Plant Cell Reports

, Volume 36, Issue 3, pp 437–445 | Cite as

CYP716A179 functions as a triterpene C-28 oxidase in tissue-cultured stolons of Glycyrrhiza uralensis

  • Keita Tamura
  • Hikaru Seki
  • Hideyuki Suzuki
  • Mareshige Kojoma
  • Kazuki Saito
  • Toshiya MuranakaEmail author
Original Article


Key message

CYP716A179, a cytochrome P450 monooxygenase expressed predominantly in tissue-cultured stolons of licorice ( Glycyrrhiza uralensis ), functions as a triterpene C-28 oxidase in the biosynthesis of oleanolic acid and betulinic acid.


Cytochrome P450 monooxygenases (P450s) play key roles in the structural diversification of plant triterpenoids. Among these, the CYP716A subfamily, which functions mainly as a triterpene C-28 oxidase, is common in plants. Licorice (Glycyrrhiza uralensis) produces bioactive triterpenoids, such as glycyrrhizin and soyasaponins, and relevant P450s (CYP88D6, CYP72A154, and CYP93E3) have been identified; however, no CYP716A subfamily P450 has been isolated. Here, we identify CYP716A179, which functions as a triterpene C-28 oxidase, by RNA sequencing analysis of tissue-cultured stolons of G. uralensis. Heterologous expression of CYP716A179 in engineered yeast strains confirmed the production of oleanolic acid, ursolic acid, and betulinic acid from β-amyrin, α-amyrin, and lupeol, respectively. The transcript level of CYP716A179 was about 500 times higher in tissue-cultured stolons than in intact roots. Oleanolic acid and betulinic acid were consistently detected only in tissue-cultured stolons. The discovery of CYP716A179 helps increase our understanding of the mechanisms of tissue-type-dependent triterpenoid metabolism in licorice and provides an additional target gene for pathway engineering to increase the production of glycyrrhizin in licorice tissue cultures by disrupting competing pathways.


Glycyrrhiza uralensis Licorice P450 RNA-seq Triterpenoid 



α-Amyrin synthase


β-Amyrin synthase


Cycloartenol synthase


Coding sequence


Cytochrome P450 reductase


Expressed sequence tag


Lupeol synthase


Oxidosqualene cyclase



We are very grateful to the Takeda Garden for Medicinal Plant Conservation, Kyoto, Japan (Takeda Pharmaceutical Company Limited) for the supply of licorice plants. We thank Dr. David R. Nelson (University of Tennessee) for the naming of CYP716A179 according to the P450 nomenclature. We also thank Mr. Tsutomu Hosouchi and Ms. Sayaka Shinpo (Kazusa DNA Research Institute) for technical support with the Illumina sequencing, Dr. Kazuto Mannen (Kazusa DNA Research Institute) and Dr. Ryosuke Sano (Nara Institute of Science and Technology) for technical advice with bioinformatics tools, Dr. Kiyoshi Ohyama (Tokyo Institute of Technology) for providing oleanolic aldehyde, ursolic aldehyde and betulinic aldehyde standards, and Dr. Ery Odette Fukushima (Osaka University) for helpful discussion on phylogenetic analysis of CYP716A subfamily enzymes. This work was supported by the “Health and Labour Sciences Research Grant” on the enhancement of “Comprehensive Medicinal Plant Database”, JSPS KAKENHI Grant Number JP26450123 and research funding from the Yamada Science Foundation to H. Seki, JSPS KAKENHI Grant Number JP15H04485 to TM, and the Doctor 21 scholarship from the Yoshida Scholarship Foundation to KT.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

299_2016_2092_MOESM1_ESM.docx (2.6 mb)
Supplementary material (DOCX 2697 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Keita Tamura
    • 1
  • Hikaru Seki
    • 1
  • Hideyuki Suzuki
    • 2
  • Mareshige Kojoma
    • 3
  • Kazuki Saito
    • 4
  • Toshiya Muranaka
    • 1
    Email author
  1. 1.Department of Biotechnology, Graduate School of EngineeringOsaka UniversitySuitaJapan
  2. 2.Department of Research and DevelopmentKazusa DNA Research InstituteKisarazuJapan
  3. 3.Faculty of Pharmaceutical SciencesHealth Sciences University of HokkaidoTobetsuJapan
  4. 4.Graduate School of Pharmaceutical SciencesChiba UniversityChibaJapan

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