Shi Yu-Song, Wang Dong, Li Rong-Sheng, Zhang Xue-Li, Dai Zhu-Bo
School of Biology and Biological Engineering, South China University of Technology Guangzhou 510006, China Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences Tianjin 300308, China Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences Tianjin 300308, China.
Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences Tianjin 300308, China Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences Tianjin 300308, China.
Zhongguo Zhong Yao Za Zhi. 2022 Feb;47(3):651-658. doi: 10.19540/j.cnki.cjcmm.20210914.101.
Ginsenoside Rh_2 is a rare active ingredient in precious Chinese medicinal materials such as Ginseng Radix et Rhizoma, Notoginseng Radix et Rhizoma, and Panacis Quinquefolii Radix. It has important pharmacological activities such as anti-cancer and improving human immunity. However, due to the extremely low content of ginsenoside Rh_2 in the source plants, the traditional way of obtaining it has limitations. This study intended to apply synthetic biological technology to develop a cell factory of Saccharomyces cerevisiae to produce Rh_2 by low-cost fermentation. First, we used the high protopanaxadiol(PPD)-yielding strain LPTA as the chassis strain, and inserted the Panax notoginseng enzyme gene Pn1-31, together with yeast UDP-glucose supply module genes[phosphoglucose mutase 1(PGM1), α-phosphoglucose mutase(PGM2), and uridine diphosphate glucose pyrophosphorylase(UGP1)], into the EGH1 locus of yeast chromosome. The engineered strain LPTA-RH2 produced 17.10 mg·g(-1) ginsenoside Rh_2. This strain had low yield of Rh_2 while accumulated much precursor PPD, which severely restricted the application of this strain. In order to further improve the production of ginsenoside Rh_2, we strengthened the UDP glucose supply module and ginsenoside Rh_2 synthesis module by engineered strain LPTA-RH2-T. The shaking flask yield of ginsenoside Rh_2 was increased to 36.26 mg·g(-1), which accounted for 3.63% of the dry weight of yeast cells. Compared with those of the original strain LPTA-RH2, the final production and the conversion efficiency of Rh_2 increased by 112.11% and 65.14%, respectively. This study provides an important basis for further obtaining the industrial-grade cell factory for the production of ginsenoside Rh_2.
人参皂苷Rh_2是人参、三七、西洋参等名贵中药材中罕见的活性成分。它具有抗癌和提高人体免疫力等重要药理活性。然而，由于源植物中人参皂苷Rh_2的含量极低，传统的获取方式存在局限性。本研究旨在应用合成生物技术开发酿酒酵母细胞工厂，通过低成本发酵生产Rh_2。首先，我们以高产原人参二醇（PPD）的菌株LPTA为底盘菌株，将三七酶基因Pn1-31与酵母UDP-葡萄糖供应模块基因[磷酸葡萄糖变位酶1（PGM1）、α-磷酸葡萄糖变位酶（PGM2）和尿苷二磷酸葡萄糖焦磷酸化酶（UGP1）]一起插入酵母染色体的EGH1位点。工程菌株LPTA-RH2产生了17.10 mg·g(-1)的人参皂苷Rh_2。该菌株Rh_2产量低，同时积累了大量前体PPD，严重限制了该菌株的应用。为了进一步提高人参皂苷Rh_2的产量，我们通过工程菌株LPTA-RH2-T强化了UDP葡萄糖供应模块和人参皂苷Rh_2合成模块。人参皂苷Rh_2的摇瓶产量提高到36.26 mg·g(-1)，占酵母细胞干重的3.63%。与原始菌株LPTA-RH2相比，Rh_2的最终产量和转化效率分别提高了112.11%和65.14%。本研究为进一步获得生产人参皂苷Rh_2的工业级细胞工厂提供了重要依据。
