Identification of a NRP with the ability to destroy cell membrane of Staphylococcus epidermidis from Rhodococcus genomes database.

With the rising infections caused by Staphylococcus epidermidis in clinical and the threat of existing antibiotic resistance, it is urgent to identify novel antimicrobial peptides (AMPs) against S. epidermidis. However, most microorganisms are difficult to culture, and it is hard to discover novel AMPs through conventional bioassay-guided fractionation. In this study, thousands of non-ribosomal peptide synthetase (NRPS) biosynthetic gene clusters (BGCs) were mined from hundreds of Rhodococcus genomes in public databases. Bioinformatic tools were employed to predict the core molecular skeletons of non-ribosomal peptides (NRPs) synthesized by selected NRPSs. A NRP analogue A8-5-line, which was obtained by chemical synthesis, showed potent activity against S. epidermidis. Based on structure-activity relationship (SAR) studies, substitution of the C-terminal hydrophilic glutamine residue with a hydrophobic alanine residue generated the derivative A8NO5, which exhibited significantly enhanced antibacterial activity. Mechanistic studies showed that A8NO5 can destroy bacterial cell membrane integrity effectively. Metabolomic analysis revealed that nucleotide metabolism pathway was disrupted upon the treatment of A8NO5. The altered nucleotide-related metabolites are likely downstream effects of membrane damage and the associated metabolic collapse. Furthermore, A8NO5 displayed negligible cytotoxicity and hemolytic activity in vitro, indicating a favorable biosafety. Collectively, these findings position A8NO5 as a promising lead compound for the development of novel bactericidal agents.