Metabolic and Biomolecular Engineering National Research Laboratory, Systems Metabolic Engineering and Systems Healthcare Cross-Generation Collaborative Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), Institute for the BioCentury, Korea Advanced Institute of Science and Technology, Yuseong-gu, 34141 Daejeon, Republic of Korea.
Metabolic and Biomolecular Engineering National Research Laboratory, Systems Metabolic Engineering and Systems Healthcare Cross-Generation Collaborative Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), Institute for the BioCentury, Korea Advanced Institute of Science and Technology, Yuseong-gu, 34141 Daejeon, Republic of Korea;
Proc Natl Acad Sci U S A. 2020 Dec 1;117(48):30328-30334. doi: 10.1073/pnas.2017483117. Epub 2020 Nov 16.
There is increasing industrial demand for five-carbon platform chemicals, particularly glutaric acid, a widely used building block chemical for the synthesis of polyesters and polyamides. Here we report the development of an efficient glutaric acid microbial producer by systems metabolic engineering of an l-lysine-overproducing BE strain. Based on our previous study, an optimal synthetic metabolic pathway comprising l-lysine monooxygenase () and 5-aminovaleramide amidohydrolase () genes and 4-aminobutyrate aminotransferase () and succinate-semialdehyde dehydrogenase () genes, was introduced into the BE strain. Through system-wide analyses including genome-scale metabolic simulation, comparative transcriptome analysis, and flux response analysis, 11 target genes to be manipulated were identified and expressed at desired levels to increase the supply of direct precursor l-lysine and reduce precursor loss. A glutaric acid exporter encoded by was discovered and overexpressed to further enhance glutaric acid production. Fermentation conditions, including oxygen transfer rate, batch-phase glucose level, and nutrient feeding strategy, were optimized for the efficient production of glutaric acid. Fed-batch culture of the final engineered strain produced 105.3 g/L of glutaric acid in 69 h without any byproduct. The strategies of metabolic engineering and fermentation optimization described here will be useful for developing engineered microorganisms for the high-level bio-based production of other chemicals of interest to industry.
工业界对五碳平台化学品的需求日益增长，特别是戊二酸，它是一种广泛用于合成聚酯和聚酰胺的基础化工原料。在这里，我们通过对赖氨酸高产 BE 菌株进行系统代谢工程改造，开发出了一种高效的戊二酸微生物生产菌。基于我们之前的研究，引入了包含 l-赖氨酸单加氧酶（）和 5-氨基戊酰胺水解酶（）基因以及 4-氨基丁酸转氨酶（）和琥珀酸半醛脱氢酶（）基因的优化合成代谢途径。通过包括基因组规模代谢模拟、比较转录组分析和通量响应分析在内的系统分析，确定了 11 个需要操作的目标基因，并在期望水平上进行表达，以增加直接前体 l-赖氨酸的供应并减少前体损失。发现并过表达了编码戊二酸外排泵的基因，以进一步提高戊二酸的产量。优化了发酵条件，包括氧传递速率、分批阶段葡萄糖水平和营养物补料策略，以实现戊二酸的高效生产。最终工程菌株的分批补料培养在 69 小时内生产了 105.3 g/L 的戊二酸，没有任何副产物。这里描述的代谢工程和发酵优化策略将有助于开发用于高水平生物基生产其他工业相关化学品的工程微生物。
