Abstract
Fungal terpenoids catalyzed by bifunctional terpene synthases (BFTSs) possess interesting bioactive and chemical properties. In this study, an integrated approach of genome mining, heterologous expression, and in vitro enzymatic activity assay was used, and these identified a unique BFTS sub-clade critical to the formation of a 5-15 trans-fused bicyclic sesterterpene preterpestacin I (1). The 5-15 bicyclic BFTS gene clusters were highly conserved but showed relatively wide phylogenetic distribution across several species of the diverged fungal classes Dothideomycetes and Sordariomycetes. Further genomic organization analysis of these homologous biosynthetic gene clusters from this clade revealed a glycosyltransferase from the graminaceous pathogen Bipolaris sorokiniana isolate BS11134, which was absent in other 5-15 bicyclic BFTS gene clusters. Targeted isolation guided by BFTS gene deletion led to the identification of two new sesterterpenoids (4, and 6) from BS11134. Compounds 2 and 4 showed moderate effects on LPS-induced nitrous oxide production in the murine macrophage-like cell line RAW264.7 with in vitro inhibition rates of 36.6 ± 2.4% and 24.9 ± 2.1% at 10 μM, respectively. The plausible biosynthetic pathway of these identified compounds was proposed as well. This work revealed that phytopathogenic fungi can serve as important sources of active terpenoids via systematic analysis of the genomic organization of BFTS biosynthetic gene clusters, their phylogenetic distribution in fungi, and cyclization properties of their metabolic products.
Key points
• Genome mining of the first BFTS BGC harboring a glycosyltransferase.
• Gene-deletion guided isolation revealed three novel 5-15 bicyclic sesterterpenoids.
• Biosynthetic pathway of isolated sesterterpenoids was proposed.
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All data generated or analyzed during this study are included in the published article and electrical supplementary material available at https://doi.org/XXXX.
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Acknowledgements
We gratefully acknowledge the financial support from the National Key Research and Development Program of China (2019YFA0906200, 2020YFA090032, and 2020YFA0907200), the National Natural Science Foundation of China (21877038, 21907031, 81903529, 21977029, 31720103901, and 81573341), the Open Project Funding of the State Key Laboratory of Bioreactor Engineering, the 111 Project (B18022), Shanghai Rising-Star Program (20QA1402800), Shanghai Science and Technology Commission (18JC1411900). Genome sequencing and assembly of in-house genomes were supported by funding from the Natural Science and Engineering Research Council of Canada to T. Hsiang.
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XL and LZ designed this project. XL, JZ, LJ, and GZ arranged the research consortium, TH sequenced and assembled genomic data. CH, ZC, XW, and XC conducted the bioinformatic analyses and analyzed the data. JZ, GZ, JH, WY, KL, XW, HY, XL, XX, and ML carried out fermentation, compound purification, and structural elucidation. LJ, CH, XZ, ZW, GT, LZ, LK, GL, and SG contributed to molecular biological experiments, protein purification, and enzymatic reaction. BR, LM, RQ, and HD contributed to biological assays. LJ, GZ, JZ, XL, and LZ wrote the manuscript, and TH revised it. All authors discussed the results and approved the manuscript.
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Jiang, L., Zhu, G., Han, J. et al. Genome-guided investigation of anti-inflammatory sesterterpenoids with 5-15 trans-fused ring system from phytopathogenic fungi. Appl Microbiol Biotechnol 105, 5407–5417 (2021). https://doi.org/10.1007/s00253-021-11192-3
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DOI: https://doi.org/10.1007/s00253-021-11192-3