中被认为会循环通过一系列基于镍的有机金属中间体，尽管这些中间体的鉴定仍然具有挑战性。在这项工作中，我们开发并表征了一种含镍的金属蛋白，该蛋白模拟了天然酶中提出的与甲基结合的有机金属中间体。使用镍 (I) 取代的蓝铜蛋白突变体，我们证明烷基结合是通过作为甲基供体的甲基碘的亲核加成发生的。最初生成的顺磁 Ni-CH 物种可以在存在外源还原剂的情况下迅速还原为高自旋 Ni-CH 物种，遵循与 ACS 中提出的反应序列类似的反应序列。这些两种不同的生物有机金属物种已通过光学、EPR、XAS 和 MCD 光谱进行了表征，并使用全局动力学模拟对描述镍蓝铜蛋白与甲基反应性的整体机制进行了定量建模。提出了镍蓝铜蛋白体系与现有 ACS 模型化合物的比较。Ni-CH Az 仅是第二例二电子加成甲基碘到 Ni 中心得到可分离物种的例子，也是第一个在生物学相关体系中形成的例子。这些结果突出了镍在两个中间体中的不同反应性，对可能的反应机制和催化相关状态在天然 ACS 酶中具有重要意义。
