Iso-Migrastatin (iso-MGS) and lactimidomycin (LTM) are glutarimide-containing polyketide natural products (NPs) that are biosynthesized by homologous acyltransferase (AT)-less type I polyketide synthase (PKS) assembly lines. The biological activities of iso-MGS and LTM have inspired numerous efforts to generate analogues via genetic manipulation of their biosynthetic machinery in both native producers and model heterologous hosts. A detailed understanding of the MGS and LTM AT-less type I PKSs would serve to inspire future engineering efforts while advancing the fundamental knowledge of AT-less type I PKS enzymology. The mgs and ltm biosynthetic gene clusters (BGCs) encode for two discrete ATs of the architecture AT-enoylreductase (AT-ER) and AT-type II thioesterase (AT-TE). Herein, we report the functional characterization of the mgsB and ltmB and the mgsH and ltmH gene products, revealing that MgsB and LtmB function as type II thioesterases (TEs) and MgsH and LtmH are the dedicated trans-ATs for the MGS and LTM AT-less type I PKSs. In vivo and in vitro experiments demonstrated that MgsB was devoid of any AT activity, despite the presence of the conserved catalytic triad of canonical ATs. Cross-complementation experiments demonstrated that MgsH and LtmH are functionally interchangeable between the MGS and LTM AT-less type I PKSs. This work sets the stage for future mechanistic studies of AT-less type I PKSs and efforts to engineer the MGS and LTM AT-less type I PKS assembly lines for novel glutarimide-containing polyketides.

中文翻译：

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异米格拉他汀和乳霉素生物合成中的离散酰基转移酶和硫酯酶

异米格拉他汀 (iso-MGS) 和拉克酰亚胺霉素 (LTM) 是含戊二酰亚胺的聚酮化合物天然产物 (NP)，由同源无酰基转移酶 (AT) 的 I 型聚酮化合物合酶 (PKS) 装配线生物合成。iso-MGS 和 LTM 的生物活性激发了许多努力，通过在本地生产者和模型异源宿主中对其生物合成机制进行遗传操作来产生类似物。对 MGS 和 LTM AT-less I 型 PKS 的详细了解将有助于激发未来的工程工作，同时推进无 AT-I 型 PKS 酶学的基础知识。mgs和ltm生物合成基因簇 (BGC) 编码结构 AT-烯酰还原酶 (AT-ER) 和 AT-II 型硫酯酶 (AT-TE) 的两个离散 AT 。在此，我们报告了mgsB和ltmB以及mgsH和ltmH基因产物的功能表征，揭示了 MgsB 和 LtmB 作为 II 型硫酯酶 (TE) 发挥作用，而 MgsH 和 LtmH 是MGS 和 LTM AT- 的专用反式AT- I 型 PKS 较少。体内和体外实验表明，尽管存在典型 AT 的保守催化三联体，但 MgsB 缺乏任何 AT 活性。交叉互补实验表明，MgsH 和 LtmH 在 MGS 和 LTM AT-less I 型 PKS 之间可以功能互换。这项工作为未来无AT I型PKS的机理研究以及为新型含戊二酰亚胺的聚酮化合物设计MGS和LTM无AT I型PKS装配线奠定了基础。