Wang Pingping, Wei Yongjun, Fan Yun, Liu Qunfang, Wei Wei, Yang Chengshuai, Zhang Lei, Zhao Guoping, Yue Jianmin, Yan Xing, Zhou Zhihua
CAS-Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, Shanghai 200032, China.
CAS-Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, Shanghai 200032, China; State Key Laboratory of Genetic Engineering, Department of Microbiology, School of Life Sciences and Institute of Biomedical Sciences, Fudan University, Shanghai 200433, China.
Metab Eng. 2015 May;29:97-105. doi: 10.1016/j.ymben.2015.03.003. Epub 2015 Mar 11.
Ginsenosides Rh2 and Rg3 represent promising candidates for cancer prevention and therapy and have low toxicity. However, the concentrations of Rh2 and Rg3 are extremely low in the bioactive constituents (triterpene saponins) of ginseng. Despite the available heterologous biosynthesis of their aglycone (protopanaxadiol, PPD) in yeast, production of Rh2 and Rg3 by a synthetic biology approach was hindered by the absence of bioparts to glucosylate the C3 hydroxyl of PPD. In this study, two UDP-glycosyltransferases (UGTs) were cloned and identified from Panax ginseng. UGTPg45 selectively transfers a glucose moiety to the C3 hydroxyl of PPD and its ginsenosides. UGTPg29 selectively transfers a glucose moiety to the C3 glucose of Rh2 to form a 1-2-glycosidic bond. Based on the two UGTs and a yeast chassis to produce PPD, yeast cell factories were built to produce Rh2 and/or Rg3 from glucose. The turnover number (kcat) of UGTPg29 was more than 2500-fold that of UGTPg45, which might explain the higher Rg3 yield than that of Rh2 in the yeast cell factories. Building yeast cell factories to produce Rh2 or Rg3 from simple sugars by microbial fermentation provides an alternative approach to replace the traditional method of extracting ginsenosides from Panax plants.
人参皂苷Rh2和Rg3是癌症预防和治疗的有潜力的候选药物，且毒性较低。然而，人参生物活性成分（三萜皂苷）中Rh2和Rg3的浓度极低。尽管已在酵母中实现了其苷元（原人参二醇，PPD）的异源生物合成，但由于缺乏将PPD的C3羟基糖基化的生物元件，通过合成生物学方法生产Rh2和Rg3受到阻碍。在本研究中，从人参中克隆并鉴定了两种尿苷二磷酸糖基转移酶（UGT）。UGTPg45选择性地将葡萄糖部分转移到PPD及其人参皂苷的C3羟基上。UGTPg29选择性地将葡萄糖部分转移到Rh2的C3葡萄糖上，形成1-2糖苷键。基于这两种UGT和一个生产PPD的酵母底盘，构建了酵母细胞工厂以从葡萄糖生产Rh2和/或Rg3。UGTPg29的转换数（kcat）是人UGTPg45的2500多倍，这可能解释了酵母细胞工厂中Rg3的产量高于Rh2的原因。通过微生物发酵构建酵母细胞工厂以从单糖生产Rh2或Rg3，为替代从人参植物中提取人参皂苷的传统方法提供了一种途径。
