构建酿酒酵母细胞工厂从头合成倍半萜类化合物<italic style="font-style: italic">α</italic>-新丁香三环烯和<italic style="font-style: italic">β</italic>-石竹烯

李晓东; 杨成帅; 王平平; 严兴; 周志华

doi:10.12211/2096-8280.2021-014

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构建酿酒酵母细胞工厂从头合成倍半萜类化合物α-新丁香三环烯和β-石竹烯

研究论文 | 更新时间：2026-03-18

- 构建酿酒酵母细胞工厂从头合成倍半萜类化合物α-新丁香三环烯和β-石竹烯
- Production of sesquiterpenoids α-neoclovene and β-caryophyllene by engineered Saccharomyces cerevisiae
- 合成生物学 2021年2卷第5期页码：792-803
- 作者机构：
  
  1.中国科学院分子植物科学卓越创新中心，合成生物学重点实验室，上海　200032
  2.中国科学院大学，北京　100049
  3.中国科学院深圳先进技术研究院，深圳合成生物学创新研究院，中国科学院定量工程生物学重点实验室，广东　深圳　518055
- 作者简介：
  
  李晓东（1992—），男，博士研究生。研究方向为天然产物合成生物学。
  周志华（1966—），女，研究员，博士，博士生导师。研究方向为天然化合物合成生物学、丝状真菌分子生物学及基因组编辑技术等。
- 基金信息：
  
  国家重点研发计划(2019YFA0904300);国家自然科学基金(31921006)
- DOI：10.12211/2096-8280.2021-014
  中图分类号： Q939.9;Q789
- 收稿：2021-01-31，
  
  修回：2021-03-10，
  
  纸质出版：2021-10-31
- 稿件说明：
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李晓东，杨成帅，王平平，严兴，周志华. 构建酿酒酵母细胞工厂从头合成倍半萜类化合物α-新丁香三环烯和β-石竹烯［J］. 合成生物学， 2021， 2（5）： 792-803

LI Xiaodong， YANG Chengshuai， WANG Pingping， YAN Xing， ZHOU Zhihua. Production of sesquiterpenoids α-neoclovene and β-caryophyllene by engineered Saccharomyces cerevisiae［J］. Synthetic Biology Journal， 2021， 2（5）： 792-803
李晓东，杨成帅，王平平，严兴，周志华. 构建酿酒酵母细胞工厂从头合成倍半萜类化合物α-新丁香三环烯和β-石竹烯［J］. 合成生物学， 2021， 2（5）： 792-803 DOI： 10.12211/2096-8280.2021-014.

LI Xiaodong， YANG Chengshuai， WANG Pingping， YAN Xing， ZHOU Zhihua. Production of sesquiterpenoids α-neoclovene and β-caryophyllene by engineered Saccharomyces cerevisiae［J］. Synthetic Biology Journal， 2021， 2（5）： 792-803 DOI： 10.12211/2096-8280.2021-014.

摘要

倍半萜类化合物（sesquiterpenoids）

-新丁香三环烯（

-neoclovene）和

-石竹烯（

-caryophyllene）都是人参挥发油中的主要组成成分，不仅具有重要的药用开发价值，而且在高能量密度生物能源的开发中也受到关注。然而，它们在人参和其他植物中含量均十分稀少且难以分离纯化，其开发与应用研究严重滞后。本研究利用天然产物合成生物学的方法，以BY4742为出发菌株构建了高产倍半萜类化合物前体FPP的通用底盘菌株SQTBY03。在此底盘菌株中异源表达了密码子优化的植物内生真菌

Hypoxylon

sp. EC38来源的多产物倍半萜合酶元件基因

ec38

和黄花蒿（

Artemisia annua

）来源的倍半萜合酶元件基因

QHS1

，分别构建了合成

-新丁香三环烯和

-石竹烯的两个倍半萜细胞工厂NCVBY01和CPLBY01。细胞工厂NCVBY01的主要倍半萜产物为

-新丁香三环烯（

39%），细胞工厂CPLBY01的主要产物为

-石竹烯（

96%）。它们的摇瓶发酵产量分别达到25.8 mg/L和250.4 mg/L。通过在1.3 L发酵罐中进行批次补料发酵，细胞工厂NCVBY01合成

-新丁香三环烯的产量达到了487.1 mg/L，为目前首次在微生物细胞工厂中合成

-新丁香三环烯；CPLBY01合成

-石竹烯的产量达到2949.1 mg/L，为目前报道的最高水平。上述研究成果为

-新丁香三环烯和

-石竹烯的规模化制备提供新的途径。

Abstract

Sesquiterpenoids

-neoclovene and

-caryophyllene are major components in volatile oils from

Panax ginseng

which have been demonstrated to play important roles in antibacteria

antitumor and cardiovascular protection. Moreover

they have attracted attentions for potential use as biofuels with high-energy-density. However

