Enzymatic Plasticity Inspired by the Diterpene Cyclase CotB2
Enzymatic Plasticity Inspired by the Diterpene Cyclase CotB2
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受二萜环化酶 CotB2 启发的酶促可塑性
DOI：
10.1021/acschembio.0c00645
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发表时间：
2020-10-16
影响因子：
4
通讯作者：
Wu, Ruibo
中科院分区：
文献类型：
--
作者：
Tang, Xiaowen;Zhang, Fan;Wu, Ruibo
酶可塑性是一个现代术语，指酶的功能转换，对于酶活性的重新设计具有重要意义。细菌二萜环化酶CotB2是一种典型的塑料酶，其天然形式精确地进行化学反应，而其突变体则使催化功能大大多样化。为了揭示 CotB2 酶催化的奥秘，人们做出了许多努力。然而，精确化学和立体选择性的催化细节和调控机制仍然难以捉摸。在这项工作中，采用多尺度模拟来阐明线性底物生物环化成具有5-8-5稠环支架的最终产物cyclooctat-9-en-7-ol的机制，并且还讨论了反应性碳阳离子中间体过早猝灭产生的脱轨产物。两个主要的调节因素，即口袋中芳香族残基或极性残基的局部静电稳定效应以及活性位点的整体特征（包括口袋轮廓和口袋疏水性），决定了 CotB2 的酶促可塑性。对代表性大戟科植物和真菌二萜环化酶（RcCS和PaFS）的进一步比较研究表明，口袋可塑性和产物多样性之间存在相关性，这启发了未来酶产物的初步预测和合理的二萜环化酶的重新设计。
Enzymatic plasticity, as a modern term referring to the functional conversion of an enzyme, is significant for enzymatic activity redesign. The bacterial diterpene cyclase CotB2 is a typical plastic enzyme by which its native form precisely conducts a chemical reaction while its mutants diversify the catalytic functions drastically. Many efforts have been made to disclose the mysteries of CotB2 enzyme catalysis. However, the catalytic details and regulatory mechanism toward the precise chemo- and stereo-selectivity are still elusive. In this work, multiscale simulations are employed to illuminate the biocydization mechanisms of the linear substrate into the final product cyclooctat-9-en-7-ol with a 5-8-5 fused ring scaffold, and the derailment products arising from the premature quenching of reactive carbocation intermediates are also discussed. The two major regulatory factors, local electrostatic stabilization effects from aromatic residues or polar residue in pocket and global features of active site including pocket-contour and pocket-hydrophobicity, are responsible for the enzymatic plasticity of CotB2. Further comparative studies of representative Euphorbiaceae and fungal diterpene cyclase (RcCS and PaFS) show a correlation between pocket plasticity and product diversity, which inspires a tentative enzyme product prediction and the rational diterpene cyclases' reengineering in the future.