Combining Genomic and Metabolomic Tools for Novel Specialized Metabolites in the Genus Actinoallomurus

In response to the escalating threat of antimicrobial resistance, research at NAICONS Srl has focused on discovering bioactive natural products (NPs) by investigating the metabolic potential of Actinoallomurus, a relatively unexplored Actinobacteria genus. By integrating traditional cell-based bioassays with advanced metabolomic approaches, the analysis of extracts from 88 Actinoallomurus strains led to the identification of allopeptimicins, a novel family of cyclic depsipeptides with promising antimicrobial activity. Isolated from Actinoallomurus sp. ID145808, allopeptimicins possess a distinctive polyketide-peptide hybrid structure. Structural and bioactivity analyses demonstrated their selective activity against pathogenic bacteria, highlighting their potential therapeutic relevance. Characterization of their biosynthetic gene cluster (BGC) apt revealed gene cassettes for specialized precursors and modular assembly lines with similarities to other known BGCs. This enabled the deposition of apt in the newly released MIBiG 4.0 database, a resource supporting the NP research community in identifying BGCs and their metabolites. Furthermore, an innovative resin-based purification method using lithium chloride elution was developed for selective purification of allopeptimicins A, optimizing yield (35%) and purity (74%). To explore the metabolic potential of Actinoallomurus, a large-scale genomic analysis of 144 strains disclosed both conserved and unique BGCs with low or no similarity to known clusters, facilitating the prioritization of promising BGCs for novel metabolite discovery. Complementing the genomic study, a computational networking approach was implemented to aid large-scale data annotation. The Molecules Gateway, a proprietary database of 150k annotated molecules, enables efficient dereplication and compound identification in complex microbial extracts. This resource supported the development of a semi-automated workflow for molecular network annotation, laying a foundation for prioritizing novel NPs. Applied to 340 Actinoallomurus extracts, this workflow identified numerous genus-specific, uncharacterized molecular families, including one exclusive to strain M23/ID150205. Integration with genomic data enabled the identification of a novel BGC, exclusive to this strain, leading to the discovery of novel compounds currently under structural characterization. This scalable framework places Actinoallomurus as a promising source for novel metabolites and provides a model for exploring underexplored microbial genera to uncover new chemical scaffolds.