文献类型： 外文期刊

作者： Li, Yihan ¹ ; Kang, Chuanzhi ² ; Xu, Jiahui ¹ ; Zhou, Wenqing ³ ; Pan, Weishan ¹ ; Xia, Daofang ³ ; Liang, Jian ¹ ; Guo, Lanping ² ; Ma, Xiao-kui ¹ ;

作者机构： 1.Shaanxi Normal Univ, Coll Life Sci, Key Lab Med Resources & Nat Pharmaceut Chem, Natl Engn Lab Resource Developing Endangered Chine, Xian 710055, Shaanxi, Peoples R China

2.China Acad Chinese Med Sci, Natl Resource Ctr Chinese Mat Med, State Key Lab Qual Ensurance & Sustainable Use Dao, Beijing 100700, Peoples R China

3.Ningxia Acad Agr & Forestry Sci, Inst Plant Protect, Yinchuan 750002, Peoples R China

4.Ningxia Forest Rights Serv & Forestry Ind Dev Ctr, Yinchuan 750001, Peoples R China

关键词： Sanghuangporus sanghuang; Sesquiterpenoids; Terpenoid synthesis (TPS); Sesquiterpene synthase; (-)-Aristolone; Fermentation kinetics

期刊名称：APPLIED MICROBIOLOGY AND BIOTECHNOLOGY （ 影响因子：4.3； 五年影响因子：5.1 ）

ISSN： 0175-7598

年卷期： 2025 年 109 卷 1 期

页码：

收录情况： SCI

摘要： (-)-Aristolone, a sesquiterpene with promising therapeutic properties such as antidiabetic and vasorelaxant effects, currently suffers from limited availability due to inefficient chemical synthesis and lack of viable extraction methods. This study presents a novel strategy for high-yield microbial (-)-aristolone production using Sanghuangporus sanghuang DM989 as a fungal chassis. Genome mining identified nine sesquiterpene synthases, among which TPS2152 was functionally linked to (-)-aristolone biosynthesis. TPS2152 harbors a rare DQxxD motif, diverging from the canonical DDxxD motif in plants, suggesting unique catalytic properties in fungi. Overexpression of farnesyl pyrophosphate synthase (FPPS) increased FPP precursor supply, resulting in a 78.79% rise in squalene content (1.18 mg/g) and enabling de novo (-)-aristolone synthesis (0.42 mg/g) in the FPPS+ strain. To enhance FPP flux toward (-)-aristolone, the Delta SQS/TPS2152+ strain was constructed by co-overexpressing TPS2152 and silencing squalene synthase (SQS), yielding a 210% increase in (-)-aristolone (1.30 mg/g) and 56.78% reduction in squalene compared to FPPS+. Further, site-directed mutagenesis converted DQxxD to DDxxD, producing TPS2152D, which retained substrate binding affinity (docking score: - 9.1 kcal/mol) and exhibited a 2.57-fold increase in catalytic efficiency. Integration of TPS2152D with SQS silencing produced the Delta SQS/TPS2152D+ strain, achieving a 217% higher (-)-aristolone yield than FPPS+. Fermentation kinetics showed product accumulation from day 5, with maximal Qp on days 8 and complete squalene suppression by day 9. These results establish S. sanghuang as a robust microbial platform for sesquiterpene production and demonstrate the feasibility of combining fungal pathway engineering and motif-based enzyme optimization for scalable biosynthesis of high-value terpenoids.