School of Biosciences, University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK.
School of Chemistry, University of St Andrews, BMS Building, North Haugh, St Andrews, KY16 9ST, UK.
Sci Rep. 2019 Feb 7;9(1):1542. doi: 10.1038/s41598-018-38038-9.
The mupirocin trans-AT polyketide synthase pathway, provides a model system for manipulation of antibiotic biosynthesis. Its final phase involves removal of the tertiary hydroxyl group from pseudomonic acid B, PA-B, producing the fully active PA-A in a complex series of steps. To further clarify requirements for this conversion, we fed extracts containing PA-B to mutants of the producer strain singly deficient in each mup gene. This additionally identified mupM and mupN as required plus the sequence but not enzymic activity of mupL and ruled out need for other mup genes. A plasmid expressing mupLMNOPVCFU + macpE together with a derivative of the producer P. fluorescens strain NCIMB10586 lacking the mup cluster allowed conversion of PA-B to PA-A. MupN converts apo-mAcpE to holo-form while MupM is a mupirocin-resistant isoleucyl tRNA synthase, preventing self-poisoning. Surprisingly, the expression plasmid failed to allow the closely related P. fluorescens strain SBW25 to convert PA-B to PA-A.
莫匹罗星反式 AT 聚酮合酶途径为抗生素生物合成的操作提供了一个模型系统。其最后阶段涉及从假单胞菌酸 B (PA-B) 中去除三级羟基，产生完全活性的 PA-A，这是一个复杂的系列步骤。为了进一步阐明这一转化的要求，我们将含有 PA-B 的提取物分别喂食给产生菌的每个 mup 基因单一缺陷的突变体。这还确定了 mupM 和 mupN 是必需的，加上 mupL 的序列但没有酶活性，并排除了其他 mup 基因的需求。一个表达 mupLMNOPVCFU + macpE 的质粒与缺乏 mup 簇的荧光假单胞菌 NCIMB10586 衍生菌株一起使用，允许将 PA-B 转化为 PA-A。MupN 将脱辅基-mAcpE 转化为全酶形式，而 MupM 是一种莫匹罗星抗性异亮氨酰 tRNA 合成酶，可防止自身中毒。令人惊讶的是，表达质粒未能使密切相关的荧光假单胞菌 SBW25 转化 PA-B 为 PA-A。
