α/β-水解酶折叠处从水解催化到氧解催化的进化适应,Chemical Science

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α/β-水解酶折叠处从水解催化到氧解催化的进化适应
Chemical Science ( IF 7.6 ) Pub Date : 2023-09-18 , DOI: 10.1039/d3sc03044j
Soi Bui ₁ , Sara Gil-Guerrero ₂ , Peter van der Linden ₃ , Philippe Carpentier _{4,

5} , Matteo Ceccarelli _{6,

7} , Pablo G Jambrina ₂ , Roberto A Steiner _{1,

8}

Affiliation

蛋白质折叠适应新的酶反应是一个基本的进化过程。降解N-杂芳族底物的不依赖辅因子的加氧酶属于 α/β-水解酶 (ABH) 折叠超家族，通常不催化氧化反应。在这里，我们将常氧和高氧条件下的晶体学分析与分子动力学和量子力学计算相结合，以研究其原型 1- H -3-羟基-4-氧代喹哪啶 2,4-双加氧酶 (HOD) 成员。O ₂定位到发生催化的“氧阴离子孔”是不利的事件，并且双氧和水之间对该位点的直接竞争由“亲核弯头”残基调节。与有机底物结合位点重叠的疏水口袋可以充当近端双氧储存器。冷冻阱加压允许确定具有底物类似物和结合在氧阴离子孔处的O _{2的三元复合物的结构。}理论计算表明O ₂方向与结合的有机配体的电荷耦合。当1- H -3-羟基-4-氧代喹哪啶不带电荷时，O ₂与其分子轴沿着配体的C2-C4方向结合，与晶体结构完全一致。His-Asp 二联体对其 3-OH 基团的去质子化触发底物激活，使 O ₂旋转约 60°。这种几何形状最大化了基底和O ₂之间的电荷转移，从而削弱了后者的双键。电子密度转移至 O ₂ (π*) 轨道可通过决定速率的系间窜越促进过氧化物中间体的形成。我们的工作详细描述了进化如何重新利用 ABH 折叠结构及其简单的催化机制来实现不依赖于金属的氧化。

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Evolutionary adaptation from hydrolytic to oxygenolytic catalysis at the α/β-hydrolase fold

Protein fold adaptation to novel enzymatic reactions is a fundamental evolutionary process. Cofactor-independent oxygenases degrading N-heteroaromatic substrates belong to the α/β-hydrolase (ABH) fold superfamily that typically does not catalyze oxygenation reactions. Here, we have integrated crystallographic analyses under normoxic and hyperoxic conditions with molecular dynamics and quantum mechanical calculations to investigate its prototypic 1-H-3-hydroxy-4-oxoquinaldine 2,4-dioxygenase (HOD) member. O₂ localization to the “oxyanion hole”, where catalysis occurs, is an unfavorable event and the direct competition between dioxygen and water for this site is modulated by the “nucleophilic elbow” residue. A hydrophobic pocket that overlaps with the organic substrate binding site can act as a proximal dioxygen reservoir. Freeze-trap pressurization allowed the structure of the ternary complex with a substrate analogue and O₂ bound at the oxyanion hole to be determined. Theoretical calculations reveal that O₂ orientation is coupled to the charge of the bound organic ligand. When 1-H-3-hydroxy-4-oxoquinaldine is uncharged, O₂ binds with its molecular axis along the ligand's C2–C4 direction in full agreement with the crystal structure. Substrate activation triggered by deprotonation of its 3-OH group by the His-Asp dyad, rotates O₂ by approximately 60°. This geometry maximizes the charge transfer between the substrate and O₂, thus weakening the double bond of the latter. Electron density transfer to the O₂(π*) orbital promotes the formation of the peroxide intermediate via intersystem crossing that is rate-determining. Our work provides a detailed picture of how evolution has repurposed the ABH-fold architecture and its simple catalytic machinery to accomplish metal-independent oxygenation.

更新日期：2023-09-18

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