摘要
人参皂苷Rh_(2)是人参、三七、西洋参等名贵中药材中珍稀活性成分,具有抗癌、提高人体免疫力等多种重要药理活性。然而人参皂苷Rh_(2)在源植物中含量极低,传统的获取方式有较大的局限性。该研究拟应用合成生物学技术开发酿酒酵母细胞工厂来低成本发酵生产人参皂苷Rh_(2),首先以高产原人参二醇(protopanaxadiol,PPD)的菌株LPTA为底盘菌,将三七来源的3位糖基转移酶基因Pn1-31,与酵母UDP-葡萄糖供给模块基因: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)一同插入酵母染色体的EGH1位点,成功获得生产17.10 mg·g^(-1)人参皂苷Rh_(2)的工程菌LPTA-RH2。通过对菌株的代谢物检测,发现目标产物人参皂苷Rh_(2)的合成前体PPD存在大量积累。为了进一步提高工程菌人参皂苷Rh_(2)的产量,研究对UDP-葡萄糖供给模块及人参皂苷Rh_(2)合成模块进行强化,构建出人参皂苷Rh_(2)摇瓶产量可达36.26 mg·g^(-1),含量占酵母细胞干重的3.63%的工程菌株LPTA-RH2-T,相较于最初改造的菌株LPTA-RH2,最终人参皂苷Rh_(2)的产量提升了112.11%,转化效率提升65.14%。该研究为进一步获得产业级人参皂苷Rh_(2)细胞工厂提供重要基础。
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).
作者
石玉松
王冬
李荣生
张学礼
戴住波
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)
出处
《中国中药杂志》
CAS
CSCD
北大核心
2022年第3期651-658,共8页
China Journal of Chinese Materia Medica
基金
国家重点研发计划项目(2020YFA0908000)。
