Functional identification of key enzymes in calycosin biosynthesis in
- 1State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Cheeloo College of Medicine, Shandong University, Jinan, 250012, China
- 2Corresponding author.
Abstract
Astragalus membranaceus , a widely used medicinal and edible herb, contains major active isoflavonoid components such as formononetin, calycosin, and their glycosides, which exhibit a broad spectrum of pharmacological effects including anti-inflammatory, immunomodulatory, and cardioprotective activities. In this study, we identified three key enzymes: isoflavone synthase (AmIFS), O -methyltransferase (AmOMT2), and isoflavone 3′-hydroxylase (AmI3′H), elucidating the critical steps in calycosin biosynthesis and revealing a novel synthetic pathway. The research showed that AmIFS efficiently catalyzed the conversion of liquiritigenin and naringenin into the isoflavone backbone. The multifunctional O -methyltransferase AmOMT2 preferentially catalyzed the 4′-O-methylation of liquiritigenin, and the resulting product was subsequently converted by AmIFS into 2,7-dihydroxy-4′-methoxyisoflavanone, thereby broadening substrate versatility of AmIFS and revealing a new biosynthetic route. Meanwhile, AmI3′H completed the biosynthesis by catalyzing the final 3′-hydroxylation of formononetin to calycosin. Furthermore, this study successfully achieved the heterologous synthesis of calycosin precursors in yeast and tobacco, confirming the functional expression of these enzymes in plant and microbial hosts. Tissue-specific expression analysis showed differential expression of AmIFS and AmI3′H in roots, stems, and leaves of A. membranaceus , which was positively correlated with formononetin accumulation. These findings not only completed the calycosin biosynthetic pathway, but also laid a theoretical foundation and provided technical support for the scalable production of bioactive isoflavones via synthetic biology.
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BibTeX
@article{cui2026functional,
title = {{Functional identification of key enzymes in calycosin biosynthesis in}},
author = {Ming-Xin Cui and Yue Hao and Jiang-Nan Li and Dan-Dan Xu and Pei-Xi Hao and Jie Fu and Ying Lu and Ya-Tong Li and Lin-Lin Zhi and Jiao-Zhen Zhang and Hong-Xiang Lou and Ai-Xia Cheng},
journal = {Pharmaceutical Science Advances},
year = {2026},
month = nov,
doi = {10.1016/j.pscia.2026.100118},
url = {https://doi.org/10.1016/j.pscia.2026.100118},
note = {CC BY},
}
RIS
TY - JOUR
TI - Functional identification of key enzymes in calycosin biosynthesis in
AU - Ming-Xin Cui
AU - Yue Hao
AU - Jiang-Nan Li
AU - Dan-Dan Xu
AU - Pei-Xi Hao
AU - Jie Fu
AU - Ying Lu
AU - Ya-Tong Li
AU - Lin-Lin Zhi
AU - Jiao-Zhen Zhang
AU - Hong-Xiang Lou
AU - Ai-Xia Cheng
PY - 2026
DA - 2026/11/06
JO - Pharmaceutical Science Advances
DO - 10.1016/j.pscia.2026.100118
UR - https://doi.org/10.1016/j.pscia.2026.100118
AB - Astragalus membranaceus , a widely used medicinal and edible herb, contains major active isoflavonoid components such as formononetin, calycosin, and their glycosides, which exhibit a broad spectrum of pharmacological effects including anti-inflammatory, immunomodulatory, and cardioprotective activities. In this study, we identified three key enzymes: isoflavone synthase (AmIFS), O -methyltransferase (AmOMT2), and isoflavone 3′-hydroxylase (AmI3′H), elucidating the critical steps in calycosin biosynthesis and revealing a novel synthetic pathway. The research showed that AmIFS efficiently catalyzed the conversion of liquiritigenin and naringenin into the isoflavone backbone. The multifunctional O -methyltransferase AmOMT2 preferentially catalyzed the 4′-O-methylation of liquiritigenin, and the resulting product was subsequently converted by AmIFS into 2,7-dihydroxy-4′-methoxyisoflavanone, thereby broadening substrate versatility of AmIFS and revealing a new biosynthetic route. Meanwhile, AmI3′H completed the biosynthesis by catalyzing the final 3′-hydroxylation of formononetin to calycosin. Furthermore, this study successfully achieved the heterologous synthesis of calycosin precursors in yeast and tobacco, confirming the functional expression of these enzymes in plant and microbial hosts. Tissue-specific expression analysis showed differential expression of AmIFS and AmI3′H in roots, stems, and leaves of A. membranaceus , which was positively correlated with formononetin accumulation. These findings not only completed the calycosin biosynthetic pathway, but also laid a theoretical foundation and provided technical support for the scalable production of bioactive isoflavones via synthetic biology.
SN - 2773-2169
ER -