摘要
人参(Panaxginseng)来源的糖基转移酶Pg UGT74AE2能够催化原人参二醇(protopanaxadiol,PPD)的C-3位羟基发生糖基化反应,生成人参皂苷Rh2,但其催化原人参三醇(protopanaxatriol,PPT)的C-3位羟基发生糖基化反应的研究至今未见报道。本研究构建重组表达质粒p ET-32a-Pg UGT74AE2,转化大肠杆菌Transetta(DE3)感受态细胞,获得重组菌株Transetta-Pg UGT74AE2。通过异丙基-β-D-硫代半乳糖苷(isopropyl-β-Dthiogalactoside, IPTG)诱导表达,获得重组Pg UGT74AE2。建立重组Pg UGT74AE2催化PPT的酶促反应体系,结果表明,Pg UGT74AE2能够催化PPT的C-3位羟基发生糖基化反应,生成产物3-O-β-D-glucopyranosyl-dammar-24-ene-3β,6α,12β,20S-tetraol。本研究通过酶促反应获得C-3位糖基化的新型人参三醇皂苷,可为创新药物研究提供原料。
UDP-glycosyltransferase PgUGT74AE2 from Panax ginseng can transfer a glucose moiety to the free C-3 hydroxyl of protopanaxadiol（PPD） to produce ginsenoside Rh2. However, no report demonstrates that PgUGT74AE2 can transfer a glucose moiety to the free C-3 hydroxyl of protopanaxatriol（PPT） to produce a PPT-type ginsenoside. In this study, the expression plasmid p ET-32 a-PgUGT74AE2 was constructed for expression of the recombinant protein and transferred into Escherichia coli Transetta（DE3） to generate the recombinant strain Transetta-PgUGT74AE2. The recombinant enzyme PgUGT74AE2 was expressed by induction of isopropyl-β-D-thiogalactoside（IPTG）. An in vitro enzymatic reaction system was established with the recombinant enzyme PgUGT74AE2 and the substrate PPT. PgUGT74AE2 catalyzed the glycosylation of the free C-3 hydroxyl of PPT to produce 3-O-β-D-glucopyranosyl-dammar-24-ene-3β,6α,12β,20 S-tetraol, a new PPT-type ginsenoside. This study provides an efficient approach for the biosynthesis of a new PPT-type ginsenoside through in vitro enzymatic reaction, which may pave a way to produce promising lead in drug discovery.
作者
张婷婷
梁会超
巩婷
胡宗风
顾安頔
杨金玲
朱平
ZHANG Ting-ting;LIANG Hui-chao;GONG Ting;HU Zong-feng;GU An-di;YANG Jin-ling;ZHU Ping(State Key Laboratory of Bioactive Substance and Function of Natural Medicines and Key Laboratory of Biosynthesis of Natural Products of National Health Commission of PRC,Institute of Materia Medica,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100050,China)
出处
《药学学报》
CAS
CSCD
北大核心
2018年第9期1565-1570,共6页
Acta Pharmaceutica Sinica
基金
国家自然科学基金资助项目(81673341)
北京市自然科学基金资助项目(7122115)
中国医学科学院医学与健康科技创新工程经费(2016-I2M-3-012)
关键词
人参
糖基转移酶
糖基化
原人参三醇
人参三醇皂苷
Panax ginseng
glycosyltransferase
glycosylation
protopanaxatriol
protopanaxatriol-typeginsenoside
