Short Lecture “High-throughput whole-cell biotransformation approach for fast and efficient chemodiversification of natural products”

Biotransformation is known to be a green, affordable and efficient way to generate chemodiversity. Previous works in our lab have shown that the use of secreted enzymes produced by fungal species (“fungal secretome”) was highly effective to generate complex molecules from simple starting materials [1], [2], [3], [4]. Despite these successes, the fungal secretome approach suffers from some drawbacks. For example, non-secreted enzymes are not recovered and enzymes requiring a cofactor lose their activity. Conventional whole-cell biotransformation can address these issues, but is time- and substrate-consuming, and therefore not suitable for screening approaches. The present work describes the use of fungal cultures in 96-well plates to perform high-throughput whole-cell biotransformations. This method allows the screening of the biotransformation capacity of a large number of fungal strains with small amounts of substrates. The approach benefits from recent advances in rapid and efficient annotation workflows using UHPLC-HRMS/MS that allow identification of the generated compounds and provide insight into their possible structure. A proof of concept of this screening method was performed with 30 strains of the necrotrophic fungal pathogen Botrytis sp. (B. cinerea and other close species) to identify those capable of hydroxylation reactions on various substrates. The most promising reactions were performed on a large scale, allowing the isolation of a series of hydroxylated terpene derivatives in a targeted manner by high-resolution chromatography. These preliminary results show that the proposed method allows a fast and efficient screening of biotransformation reactions using living fungi in order to generate libraries of compounds for biological screening.

Publikationsverlauf

Artikel online veröffentlicht:
12. Dezember 2022

• References

• 1 Queiroz EF, Gindro K. Improve the chemical biodiversity of natural products for drug discovery by fungal secretome-assisted biotransformation. The Swiss National Science Foundation provided financial support for this project. Grant 205321_182438/1.
• 2 Righi D, Huber R, Koval A. et al. Generation of stilbene antimicrobials against multiresistant strains of Staphylococcus aureus through biotransformation by the enzymatic secretome of Botrytis cinerea. J Nat Prod 2020; 83: 2347-2356
• 3 Huber R, Marcourt L, Koval A. et al. Chemoenzymatic synthesis of complex phenylpropanoid derivatives by the Botrytis cinerea secretome and evaluation of their Wnt inhibition activity. Front Plant Sci 2022; 12: 805610
• 4 Huber R, Koval A, Marcourt L. et al. Chemoenzymatic synthesis of original stilbene dimers possessing Wnt inhibition activity in triple-negative breast cancer cells using the enzymatic secretome of Botrytis cinerea Pers. Front Chem 2022; 10: 881298