Molecular basis for short-chain thioester hydrolysis by acyl hydrolase domains in trans -acyltransferase polyketide synthases
Molecular basis for short-chain thioester hydrolysis by acyl hydrolase domains in trans -acyltransferase polyketide synthases
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反式酰基转移酶聚酮合酶中酰基水解酶结构域水解短链硫酯的分子基础
DOI：
10.1101/2023.08.11.552765
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Fage C
中科院分区：
文献类型：
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作者：
Fage C
聚酮合成酶（pks）是一种多结构域的酶装配线，可以从简单的构建块中生物合成多种生物活性天然产物。与顺式酰基转移酶（AT）相比，反式AT PKS依赖于独立的AT结构域将扩展器单元加载到嵌入核心PKS机制的酰基载体蛋白（ACP）结构域上。Trans-AT PKS基因簇也编码酰基水解酶（AH）结构域，预计这些结构域共享AT结构域的整体折叠，但水解来自ACP结构域的异常酰基链，从而确保高效的聚酮生物合成。这些结构域如何专门针对短酰基链，特别是乙酰基，作为硫酯系在底物穿梭ACP结构域上，具有水解活性而不是酰基转移活性，目前尚不清楚。为了回答这些问题，我们解决了AH结构域的第一个结构，并对活性位点变体进行了结构指导的活性分析。我们的研究结果为对辅酶a系链底物的链长控制和选择提供了关键的见解，并阐明了AH和ACP结构域之间的相互作用界面如何有助于识别同源和非同源ACP结构域。将我们的实验结果与分子动力学模拟相结合，可以产生一个数据驱动的AH:ACP结构域复合物模型。我们的研究结果促进了目前对trans- at pks合成聚酮生物的不完全理解，并为未来的生物工程工作提供了基础。
Polyketide synthases (PKSs) are multi-domain enzymatic assembly lines that biosynthesise a wide selection of bioactive natural products from simple building blocks. In contrast to theircis-acyltransferase (AT) counterparts,trans-AT PKSs rely on stand-alone AT domains to load extender units onto acyl carrier protein (ACP) domains embedded in the core PKS machinery.Trans-AT PKS gene clusters also encode acyl hydrolase (AH) domains, which are predicted to share the overall fold of AT domains, but hydrolyse aberrant acyl chains from ACP domains, thus ensuring efficient polyketide biosynthesis. How such domains specifically target short acyl chains, in particular acetyl groups, tethered as thioesters to the substrate-shuttling ACP domains, with hydrolytic rather than acyl transfer activity, has remained unclear. To answer these questions, we solved the first structure of an AH domain and performed structure-guided activity assays on active site variants. Our results offer key insights into chain length control and selection against coenzyme A-tethered substrates, and clarify how the interaction interface between AH and ACP domains contributes to recognition of cognate and non-cognate ACP domains. Combining our experimental findings with molecular dynamics simulations allowed for the production of a data-driven model of an AH:ACP domain complex. Our results advance the currently incomplete understanding of polyketide biosynthesis bytrans-AT PKSs, and provide foundations for future bioengineering efforts.