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学科主题: 物理化学
题名: Modular Pathway Engineering of Diterpenoid Synthases and the Mevalonic Acid Pathway for Miltiradiene Production
作者: Zhou, Yongjin J.1, 6;  Gao, Wei3, 4;  Rong, Qixian3;  Jin, Guojie1, 6;  Chu, Huiying2;  Liu, Wujun1;  Yang, Wei1;  Zhu, Zhiwei1, 6;  Li, Guohui2;  Zhu, Guofeng5;  Huang, Luqi3;  Zhao, Zongbao K.1
通讯作者: HuangLuqi
刊名: JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
发表日期: 2012-02-15
DOI: 10.1021/ja2114486
卷: 134, 期:6, 页:3234-3241
收录类别: SCI
文章类型: Article
部门归属: 11
项目归属: 1816;1106
产权排名: 1,1
WOS标题词: Science & Technology ;  Physical Sciences
类目[WOS]: Chemistry, Multidisciplinary
研究领域[WOS]: Chemistry
英文摘要: Microbial production can be advantageous over the extraction of phytoterpenoids from natural plant sources, but it remains challenging to rationally and rapidly access efficient pathway variants. Previous engineering attempts mainly focused on the mevalonic acid (MVA) or methyl-D-erythritol phosphate (MEP) pathways responsible for the generation of precursors for terpenoids biosynthesis, and potential interactions between diterpenoids synthases were unexplored. Miltiradiene, the product of the stepwise conversion of (E,E,E)-geranylgeranyl diphosphate (GGPP) catalyzed by diterpene synthases SmCPS and SmKSL, has recently been identified as the precursor to tanshionones, a group of abietane-type norcliterpenoids rich in the Chinese medicinal herb Salvia miltiorrhiza. Here, we present the modular pathway engineering (MOPE) strategy and its application for rapid assembling synthetic miltiradiene pathways in the yeast Saccharomyces cerevisiae. We predicted and analyzed the molecular interactions between SmCPS and SmKSL, and engineered their active sites into close proximity for enhanced metabolic flux channeling to miltiradiene biosynthesis by constructing protein fusions. We show that the fusion of SmCPS and SmKSL, as well as the fusion of BTS1 (GGPP synthase) and ERG20 (farnesyl diphosphate synthase), led to significantly improved miltiradiene production and reduced byproduct accumulation. The MOPE strategy facilitated a comprehensive evaluation of pathway variants involving multiple genes, and, as a result, our best pathway with the diploid strain YJ2X reached miltiradiene titer of 365 mg/L in a 15-L bioreactor culture. These results suggest that terpenoids synthases and the precursor supplying enzymes should be engineered systematically to enable an efficient microbial production of phytoterpenoids.
关键词[WOS]: SACCHAROMYCES-CEREVISIAE ;  METABOLIC ENZYMES ;  YEAST ;  OVERPRODUCTION ;  EXPRESSION ;  REDUCTASE ;  BIOSYNTHESIS ;  ORGANIZATION ;  COMPLEXES ;  COLI
语种: 英语
WOS记录号: WOS:000301161500065
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/117888
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

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作者单位: 1.Chinese Acad Sci, Dalian Inst Chem Phys, Div Biotechnol, Dalian 116023, Peoples R China
2.Chinese Acad Sci, Dalian Inst Chem Phys, State Key Lab Mol React Dynam, Dalian 116023, Peoples R China
3.CACMS, Inst Chinese Mat Med, Beijing 100700, Peoples R China
4.Capital Med Univ, Sch Tradit Chinese Med, Beijing 100069, Peoples R China
5.CAMS, Inst Pathogen Biol, Beijing 100730, Peoples R China
6.Chinese Acad Sci, Grad Univ, Beijing 100049, Peoples R China

Recommended Citation:
Zhou, Yongjin J.,Gao, Wei,Rong, Qixian,et al. Modular Pathway Engineering of Diterpenoid Synthases and the Mevalonic Acid Pathway for Miltiradiene Production[J]. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY,2012,134(6):3234-3241.
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