Deep Functional Profiling Facilitates the Evaluation of the Antibacterial Potential of the Antibiotic Amicoumacin

Abstract

The global spread of antibiotic resistance is forcing the scientific community to find new molecular strategies to counteract it. Deep functional profiling of microbiomes provides an alternative source for the discovery of novel antibiotic producers and probiotics. Recently, we implemented this ultrahigh-throughput screening approach for the isolation of Bacillus pumilus strains efficiently producing the ribosome-targeting antibiotic amicoumacin A (Ami). Proteomics and metabolomics revealed essential insight into the activation of Ami biosynthesis. Here, we applied omics to boost Ami biosynthesis, providing the optimized cultivation conditions for high-scale production of Ami. Ami displayed a pronounced activity against Lactobacillales and Staphylococcaceae, including methicillin-resistant Staphylococcus aureus (MRSA) strains, which was determined using both classical and massive single-cell microfluidic assays. However, the practical application of Ami is limited by its high cytotoxicity and particularly low stability. The former is associated with its self-lactonization, serving as an improvised intermediate state of Ami hydrolysis. This intramolecular reaction decreases Ami half-life at physiological conditions to less than 2 h, which is unprecedented for a terminal amide. While we speculate that the instability of Ami is essential for Bacillus ecology, we believe that its stable analogs represent attractive lead compounds both for antibiotic discovery and for anticancer drug development.

Mechanism of the Potential Therapeutic Candidate Bacillus subtilis BSXE-1601 Against Shrimp Pathogenic Vibrios and Multifunctional Metabolites Biosynthetic Capability of the Strain as Predicted by Genome Analysis

“…The strong anti-inflammatory and antiulcer properties of amicoumacin A were reported in the 1980s (Itoh et al, 1981;Iron et al, 1982), which was subsequently associated with its anticancer activity (Li et al, 2012;Prokhorova et al, 2016). Furthermore, amicoumacin A was documented to display a pronounced bactericidal activity against clinically relevant bacterial pathogens, such as Helicobacter pylori (Pinchuk et al, 2001) and the "superbug" methicillin-resistant Staphylococcus aureus (MRSA) (Berrue et al, 2009;Lama et al, 2012;Han et al, 2013;Terekhov et al, 2020). Noteworthily, besides the clinical applications, amicoumacin A produced by B. pumilus H2 also exhibited antagonistic effect toward the fish pathogen V. vulnificus CZ-A2 in vitro (Gao et al, 2017).…”

Liquid drop of DNA libraries reveals total genome information

“…The most refined method to avoid mispairing is based on amplification of single DNA molecules encapsulated inside isolated microcompartments of water-in-oil emulsions. Compartmentalization provides critical advantages to droplet-based single-cell technologies, namely the ability to distinguish and select rare individual clones from the enormous biodiversity ( 7 – 10 ). Similarly, pioneering work on single-molecule PCR amplification opened the era of emulsion PCR (ePCR) ( 11 ), which was successfully applied to amplifying complex gene libraries ( 12 ), PCR on microparticles in water-in-oil emulsions ( 13 ), and directed evolution driven by in vitro compartmentalization ( 14 ).…”

Genome Mining, Heterologous Expression, Antibacterial and Antioxidant Activities of Lipoamides and Amicoumacins from Compost-Associated Bacillus subtilis fmb60

“…In the present work, five novel lipoamides and amicoumacins were identified from B. subtilis fmb60 by genome-directed isolation. Amicoumacins belong to a family of 3, 4-dihydroisocoumarin derivatives produced by the genus Bacillus , which have shown antibacterial, anti-inflammatory, and antiulcer activities and potent gastroprotective and antiulcerogenic activities [ 30 , 31 ]. To identify the genetic determinants of lipoamide and amicoumacins biosynthesis in B. subtilis fmb60, the NRPS/PKS gene cluster of B. subtilis fmb60 genome was submitted to BLAST.…”