Theoretical Study on the Mechanism of the Oxygen Activation Process in Cysteine Dioxygenase Enzymes
Theoretical Study on the Mechanism of the Oxygen Activation Process in Cysteine Dioxygenase Enzymes
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DOI：
10.1021/ja107514f
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发表时间：
2011-03-23
影响因子：
15
通讯作者：
de Visser, Sam P.
中科院分区：
文献类型：
--
作者：
Kumar, Devesh;Thiel, Walter;de Visser, Sam P.
半胱氨酸双加氧酶(CDO)是一种对人类健康至关重要的酶，参与了有毒半胱氨酸的生物降解，从而调节体内半胱氨酸的浓度。该酶属于非血红素铁双加氧酶类，利用分子氧将两个氧原子转移到半胱氨酸上，形成半胱氨酸亚磺酸产物。该反应的机理目前存在争议，晶体研究表明过硫酸盐中间体存在于催化循环中。为了解决这一争议，我们使用一个酶单体和一个大的QM活性区域，对CDO酶激活底物进行了量子力学/分子力学(QM/MM)计算。我们发现了一个分步机理，铁(II)-超氧配合物的远端氧原子攻击半胱氨酸的硫原子形成环状结构，然后氧氧键断裂，形成与铁(IV)-氧配合物结合的亚硫氧化物。亚砜在第二个氧原子转移到底物之前发生旋转，生成半胱氨酸亚磺酸产物。该反应通过多态反应模式在几个低位自旋态表面上进行。它开始于铁(II)-超氧基反应物的单重态，然后主要在五重态和三重态表面进行。超氧基团对半胱氨酸硫原子的初始和决定速率的攻击包括从单重态到五重态的自旋态跨越。我们还研究了通过过硫酸盐中间体的另一种机制，即底物结合口袋中氢键相互作用的重新排列。然而，这种近端氧原子攻击半胱氨酸硫原子的替代机制被计算为高能途径，因此，过硫酸盐中间体不太可能参与CDO酶的催化循环。
Cysteine dioxygenase (CDO) is a vital enzyme for human health involved in the biodegradation of toxic cysteine and thereby regulation of the cysteine concentration in the body. The enzyme belongs to the group of nonheme iron dioxygenases and utilizes molecular oxygen to transfer two oxygen atoms to cysteinate to form cysteine sulfinic acid products. The mechanism for this reaction is currently disputed, with crystallographic studies implicating a persulfenate intermediate in the catalytic cycle. To resolve the dispute we have performed quantum mechanics/molecular mechanics (QM/MM) calculations on substrate activation by CDO enzymes using an enzyme monomer and a large QM active region. We find a stepwise mechanism, whereby the distal oxygen atom of the iron(II)-superoxo complex attacks the sulfur atom of cysteinate to form a ring structure, followed by dioxygen bond breaking and the formation of a sulfoxide bound to an iron(IV)-oxo complex. A sulfoxide rotation precedes the second oxygen atom transfer to the substrate to give cysteine sulfinic acid products. The reaction takes place on several low-lying spin-state surfaces via multistate reactivity patterns. It starts in the singlet ground state of the iron(II)-superoxo reactant and then proceeds mainly on the quintet and triplet surfaces. The initial and rate-determining attack of the superoxo group on the cysteinate sulfur atom involves a spin-state crossing from singlet to quintet. We have also investigated an alternative mechanism via a persulfenate intermediate, with a realignment of hydrogen bonding interactions in the substrate binding pocket. However, this alternative mechanism of proximal oxygen atom attack on the sulfur atom of cysteinate is computed to be a high-energy pathway, and therefore, the persulfenate intermediate is unlikely to participate in the catalytic cycle of CDO enzymes.