Institute of Systems Biotechnology, Saarland University, Saarbrücken, Germany.
Department of Pharmacy, Pharmaceutical Biotechnology, Saarland University, Saarbrücken, Germany.
Biotechnol Bioeng. 2020 Dec;117(12):3858-3875. doi: 10.1002/bit.27537. Epub 2020 Sep 1.
Streptomyces spp. are a rich source for natural products with recognized industrial value, explaining the high interest to improve and streamline the performance of in these microbes. Here, we studied the production of pamamycins, macrodiolide homologs with a high activity against multiresistant pathogenic microbes, using recombinant Streptomyces albus J1074/R2. Talc particles (hydrous magnesium silicate, 3MgO·4SiO ·H O) of micrometer size, added to submerged cultures of the recombinant strain, tripled pamamycin production up to 50 mg/L. Furthermore, they strongly affected morphology, reduced the size of cell pellets formed by the filamentous microbe during the process up to sixfold, and shifted the pamamycin spectrum to larger derivatives. Integrated analysis of transcriptome and precursor (CoA thioester) supply of particle-enhanced and control cultures provided detailed insights into the underlying molecular changes. The microparticles affected the expression of 3,341 genes (56% of all genes), revealing a global and fundamental impact on metabolism. Morphology-associated genes, encoding major regulators such as SsgA, RelA, EshA, Factor C, as well as chaplins and rodlins, were found massively upregulated, indicating that the particles caused a substantially accelerated morphogenesis. In line, the pamamycin cluster was strongly upregulated (up to 1,024-fold). Furthermore, the microparticles perturbed genes encoding for CoA-ester metabolism, which were mainly activated. The altered expression resulted in changes in the availability of intracellular CoA-esters, the building blocks of pamamycin. Notably, the ratio between methylmalonyl CoA and malonyl-CoA was increased fourfold. Both metabolites compete for incorporation into pamamycin so that the altered availability explained the pronounced preference for larger derivatives in the microparticle-enhanced process. The novel insights into the behavior of S. albus in response to talc appears of general relevance to further explore and upgrade the concept of microparticle enhanced cultivation, widely used for filamentous microbes.
链霉菌属是天然产物的丰富来源，具有公认的工业价值，这解释了为什么人们高度关注提高和简化这些微生物的性能。在这里，我们使用重组白色链霉菌 J1074/R2 研究了 Pamamycins 的生产，Pamamycins 是一种具有高抗多药抗性致病微生物活性的大环二萜类似物。微米大小的滑石颗粒（水合硅酸镁，3MgO·4SiO ·H O）添加到重组菌株的液体培养物中，使 Pamamycin 的产量增加了两倍，达到 50mg/L。此外，它们强烈影响形态，将丝状微生物在培养过程中形成的细胞团的大小缩小了六倍，并将 Pamamycin 谱转移到更大的衍生物上。对颗粒增强和对照培养物的转录组和前体（CoA 硫酯）供应的综合分析提供了对潜在分子变化的详细了解。微粒子影响了 3341 个基因（所有基因的 56%）的表达，显示出对新陈代谢的全面和根本影响。形态相关基因，编码主要调节剂如 SsgA、RelA、EshA、Factor C 以及 chaplins 和 rodlins，被大量上调，表明颗粒导致了形态发生的大大加速。与此一致，Pamamycin 簇被强烈上调（高达 1024 倍）。此外，微粒子扰乱了 CoA-酯代谢的基因表达，这些基因主要被激活。这种改变的表达导致细胞内 CoA-酯的可用性发生变化，CoA-酯是 Pamamycin 的构建块。值得注意的是，甲基丙二酰辅酶 A 和丙二酰辅酶 A 的比例增加了四倍。这两种代谢物竞争掺入 Pamamycin 中，因此改变的可用性解释了在微粒子增强过程中对更大衍生物的明显偏好。对白色链霉菌对滑石的反应行为的新见解似乎具有普遍的相关性，可以进一步探索和升级广泛用于丝状微生物的微粒子增强培养的概念。
