Coze Fabien, Gilard Françoise, Tcherkez Guillaume, Virolle Marie-Joëlle, Guyonvarch Armel
Unité Mixte de Recherche 8621, Institut de Génétique et Microbiologie, Université Paris-Sud, Orsay, France ; Centre National de la Recherche Scientifique, Orsay, France.
Unité Mixte de Recherche 8618, Institut de Biotechnologie des Plantes, Université Paris-Sud, Orsay, France ; Centre National de la Recherche Scientifique, Orsay, France.
PLoS One. 2013 Dec 23;8(12):e84151. doi: 10.1371/journal.pone.0084151. eCollection 2013.
Metabolic Flux Analysis is now viewed as essential to elucidate the metabolic pattern of cells and to design appropriate genetic engineering strategies to improve strain performance and production processes. Here, we investigated carbon flux distribution in two Streptomyces coelicolor A3 (2) strains: the wild type M145 and its derivative mutant M1146, in which gene clusters encoding the four main antibiotic biosynthetic pathways were deleted. Metabolic Flux Analysis and (13)C-labeling allowed us to reconstruct a flux map under steady-state conditions for both strains. The mutant strain M1146 showed a higher growth rate, a higher flux through the pentose phosphate pathway and a higher flux through the anaplerotic phosphoenolpyruvate carboxylase. In that strain, glucose uptake and the flux through the Krebs cycle were lower than in M145. The enhanced flux through the pentose phosphate pathway in M1146 is thought to generate NADPH enough to face higher needs for biomass biosynthesis and other processes. In both strains, the production of NADPH was higher than NADPH needs, suggesting a key role for nicotinamide nucleotide transhydrogenase for redox homeostasis. ATP production is also likely to exceed metabolic ATP needs, indicating that ATP consumption for maintenance is substantial.Our results further suggest a possible competition between actinorhodin and triacylglycerol biosynthetic pathways for their common precursor, acetyl-CoA. These findings may be instrumental in developing new strategies exploiting S. coelicolor as a platform for the production of bio-based products of industrial interest.
代谢通量分析现在被视为阐明细胞代谢模式以及设计合适的基因工程策略以改善菌株性能和生产过程的关键。在此，我们研究了两种天蓝色链霉菌A3(2)菌株中的碳通量分布：野生型M145及其衍生突变体M1146，后者缺失了编码四种主要抗生素生物合成途径的基因簇。代谢通量分析和(13)C标记使我们能够重建两种菌株在稳态条件下的通量图。突变菌株M1146显示出更高的生长速率、通过磷酸戊糖途径的更高通量以及通过回补磷酸烯醇丙酮酸羧化酶的更高通量。在该菌株中，葡萄糖摄取和通过三羧酸循环的通量低于M145。M1146中通过磷酸戊糖途径增强的通量被认为能够产生足够的NADPH以满足对生物量生物合成和其他过程的更高需求。在两种菌株中，NADPH的产生均高于NADPH需求，这表明烟酰胺核苷酸转氢酶在氧化还原稳态中起关键作用。ATP的产生也可能超过代谢对ATP的需求，这表明维持所需的ATP消耗很大。我们的结果进一步表明，放线紫红素和三酰甘油生物合成途径可能会竞争它们的共同前体乙酰辅酶A。这些发现可能有助于开发新策略，利用天蓝色链霉菌作为生产具有工业价值的生物基产品的平台。
