Microbial de novo production of l-serine, which is widely used in a range of cosmetic and pharmaceutical products, has attracted increasing attention due to its environmentally friendly characteristics. Previous pioneering work mainly focused on l-serine anabolism; however, in this study, it was found that l-serine could be reimported through the l-serine uptake system, thus hampering l-serine production. To address this challenge, engineering via deletion of four genes, namely, sdaC, cycA, sstT and tdcC, which have been reported to be involved in l-serine uptake in Escherichia coli, was first carried out in the l-serine producer E. coli ES. Additionally, the effects of these genes on l-serine uptake activity and l-serine production were investigated. The data revealed an abnormal phenomenon regarding serine uptake activity. The serine uptake activity of the ΔsdaC mutant was 0.798 nmol min−1 (mg dry weight) −1 after 30 min, decreasing by 23.34% compared to that of the control strain. However, the serine uptake activity of the single sstT, cycA and tdcC mutants increased by 34.29%, 78.29% and 48.03%, respectively, compared to that of the control strain. This finding may be the result of the increased level of sdaC expression in these mutants. In addition, multigene-deletion strains were constructed based on an sdaC knockout mutant. The ΔsdaCΔsstTΔtdcC mutant strain exhibited 0.253 nmol min−1 (mg dry weight) −1l-serine uptake activity and the highest production titer of 445 mg/L in shake flask fermentation, which was more than three-fold the 129 mg/L production observed for the parent. Furthermore, the ΔsdaCΔsstTΔtdcC mutant accumulated 34.8 g/L l-serine with a yield of 32% from glucose in a 5-L fermenter after 36 h. The results indicated that reuptake of l-serine impairs its production and that an engineered cell with reduced uptake can address this problem and improve the production of l-serine in E. coli.

中文翻译：

---

通过减少摄取的生产的l-丝氨酸改善大肠杆菌的l-丝氨酸形成

从头开始生产微生物化的L-丝氨酸，由于其环保特性，已广泛用于化妆品和药品。先前的开拓性工作主要集中在l-丝氨酸合成代谢上。然而，在这项研究中，发现可以通过l-丝氨酸摄取系统重新导入l-丝氨酸，从而阻碍了l-丝氨酸的产生。为了应对这一挑战，首先在l-丝氨酸生产商E中进行了通过缺失sdaC，cycA，sstT和tdcC四个基因的工程改造，据报道这些基因与大肠杆菌中的l-丝氨酸摄取有关。大肠杆菌 另外，研究了这些基因对l-丝氨酸摄取活性和l-丝氨酸产生的影响。数据揭示了关于丝氨酸摄取活性的异常现象。30分钟后，ΔsdaC突变体的丝氨酸摄取活性为0.798nmol min-1（mg干重）-1，与对照菌株相比降低了23.34％。然而，与对照菌株相比，单个sstT，cycA和tdcC突变体的丝氨酸吸收活性分别增加了34.29％，78.29％和48.03％。这一发现可能是这些突变体中sdaC表达水平提高的结果。另外，基于sdaC敲除突变体构建了多基因缺失菌株。ΔsdaCΔsstTΔtdcC突变菌株在摇瓶发酵中表现出0.253 nmol min-1（mg干重）-1l-丝氨酸吸收活性和最高滴定度445 mg / L，是观察到的129 mg / L产量的三倍以上为父母。此外，ΔsdaCΔsstTΔtdcC突变体累积了34。36 h后，在5 L发酵罐中，葡萄糖的含量为8 g / L l-丝氨酸，产率为32％。结果表明，L-丝氨酸的再摄取会损害其产量，而摄取减少的工程化细胞可以解决该问题并提高大肠杆菌中L-丝氨酸的产量。