文献类型：期刊文献

英文题名：Function Switch of a Fungal Sesterterpene Synthase through Molecular Dynamics Simulation Assisted Alteration of an Aromatic Residue Cluster in the Active Pocket of PfNS

作者：Zhang, Weiyan[1];Wang, Xinye[1,2];Zhu, Guoliang[1];Zhu, Bin[1];Peng, Kaitong[1];Hsiang, Tom[3];Zhang, Lixin[1];Liu, Xueting[1]

机构：[1]East China Univ Sci & Technol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[2]Ludong Univ, Sch Life Sci, Yantai 264025, Shandong, Peoples R China;[3]Univ Guelph, Sch Environm Sci, 50 Stone Rd East, Guelph, ON N1G 2W1, Canada

年份：2024

卷号：63

期号：37

外文期刊名：ANGEWANDTE CHEMIE-INTERNATIONAL EDITION

收录：;EI(收录号：20243316872192);WOS:【SCI-EXPANDED(收录号:WOS:001288477600001)】；

基金：This work was supported by the National Natural Science Foundation of China (21977029, 81903529, 31720103901, 22307037), the National Key Research and Development Program of China (2020YFA090032 and 2022YFC2105400), the China Postdoctoral Science Foundation funded project (2019M661403), the 111 Project (B18022), the Science and Technology Commission of Shanghai Municipality (21NL2600100), and the Open Project Funding of the State Key Laboratory of Bioreactor Engineering.

语种：英文

外文关键词：aromatic residue cluster; sesterterpene synthase; molecular dynamics; mutagenesis; terpenoids

摘要：Terpene synthases (TPSs) play pivotal roles in generating diverse terpenoids through complex cyclization pathways. Protein engineering of TPSs offers a crucial approach to expanding terpene diversity. However, significant potential remains untapped due to limited understanding of the structure-function relationships of TPSs. In this investigation, using a joint approach of molecular dynamics simulations-assisted engineering and site-directed mutagenesis, we manipulated the aromatic residue cluster (ARC) of a bifunctional terpene synthase (BFTPS), Pestalotiopsis fici nigtetraene synthase (PfNS). This led to the discovery of previously unreported catalytic functions yielding different cyclization patterns of sesterterpenes. Specifically, a quadruple variant (F89A/Y113F/W193L/T194W) completely altered PfNS ' s function, converting it from producing the bicyclic sesterterpene nigtetraene to the tricyclic ophiobolin F. Additionally, analysis of catalytic profiles by double, triple, and quadruple variants demonstrated that the ARC functions as a switch, unprecedently redirecting the production of 5/11 bicyclic (Type B) sesterterpenes to 5/15 bicyclic (Type A) ones. Molecular dynamics simulations and theozyme calculations further elucidated that, in addition to cation-pi interactions, C-H & sdot;& sdot;& sdot;pi interactions also play a key role in the cyclization patterns. This study offers a feasible strategy in protein engineering of TPSs for various industrial applications. A combined strategy involving molecular dynamics simulations and site-directed mutagenesis to manipulate the aromatic residue cluster (ARC) of PfNS protein resulted in the discovery of previously unreported catalytic functions, such as the conversion between bicyclic and tricyclic structures and the transition from Type B to Type A function, thus resulting in the production of new sesterterpenes. image