Discovery of actinomycin L, a new member of the actinomycin family of antibiotics

Nataliia V Machushynets,Gilles P Van Wezel,Mercedes De La Cruz,Thomas Hankemeier,Olga Genilloud,Chao Du,Maxime A Siegler,Somayah S Elsayed

doi:10.1038/s41598-022-06736-0

Abstract

Streptomycetes are major producers of bioactive natural products, including the majority of the naturally produced antibiotics. While much of the low-hanging fruit has been discovered, it is predicted that less than 5% of the chemical space of natural products has been mined. Here, we describe the discovery of the novel actinomycins L1 and L2 produced by Streptomyces sp. MBT27, via application of metabolic analysis and molecular networking. Actinomycins L1 and L2 are diastereomers, and the structure of actinomycin L2 was resolved using NMR and single crystal X-ray crystallography. Actinomycin L is formed via spirolinkage of anthranilamide to the 4-oxoproline moiety of actinomycin X2, prior to the condensation of the actinomycin halves. Such a structural feature has not previously been identified in naturally occurring actinomycins. Adding anthranilamide to cultures of the actinomycin X2 producer Streptomyces antibioticus, which has the same biosynthetic gene cluster as Streptomyces sp. MBT27, resulted in the production of actinomycin L. This supports a biosynthetic pathway whereby actinomycin L is produced from two distinct metabolic routes, namely those for actinomycin X2 and for anthranilamide. Actinomycins L1 and L2 showed significant antimicrobial activity against Gram-positive bacteria. Our work shows how new molecules can still be identified even in the oldest of natural product families.

Highlights

Streptomycetes are major producers of bioactive natural products, including the majority of the naturally produced antibiotics
The pentapeptide precursors are biosynthesized by a non-ribosomal peptide synthetase (NRPS) assembly line, and actinomycins are formed through oxidative condensation of two 3-hydroxy-4-methylanthranilic acid (4-MHA) pentapeptide lactones (PPLs)[27]
PPL is biosynthesized by an NRPS assembly line with the 4-MHA as the initiating unit38,39. 4-MHA is derived from 3-hydroxy-4-methylkynurenine (4-MHK), which is formed by methylation of 3-hydroxykynurenine (3-HK)[26]

Summary

Introduction

Streptomycetes are major producers of bioactive natural products, including the majority of the naturally produced antibiotics. Triggering the expression of silent BGCs by genetic and cultivationbased techniques should facilitate unlocking this yet unexplored chemical diversity, allowing the discovery of novel molecules[8,9] This strategy relies on altering the regulatory networks of the producing organism in response to fluctuating culturing conditions, such as carbon, nitrogen or phosphate concentration[10,11,12]. Metabolic profiling of crude extracts obtained under different growth conditions represents a challenging analytical task since these mixtures are composed of hundreds of natural products Metabolomics, those based on mass spectrometry (MS), became more and more valuable and greatly increased the efficiency of such screenings[19]. The pentapeptide precursors are biosynthesized by a non-ribosomal peptide synthetase (NRPS) assembly line, and actinomycins are formed through oxidative condensation of two 3-hydroxy-4-methylanthranilic acid (4-MHA) pentapeptide lactones (PPLs)[27]

Methods

Results

Conclusion