the industrial production of

-neoclovene and

-caryophyllene as well as other sesquiterpenoids are mainly relied on extraction from plant materials

which is too costly for applications at a large scale. Currently

this challenge could be addressed by advances in synthetic biology for natural product biosynthesis. Through hetero

logously assembling and integrating of their biosynthetic pathways into microbial chassis cells

targeted natural compounds from plants could be produced by microbial fermentation in a sustainable

low-cost and large-scale way. In this study

by comparing the production potential of sesquiterpenes between different

Saccharomyces cerevisiae

strains

and followed by enhancing the endogenous mevalonate pathway

a yeast sesquiterpene chassis strain (SQTBY03) with an increase of 458 times in farnesyl pyrophosphate production was constructed. Then by inserting the codon-optimized sesquiterpene synthase gene

ec38-cs

from the endophytic fungi

Hypoxylon

sp. EC38 and the codon-optimized caryophyllene synthase gene

QHS1

from

Artemisia annua

into SQTBY03

respectively

we built yeast cell factories NCVBY01 and CPLBY01 for

de novo

production of

-neoclovene and

-caryophyllene at their titers of 25.8 mg/L and 250.4 mg/L

respectively

in shake flasks. Furthermore

fed-batch fermentation using NCVBY01 and CPLBY01 resulted in the

de novo

production of 487.1 mg/L

-neoclovene and 2949.1 mg/L

-caryophyllene from glucose. It is also possible to further chemically catalyze

-caryophyllene to produce

-neoclovene. Our work provides strategies for the sustainable production of

-neoclovene and

-caryophyllene from glucose through microbial fermentation

which would benefit their applications as medicine and other functional products. In addition

our yeast chassis for the sesquiterpene production could offer a platform for the sustainable production of other valuable sesquiterpenoids

via

synthetic biology approach.

关键词

Keywords

references

RU W W , WANG D L , XU Y P , et al . Chemical constituents and bioactivities of Panax ginseng (C. A. Mey.) [J ] . Drug Discoveries and Therapeutics , 2015 , 9 ( 1 ): 23 - 32 .

GUO M K , SHAO S , WANG D D , et al . Recent progress in polysaccharides from Panax ginseng C. A. Meyer [J ] . Food and Function , 2020 , 12 ( 2 ): 494 - 518 .

JIANG R , SUN L W , WANG Y B , et al . Chemical composition, and cytotoxic, antioxidant and antibacterial activities of the essential oil from ginseng leaves [J ] . Natural Product Communications , 2014 , 9 ( 6 ): 865 - 868 .

赵岩 , 王红 , 蔡恩博 , 等 . 人参挥发油化学成分及其主要活性成分聚乙炔醇类药理作用研究进展 [J ] . 中国药房 , 2017 , 28 ( 13 ): 1856 - 1859 .

ZHAO Y , WANG H , CAI E B , et al . Research progress on chemical constituents of essential oil from Panax ginseng and pharmacological effects of polyacetylenol [J ] . China Pharmacy , 2017 , 28 ( 13 ): 1856 - 1859 .

彭雪 , 李超英 . 人参挥发油研究 [J ] . 吉林中医药 , 2017 , 37 ( 1 ): 71 - 74 .

PENG X , LI C Y . Advances in the research on volatile oil from ginseng [J ] . Jilin Journal of Traditional Chinese Medicine , 2017 , 37 ( 1 ): 71 - 74 .

IN H C . Volatile compounds of ginseng ( Panax sp.): a review [J ] . Journal of the Korean Society for Applied Biological Chemistry , 2015 , 58 ( 1 ): 67 - 75 .

JUNG J I , KIM E J , KWON G T , et al . β -Caryophyllene potently inhibits solid tumor growth and lymph node metastasis of B16F10 melanoma cells in high-fat diet-induced obese C57BL/6N mice [J ] . Carcinogenesis , 2015 , 36 ( 9 ): 1028 - 1039 .

SOTTO A D , MANCINELLI R , GULLI M , et al . Chemopreventive potential of caryophyllane sesquiterpenes: an overview of preliminary evidence [J ] . Cancers , 2020 , 12 ( 10 ): 3034 .

LEGAULT J , PICHETTE A . Potentiating effect of β -caryophyllene on anticancer activity of α -humulene, isocaryophyllene and paclitaxel [J ] . Journal of Pharmacy and Pharmacology , 2010 , 59 ( 12 ): 1643 - 1647 .

EDWARDS T , MOSES C , DRYER F . Evaluation of combustion performance of alternative aviation fuels [C ] // 46th AIAA/ASME/SAE/ASEE Joint Propulsion Conference & Exhibit . Nashville, TN . Reston, Virginia : AIAA , 2010 .

