J Integr Plant Biol ›› 2024, Vol. 66 ›› Issue (8): 1703-1717.DOI: 10.1111/jipb.13724

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  • 收稿日期:2024-04-09 接受日期:2024-05-28 出版日期:2024-08-01 发布日期:2024-08-19

Identification of the cytochrome P450s responsible for the biosynthesis of two types of aporphine alkaloids and their de novo biosynthesis in yeast

Qishuang Li1,2†, Xiang Jiao3†, Xinyi Li2,4†, Wenlong Shi2, Ying Ma2, Xiangmei Tan2, Jingyi Gan2, Jimei Liu5, Jian Yang2, Jian Wang2, Baolong Jin2, Tong Chen2 , Ping Su2, Yujun Zhao2, Yifeng Zhang2, Jinfu Tang2, Guanghong Cui2, Yun Chen3*, Juan Guo2* and Luqi Huang2*    

  1. 1. School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, China
    2. State Key Laboratory for Quality Ensurance and Sustainable Use of Dao‐di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
    3. Department of Life Sciences, Chalmers University of Technology, Gothenburg SE‐41296, Sweden
    4. State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China
    5. State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, The Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
    These authors contributed equally to this work.
    *Correspondences: Yun Chen (yunc@chalmers.se); Juan Guo (guojuanzy@163.com, Juan Guo is fully responsible for the distributions of all materials associated with this article); Luqi Huang (huangluqi01@163.com)
  • Received:2024-04-09 Accepted:2024-05-28 Online:2024-08-01 Published:2024-08-19
  • Supported by:
    This work was supported by the National Key R&D Program of China (2020YFA0908000), the National Natural Science Foundation of China (82011530137, 31961133007), Scientific and technological innovation project of CACMS (CI2023D002, CI2023E002), Key project at central government level: The ability to establish sustainable use of valuable Chinese medicine resources (2060302), and Vetenskapsradet (2018‐06003), Stiftelsen for internationalisering av hogre utbildning och forskning.

Abstract: Aporphine alkaloids have diverse pharmacological activities; however, our understanding of their biosynthesis is relatively limited. Previous studies have classified aporphine alkaloids into two categories based on the configuration and number of substituents of the D-ring and have proposed preliminary biosynthetic pathways for each category. In this study, we identified two specific cytochrome P450 enzymes (CYP80G6 and CYP80Q5) with distinct activities toward (S)-configured and (R)-configured substrates from the herbaceous perennial vine Stephania tetrandra, shedding light on the biosynthetic mechanisms and stereochemical features of these two aporphine alkaloid categories. Additionally, we characterized two CYP719C enzymes (CYP719C3 and CYP719C4) that catalyzed the formation of the methylenedioxy bridge, an essential pharmacophoric group, on the A- and D-rings, respectively, of aporphine alkaloids. Leveraging the functional characterization of these crucial cytochrome P450 enzymes, we reconstructed the biosynthetic pathways for the two types of aporphine alkaloids in budding yeast (Saccharomyces cerevisiae) for the de novo production of compounds such as (R)-glaziovine, (S)-glaziovine, and magnoflorine. This study provides key insight into the biosynthesis of aporphine alkaloids and lays a foundation for producing these valuable compounds through synthetic biology.

Key words: aporphine alkaloids, biosynthesis, CYP719C, CYP80, engineered yeast

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