Mechanism of Action of Flavin-Dependent Halogenases.
Mechanism of Action of Flavin-Dependent Halogenases.
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DOI：
10.1021/acscatal.2c05231
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发表时间：
2022-12-16
期刊：
影响因子：
12.9
通讯作者：
Scrutton, Nigel S.
中科院分区：
文献类型：
--
作者：
Barker, Rhys D.;Yu, Yuqi;De Maria, Leonardo;Johannissen, Linus O.;Scrutton, Nigel S.
要合理设计黄素依赖性卤化酶（FDHs）的底物范围和选择性，必须首先了解活性位点的反应机理和底物相互作用。长期以来，人们一直知道FDH通过在天然底物Trp的C7处的亲电芳香取代来实现区域选择性，但关键活性位点Lys残基的确切作用仍然不明确。在辅因子结合位点形成次氯酸（HOCl）是通过分子氧和单个氯离子分别与还原的FAD和氢氧化黄素直接反应来实现的。然后HOCl被引导10 μ m进入卤化活性部位。位于此位置的Lys 79被认为可以通过氢键或与HOCl直接反应形成− NH 2Cl+中间体，将HOCl导向Trp C7。在这里，我们提出了最有可能的卤化机制的基础上的分子动力学（MD）模拟和活性位点密度泛函理论“簇”模型的FDH PrnA在复杂的天然底物l-色氨酸，次氯酸，和FAD辅因子。MD模拟与不同的质子化状态的关键活性位点残基表明，Lys 79指导HOCl通过氢键，这是证实了这两个建议的机制的反应曲线的计算。
To rationally engineer the substrate scope and selectivity of flavin-dependent halogenases (FDHs), it is essential to first understand the reaction mechanism and substrate interactions in the active site. FDHs have long been known to achieve regioselectivity through an electrophilic aromatic substitution at C7 of the natural substrate Trp, but the precise role of a key active-site Lys residue remains ambiguous. Formation of hypochlorous acid (HOCl) at the cofactor-binding site is achieved by the direct reaction of molecular oxygen and a single chloride ion with reduced FAD and flavin hydroxide, respectively. HOCl is then guided 10 Å into the halogenation active site. Lys79, located in this site, has been proposed to direct HOCl toward Trp C7 through hydrogen bonding or a direct reaction with HOCl to form an −NH2Cl+ intermediate. Here, we present the most likely mechanism for halogenation based on molecular dynamics (MD) simulations and active-site density functional theory “cluster” models of FDH PrnA in complex with its native substrate l-tryptophan, hypochlorous acid, and the FAD cofactor. MD simulations with different protonation states for key active-site residues suggest that Lys79 directs HOCl through hydrogen bonding, which is confirmed by calculations of the reaction profiles for both proposed mechanisms.
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影响因子：
5.7
作者：
de Carvalho CC
通讯作者：
de Carvalho CC
影响因子：
16.6
作者：
Flecks, Silvana;Patallo, Eugenio R.;Zhu, Xiaofeng;Ernyei, Aliz J.;Seifert, Gotthard;Schneider, Alexander;Dong, Changjiang;Naismith, Janies H.;van Pee, Karl-Heinz
通讯作者：
van Pee, Karl-Heinz
影响因子：
14.8
作者：
Coin, Irene;Beyermann, Michael;Bienert, Michael
通讯作者：
Bienert, Michael
影响因子：
2.9
作者：
Bitto, Eduard;Huang, Yu;Phillips, George N., Jr.
通讯作者：
Phillips, George N., Jr.
色氨酸7-卤代酶（PRNA）结构提出了一种区域选择性氯化的机制。
DOI：
10.1126/science.1116510
发表时间：
2005-09-30
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Dong C;Flecks S;Unversucht S;Haupt C;van Pée KH;Naismith JH
通讯作者：
Naismith JH