Media and strain studies for the scaled production of cis-enone resorcylic acid lactones as feedstocks for semisynthesis

Zeinab Y Al Subeh,Nicholas H Oberlies,Huzefa A Raja,Cedric J Pearce,Jennifer C Obike,Mitchell P Croatt

doi:10.1038/s41429-021-00432-3

Abstract

Resorcylic acid lactones (RALs) with a cis-enone moiety, represented by hypothemycin (1) and (5Z)-7-oxozeaenol (2), are fungal secondary metabolites with irreversible inhibitory activity against protein kinases, with particularly selective activity for inhibition of TAK1 (transforming growth factor beta-activated kinase 1). Gram-scale quantities of these compounds were needed as feedstock for semi-synthesizing RAL-analogues in a step-economical fashion. To do so, this study had three primary goals: identifying fungi that biosynthesized 1 and 2, enhancing their production by optimizing the fermentation conditions on the lab scale, and developing straight forward purification processes. After evaluating 536 fungal extracts via an in-house dereplication protocol, three strains were identified as producing cis-enone RALs (i.e., MSX78495, MSX63935, MSX45109). Screening these fungal strains on three grain-based media revealed enhanced production of 1 by strain MSX78495 on oatmeal medium, while rice medium increased the biosynthesis of 2 by strain MSX63935. Furthermore, the purification processes were improved, moving away from HPLC purification to utilizing two to four cycles of resuspension and centrifugation in small volumes of organic solvents, generating gram-scale quantities of these metabolites readily. In addition, studying the chemistry profiles of strains MSX78495 and MSX63935 resulted in the isolation of ten other RALs (3-12), two radicinin analogues (13-14), and six benzopyranones (15-20), with 19 and 20 being newly described chlorinated benzopyranones.

Highlights

The fungal kingdom, which is estimated to include between 2.2 and 5.1 million species represents a large reservoir for a variety of bioactive compounds [1, 2]
Seven fungal strains were identified as biosynthesizing Resorcylic acid lactones (RALs), and three of those produced cisenone RALs (i.e., hypothemycin (1) and (5Z)-7-oxozeaenol (2)) (Fig. 1 and S1)
The One-Strain-Many Compounds” approach (OSMAC) approach was used to enhance the biosynthesis of cis-enone RALs from three different fungal strains

Summary

Introduction

The fungal kingdom, which is estimated to include between 2.2 and 5.1 million species represents a large reservoir for a variety of bioactive compounds [1, 2]. It is well known that growth conditions and fermentation media have a significant impact on the fungal biosynthetic machinery [3]. Both the yield and the composition of fungal secondary metabolites are affected by environmental factors [4,5,6,7,8,9,10].

Methods

Results

Conclusion