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<p><i>Rhododendron molle</i> G. Don contains a rich array of secondary metabolites, with grayanane diterpenoids as its representative components which exhibit diverse pharmacological activities, including anti-inflammatory, analgesic, antihypertensive, antiarrhythmic, antiviral and insecticidal properties. It has been reported that the 6/6/6/5 tetracyclic structure of <i>ent</i>-kaurene undergoes an A/B ring epoxide rearrangement, forming the 5/7/6/5 tetracyclic skeleton characteristic of grayanane diterpenoids. To identify CYP450s that may be involved in biosynthetic pathway, we successfully engineered a yeast chassis strain capable of producing 16α-hydroxy-<i>ent</i>-kaurane and <i>ent</i>-kaurene for functional validation. For the first time, we cloned three NADPH-cytochrome P450 reductases (RmCPRs) and identified RmCPR1 as the most active in the yeast system. Moreover, RmKO was cloned and confirmed which catalyzes the conversion of 16α-hydroxy-<i>ent</i>-kaurane into 16α-hydroxy-<i>ent</i>-kaurenoic acid. These findings provide valuable insights that may facilitate the discovery of more CYP450s from <i>Rhododendron molle</i> G. Don and lay the foundation for future elucidation of the biosynthetic pathway of grayanane diterpenoids.</p>

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Cloning and functional study of NADPH-cytochrome P450 reductases and ent-kaurene oxidase from Rhododendron Molle G. Don

  • 期刊论文

  • 发表日期：

    2025年12月9日

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  • Xinmeng Wang,
  • Zheng Liu,
  • Rongfeng Wang,
  • Hao Tang,
  • Ping Su,
  • Xianan Zhang,
  • Xiaoyi Wu,
  • Shurui Zhao,
  • Jiadian Wang,
  • Yating Hu,
  • Luqi Huang

摘要

Rhododendron molle G. Don contains a rich array of secondary metabolites, with grayanane diterpenoids as its representative components which exhibit diverse pharmacological activities, including anti-inflammatory, analgesic, antihypertensive, antiarrhythmic, antiviral and insecticidal properties. It has been reported that the 6/6/6/5 tetracyclic structure of ent-kaurene undergoes an A/B ring epoxide rearrangement, forming the 5/7/6/5 tetracyclic skeleton characteristic of grayanane diterpenoids. To identify CYP450s that may be involved in biosynthetic pathway, we successfully engineered a yeast chassis strain capable of producing 16α-hydroxy-ent-kaurane and ent-kaurene for functional validation. For the first time, we cloned three NADPH-cytochrome P450 reductases (RmCPRs) and identified RmCPR1 as the most active in the yeast system. Moreover, RmKO was cloned and confirmed which catalyzes the conversion of 16α-hydroxy-ent-kaurane into 16α-hydroxy-ent-kaurenoic acid. These findings provide valuable insights that may facilitate the discovery of more CYP450s from Rhododendron molle G. Don and lay the foundation for future elucidation of the biosynthetic pathway of grayanane diterpenoids.