In Situ Biofilm Affinity-Based Protein Profiling Identifies the Streptococcal Hydrolase GbpB as the Target of a Carolacton-Inspired Chemical Probe.

Natural products are important precursors for antibiotic drug design. These chemical scaffolds serve as synthetic inspiration for chemists who leverage their structures to develop novel antibacterials and chemical probes. We have previously studied carolacton, a natural product macrolactone fromSorangium cellulosum, and discovered a simplified derivative, A2, that maintained apparent biofilm inhibitory activity, although the biological target was unknown. Herein, we utilize affinity-based protein profiling (AfBPP) in situ during biofilm formation to identify the protein target using a photoexcitable cross-linking derivative of A2. From these studies, we identified glucan binding protein B (GbpB), a peptidoglycan hydrolase, as the primary target of A2. Further characterization of the interaction between A2 and GbpB, as well as PcsB, a closely related homologue from the more pathogenic S. pneumoniae, revealed binding to the catalytic CHAP (cysteine, histidine, aminopeptidase) domain. To the best of our knowledge, this is the first report of a small-molecule binder of a conserved and essential bacterial CHAP hydrolase, revealing its potential as an antibiotic target. This work also highlights A2 as a useful tool compound for streptococci and as an initial scaffold for the design of more potent CHAP binders.

Quaternary Phosphonium Compounds: An Examination of Non-Nitrogenous Cationic Amphiphiles That Evade Disinfectant Resistance.

Quaternary ammonium compounds (QACs) serve as mainstays in the formulation of disinfectants and antiseptics. However, an over-reliance and misuse of our limited QAC arsenal has driven the development and spread of resistance to these compounds, as well as co-resistance to common antibiotics. Extensive use of these compounds throughout the COVID-19 pandemic thus raises concern for the accelerated proliferation of antimicrobial resistance and demands for next-generation antimicrobials with divergent architectures that may evade resistance. To this end, we endeavored to expand beyond canonical ammonium scaffolds and examine quaternary phosphonium compounds (QPCs). Accordingly, a synthetic and biological investigation into a library of novel QPCs unveiled biscationic QPCs to be effective antimicrobial scaffolds with improved broad-spectrum activities compared to commercial QACs. Notably, a subset of these compounds was found to be less effective against a known QAC-resistant strain of MRSA. Bioinformatic analysis revealed the unique presence of a family of small multiresistant transporter proteins, hypothesized to enable efflux-mediated resistance to QACs and QPCs. Further investigation of this resistance mechanism through efflux-pump inhibition and membrane depolarization assays illustrated the superior ability of P6P-10,10 to perturb the cell membrane and exert the observed broad-spectrum potency compared to its commercial counterparts. Collectively, this work highlights the promise of biscationic phosphonium compounds as next-generation disinfectant molecules with potent bioactivities, thereby laying the foundation for future studies into the synthesis and biological investigation of this nascent antimicrobial class.

Targeting S. mutans biofilms: a perspective on preventing dental caries.

The prevalence of biofilm diseases, and dental caries in particular, have encouraged extensive research on S. mutans biofilms, including methods of preventing its formation. Numerous small molecules with specific anti-biofilm activity against this pathogen have been isolated and synthesized. Generally, these molecules can be characterized into three categories: sucrose-dependent anti-adhesion, sucrose-independent anti-adhesion and cellular signaling interference. This review aims to provide an overview of the current small molecule strategies used for targeting S. mutans biofilms, and a perspective of the future for the field.

Synthetic Simplification of Carolacton Enables Chemical Genetic Studies in Streptococcus mutans.

Understanding the broader biological impact of carolacton, a macrolactone natural product, has been ongoing for the past decade. Multiple studies have shown connections to regulatory systems, acid tolerance mechanisms, biofilm formation, and recently folate dehydrogenase (FolD). Progress elucidating the cause of biofilm-specific activity in Streptococcus mutans has been limited due to low-throughput analyses of carolacton-treated cells. We disclose the discovery of a simplified carolacton-inspired analog that demonstrates inhibitory activity against S. mutans biofilm cells. This discovery permitted a proof of concept chemical genetic screen of S. mutans mutants identifying the carbon catabolite protein A signaling pathway as a putative target.