Université de Lorraine, CNRS, IMoPA, F-54000, Nancy, France.
OCI & BMWZ, Leibniz Universität Hannover, Schneiderberg 38, 30167, Hannover, Germany.
Nat Commun. 2023 Mar 10;14(1):1327. doi: 10.1038/s41467-023-36974-3.
During biosynthesis by multi-modular trans-AT polyketide synthases, polyketide structural space can be expanded by conversion of initially-formed electrophilic β-ketones into β-alkyl groups. These multi-step transformations are catalysed by 3-hydroxy-3-methylgluratryl synthase cassettes of enzymes. While mechanistic aspects of these reactions have been delineated, little information is available concerning how the cassettes select the specific polyketide intermediate(s) to target. Here we use integrative structural biology to identify the basis for substrate choice in module 5 of the virginiamycin M trans-AT polyketide synthase. Additionally, we show in vitro that module 7, at minimum, is a potential additional site for β-methylation. Indeed, analysis by HPLC-MS coupled with isotopic labelling and pathway inactivation identifies a metabolite bearing a second β-methyl at the expected position. Collectively, our results demonstrate that several control mechanisms acting in concert underpin β-branching programming. Furthermore, variations in this control - whether natural or by design - open up avenues for diversifying polyketide structures towards high-value derivatives.
在多模块反式 AT 聚酮合酶的生物合成过程中，聚酮结构空间可以通过将最初形成的亲电β-酮转化为β-烷基来扩展。这些多步转化由酶的 3-羟基-3-甲基戊二酰基合酶盒催化。虽然这些反应的机制方面已经被描绘出来，但关于盒如何选择特定的聚酮中间产物作为目标，知之甚少。在这里，我们使用综合结构生物学来确定维吉尼亚霉素 M 反式 AT 聚酮合酶第 5 个模块中底物选择的基础。此外，我们在体外表明，至少模块 7 是潜在的额外β-甲基化位点。实际上，通过 HPLC-MS 分析与同位素标记和途径失活相结合，鉴定出一种在预期位置带有第二个β-甲基的代谢产物。总的来说，我们的结果表明，几种协同作用的控制机制为β-支化编程提供了支持。此外，这种控制的变化——无论是自然的还是设计的——都为聚酮结构向高价值衍生物的多样化开辟了途径。
