Department of Chemistry, NC State University, 2620 Yarbrough Drive, Raleigh, NC 27607, USA.
Comparative Medicine Institute, NC State University, 1001 William Moore Drive, Raleigh, NC 27606, USA.
Chembiochem. 2024 Nov 4;25(21):e202400532. doi: 10.1002/cbic.202400532. Epub 2024 Sep 9.
Enzymatic platforms for producing malonyl-CoA-based extender units required for polyketide biosynthesis are often based on malonyl-CoA ligases such as MatB from Rhizobium trifolii and Rhodopseudomonas palustris. However, despite broad interest in the fluorination of polyketides and prior success with engineering MatB homologs, the suitability of MatB for accessing the tertiary substituted fluoromethylmalonyl-CoA needed to produce flurithromycin and solithromycin has not yet been reported. Herein, we report the structure-guided engineering of a MatB homolog to optimize the production of fluoromethylmalonyl-CoA, resulting in a variant with increased conversion and providing a platform to produce a suitable building block mixture for fluorinated macrolide production. Additionally, the mutant demonstrated broad utility for various substituted malonyl-CoAs. The MatB mutant sets the stage to access fluorinated macrolides by coupling it with altered PKS machinery to install fluorinated malonyl-CoA into macrolide scaffolds.
用于聚酮生物合成的基于丙二酰辅酶 A 的延伸单元的酶促平台通常基于丙二酰辅酶 A 连接酶，例如 Rhizobium trifolii 和 Rhodopseudomonas palustris 中的 MatB。然而，尽管人们对聚酮的氟化广泛感兴趣，并且在工程化 MatB 同源物方面取得了成功，但 MatB 用于获得生产 flurithromycin 和 solithromycin 所需的三级取代的氟甲基丙二酰辅酶 A 的适用性尚未得到报道。在此，我们报告了一种 MatB 同源物的结构导向工程改造，以优化氟甲基丙二酰辅酶 A 的生产，从而产生转化率提高的变体，并为生产适合氟化大环内酯生产的合适砌块混合物提供了一个平台。此外，该突变体对各种取代的丙二酰辅酶 A 表现出广泛的用途。MatB 突变体通过与改变的 PKS 机制结合，将其用于将氟化丙二酰辅酶 A 安装到大环内酯支架中，为获得氟化大环内酯奠定了基础。