HARVEY B G , MERRIMAN W W , KOONTZ T A . High-density renewable diesel and jet fuels prepared from multicyclic sesquiterpanes and a 1-hexene-derived synthetic paraffinic kerosene [J ] . Energy & Fuels , 2015 , 29 ( 4 ): 2431 - 2436 .

LIU H B , LU X Y , HU Y H , et al . Chemical constituents of Panax ginseng and Panax notoginseng explain why they differ in therapeutic efficacy [J ] . Pharmacological Research , 2020 , 161 : 1052 - 1063 .

GALANIE S , THODEY K , TRENCHARD I J , et al . Complete biosynthesis of opioids in yeast [J ] . Science , 2015 , 349 ( 6252 ): 1095 - 1100 .

PADDON C J , WESTFALL P J , PITERA D J , et al . High-level semi-synthetic production of the potent antimalarial artemisinin [J ] . Nature , 2013 , 496 ( 7446 ): 528 - 532 .

YAN X , FAN Y , WEI W , et al . Production of bioactive ginsenoside compound K in metabolically engineered yeast [J ] . Cell Research , 2014 , 24 ( 6 ): 770 - 773 .

PING Y , LI X D , YOU W J , et al . De novo production of the plant-derived tropine and pseudotropine in yeast [J ] . ACS Synthetic Biology , 2019 , 8 ( 6 ): 1257 - 1262 .

BAI Y F , YIN H , BI H P , et al . De novo biosynthesis of Gastrodin in Escherichia coli [J ] . Metabolic Engineering , 2016 , 35 : 138 - 147 .

WU W H , TRAN W , TAATJES C , et al . Rapid discovery and functional characterization of terpene synthases from four endophytic Xylariaceae [J ] . PLoS One , 2016 , 11 ( 2 ): e0146983 .

CAI Y , JIA J W , CROCK J , et al . A cDNA clone for beta-caryophyllene synthase from Artemisia annua [J ] . Phytochemistry , 2002 , 61 ( 5 ): 523 - 529 .

GIETZ R D , SCHIESTL R H . Large-scale high-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method [J ] . Nature Protocols , 2007 , 2 ( 1 ): 38 - 41 .

WANG P P , WEI W , YE W , et al . Synthesizing ginsenoside Rh2 in Saccharomyces cerevisiae cell factory at high-efficiency [J ] . Cell Discovery , 2019 , 5 : 5 .

PARAMASIVAN K , RAJAGOPAL K , MUTTURI S . Studies on squalene biosynthesis and the standardization of its extraction methodology from Saccharomyces cerevisiae [J ] . Applied Biochemistry and Biotechnology , 2019 , 187 ( 3 ): 691 - 707 .

IGNEA C , TRIKKA F A , NIKOLAIDIS A K , et al . Efficient diterpene production in yeast by engineering Erg20p into a geranylgeranyl diphosphate synthase [J ] . Metabolic Engineering , 2015 , 27 : 65 - 75 .

APEL A R , D'ESPAUX L , WEHRS M , et al . A Cas9-based toolkit to program gene expression in Saccharomyces cerevisiae [J ] . Nucleic Acids Research , 2017 , 45 ( 1 ): 496 - 508 .

PILAURI V , BEWLEY M , DIEP C , et al . Gal80 dimerization and the yeast GAL gene switch [J ] . Genetics , 2005 , 169 ( 4 ): 1903 - 1914 .

WESTFALL P , PITERA D J , LENIHAN J R , et al . Production of amorphadiene in yeast, and its conversion to dihydroartemisinic acid, precursor to the antimalarial agent artemisinin [J ] . Proceedings of the National Academy of Sciences of the United States of America , 2012 , 109 ( 3 ): E111 - 8 .

YANG J M , NIE Q J . Engineering Escherichia coli to convert acetic acid to β -caryophyllene [J ] . Microbial Cell Factory , 2016 , 15 ( 5 ): 74 - 82 .

WU W H , LIU F , DIVIS R W . Engineering Escherichia coli for the production of terpene mixture enriched in caryophyllene and caryophyllene alcohol as potential aviation fuel compounds [J ] . Metabolic Engineering Communications , 2018 , 5 ( 6 ): 13 - 21 .

PARKER W , RAPHAEL R A , ROBERTS J S . Neoclovene a novel rearrangement product of caryophyllene [J ] . Tetrahedron Letters , 1965 , 6 ( 27 ): 2313 - 2315 .

MCKILLOP T F W , MARTIN J , PARKER W , et al . The synthesis of neoclovene [J ] . Journal of the Chemical Society , 1971 ( 2 ): 162 - 162 .

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构建酿酒酵母细胞工厂从头合成倍半萜类化合物α-新丁香三环烯和β-石竹烯

Production of sesquiterpenoids α-neoclovene and β-caryophyllene by engineered Saccharomyces cerevisiae

DOI：10.12211/2096-8280.2021-014

摘要

Abstract

关键词

Keywords

references