Discovery of Demurilactone A: A Specific Growth Inhibitor of L-Form Bacillus subtilis

Abstract

Metabolic profiling of the extracts from a library of actinobacteria led to the identification of a novel polyketide, demurilactone A, produced by Streptomyces strain DEM21308. The structure of the compound was assigned based on a detailed investigation of 1D/2D NMR spectra and HR-MS. Whole genome DNA sequencing, followed by bioinformatics analysis and insertional mutagenesis, identified type I polyketide synthases encoded by the dml gene cluster to direct the biosynthesis of this polyene macrolide. While the number of modules is consistent with the carbon backbone of the assigned structure, some discrepancies were identified in the domain organization of five modules. Close investigation of the amino acid sequences identified several mutations in the conserved motifs of nonfunctional domains. Furthermore, the absolute configuration of hydroxy-bearing stereocenters was proposed based on analyses of the ketoreductase domains. Remarkably, although demurilactone A has little detectable activity against normal-walled bacteria, it specifically inhibits the growth of cell wall-deficient "L-form" Bacillus subtilis at a minimum inhibitory concentration value of 16 μg/mL. Time-lapse microscopy analyses revealed that demurilactone affects membrane dynamics, probably by reducing membrane fluidity. This compound could be a powerful reagent for studying longstanding questions about the involvement of L-forms in recurrent infection.

Repurposing drugs with specific activity against L-form bacteria

“…However, our main focus will be to evaluate whether the DHP or DMPM compounds identified as specific inhibitors of L-forms can be used to test the role of L-forms in models of recurrent infection, such as that recently described by Ganguli et al (2022) . While this work was in progress we serendipitously found that a novel polyene compound called demurilactone also differentially kills L-forms of B. subtilis ( Dashti et al, 2022 ).…”