Chemical Engineering Science ( IF 4.1 ) Pub Date : 2023-05-18 , DOI: 10.1016/j.ces.2023.118885 Nan Ye , Jiaxin Du , Xueke Bian , Xiaomeng Zhao , Chuanbo Zhang , Wenyu Lu
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Ginsenoside F2 (F2) is a rare tetracyclic triterpenoid saponin with a variety of bioactivities, such as anti-cancer, anti-inflammatory, antioxidant etc. In this study, we constructed the biosynthetic pathway of F2 in Saccharomyces cerevisiae by expressing UDP-glycosyltransferase 1 (UGT1) and glycosyltransferase yojK1 (GTK1) in a high protopanaxadiol (PPD) production strain WLT-MVA5. Subsequently, we fused the two glycosyltransferases and modified the endogenous pathways of S. cerevisiae, increasing the titer of F2 to 56.31 mg/L. Then, a combined semi-rational design and directed evolution method was applied to engineer GTK1 and a mutant GTK1F81W/F178S was finally obtained which further increased the F2 titer to 86.80 mg/L. Finally, the best performance strain was applied to scale up in the 5 L bioreactor and F2 titer reached 375.38 mg/L. This research realizes the high-production of F2 in S. cerevisiae and provides a reference method for the biosynthesis of other triterpenoid saponins.
中文翻译:
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通过途径和 UDP-糖基转移酶工程在酿酒酵母中异源合成人参皂苷 F2
人参皂苷 F2 (F2) 是一种稀有的四环三萜皂苷,具有抗癌、抗炎、抗氧化等多种生物活性。本研究通过表达 UDP-糖基转移酶 1 构建了酿酒酵母中 F2 的生物合成途径(UGT1) 和糖基转移酶 yojK1 (GTK1) 在原人参二醇 (PPD) 高产菌株 WLT-MVA5 中。随后,我们融合了两种糖基转移酶并修改了酿酒酵母的内源途径,将 F2 的滴度提高到 56.31 mg/L。然后,将半理性设计和定向进化相结合的方法应用于工程 GTK1 和突变体 GTK1 F81W/F178S最终获得进一步增加 F2 滴度至 86.80 mg/L。最后,将性能最好的菌株应用于5 L生物反应器中放大,F2滴度达到375.38 mg/L。该研究实现了酿酒酵母F2的高产,为其他三萜皂甙的生物合成提供了参考方法。