PgGH介导原人参二醇型皂苷和次级苷的转化提高人参品质的机制研究

Ginseng has highly medicinal and economic value. However, the regulating mechanism research on fine quality formation of ginseng has not been fully clarified. The composition and contents of ginsenosides are the performance of ginseng real quality. Actually, the pharmacological activities of secondary glycosides are more powerful than that of their precursors, and are also the important factors of fine quality of ginseng. Previous studies have demonstrated that a panaxtrol type ginsenoside can be biotransformated to its secondary glycoside of by glycosidase. We therefore hypothesize that Panax ginseng Glycoside Hydrolase (PgGH) can catalyze hydrolysis of panaxtrol type ginsenoside to its secondary glycosides in Panax ginseng, and thus improves the fine quality formation of ginseng. The present work screens and clones the whole cDNA sequence of PgGH by mining of ginseng database, and then will verify the candidate gene function by prokaryotic expression system and enzymatic reaction in vitro. At the same time, the catalytic effects of PgGH in biotransformation of panaxtrol type ginsenoside will be illustrated by overexpression and RNA interference strategies in vivo. The controlled experiments would be implemented to elucidate the regulatory effects of PgGH in the fine quality formation of ginseng by analysis the relevance of PgGH expression, variety of ginsenoside amount. With the help of a comprehensive chemistry, bioinformatics and molecular genetics method, the project will deeply illuminate the function of PgGH, primarily revealing the mechanism of improving the quality formation of ginseng, which will provide theoretical basis for cultivating fine quality of ginseng and exploring related planting techniques by directed evolving PgGH strategy.

人参有较高的药用和经济价值，但其优良品质形成的调控机制尚不完全清晰。人参皂苷的种类和含量是人参内在品质的体现，其中次级苷具有比原人参皂苷更显著的药理活性，是人参优良品质形成的重要因素。前期研究发现，植物糖苷酶可通过催化原人参二醇型皂苷生物代谢为次级苷。据此我们认为，人参糖苷酶（PgGH）能体内催化原人参二醇型皂苷进行生物代谢产生次级苷，以此促进人参优良品质的形成。本研究拟以人参组培体系为材料，筛选获得PgGH基因全长，通过原核表达、体外酶促等方法进行功能验证；同时从体内PgGH基因过表达和表达抑制两个角度阐明PgGH在二醇型皂苷生物代谢中的作用；并通过分析不同条件下PgGH表达、人参皂苷成分的变化关联规律以明确PgGH的调控机制。本项目综合生物信息学、分子遗传学和化学分析的技术，初步解析PgGH提高人参品质的作用机制，为通过定向进化PgGH表达策略培育优质人参和栽培技术的开发提供理论依据。