Department of Organic Chemistry, Arrhenius Laboratory, Stockholm University, Stockholm, Sweden.
J Inorg Biochem. 2011 Jul;105(7):927-36. doi: 10.1016/j.jinorgbio.2011.03.020. Epub 2011 Apr 5.
Formaldehyde ferredoxin oxidoreductase from Pyrococcus furiosus is a tungsten-dependent enzyme that catalyzes the oxidation of formaldehyde to formic acid. In the present study, quantum chemical calculations are used to elucidate the reaction mechanism of this enzyme. Several possible mechanistic scenarios are investigated with a large model of the active site designed on the basis of the X-ray crystal structure of the native enzyme. Based on the calculations, we propose a new mechanism in which the formaldehyde substrate binds directly to the tungsten ion. W(VI)=O then performs a nucleophilic attack on the formaldehyde carbon to form a tetrahedral intermediate. In the second step, which is calculated to be rate limiting, a proton is transferred to the second-shell Glu308 residue, coupled with a two-electron reduction of the tungsten ion. The calculated barriers for the mechanism are energetically very feasible and in relatively good agreement with experimental rate constants. Three other second-shell mechanisms, including one previously proposed based on experimental findings, are considered but ruled out because of their high barriers.
来源于 Pyrococcus furiosus 的甲醛铁氧还蛋白氧化还原酶是一种依赖钨的酶，能够催化甲醛氧化生成甲酸。在本研究中，我们使用量子化学计算阐明了该酶的反应机制。基于 X 射线晶体结构设计了一个大型活性位点模型，研究了几种可能的机制场景。根据计算结果，我们提出了一个新的机制，其中甲醛底物直接与钨离子结合。W(VI)=O 然后对甲醛碳进行亲核攻击，形成四面体中间体。在第二步中，计算表明这是限速步骤，质子转移到第二壳层的 Glu308 残基上，同时钨离子发生两电子还原。计算得到的机制的能垒在能量上是非常可行的，并且与实验速率常数相对吻合。还考虑了其他三种第二壳层机制，包括一个之前基于实验结果提出的机制，但由于其能垒较高而被排除。
