Tuning the Moenomycin Pharmacophore To Enable Discovery of Bacterial Cell Wall Synthesis Inhibitors

New antibiotic drugs need to be identified to address rapidly developing resistance of bacterial pathogens to common antibiotics. The natural antibiotic moenomycin A is the prototype for compounds that bind to bacterial peptidoglycan glycosyltransferases (PGTs) and inhibit cell wall biosynthesis, but it cannot be used as a drug. Here we report the chemoenzymatic synthesis of a fluorescently labeled, truncated analogue of moenomycin based on the minimal pharmacophore. This probe, which has optimized enzyme binding properties compared to moenomycin, was designed to identify low-micromolar inhibitors that bind to conserved features in PGT active sites. We demonstrate its use in displacement assays using PGTs from <i>S. aureus</i>, <i>E. faecalis</i>, and <i>E. coli</i>. 110,000 compounds were screened against <i>S. aureus</i> SgtB, and we identified a non-carbohydrate based compound that binds to all PGTs tested. We also show that the compound inhibits <i>in vitro</i> formation of peptidoglycan chains by several different PGTs. Thus, this assay enables the identification of small molecules that target PGT active sites, and may provide lead compounds for development of new antibiotics

Diversity-oriented synthesis encoded by deoxyoligonucleotides

Abstract Diversity-oriented synthesis (DOS) is a powerful strategy to prepare molecules with underrepresented features in commercial screening collections, resulting in the elucidation of novel biological mechanisms. In parallel to the development of DOS, DNA-encoded libraries (DELs) have emerged as an effective, efficient screening strategy to identify protein binders. Despite recent advancements in this field, most DEL syntheses are limited by the presence of sensitive DNA-based constructs. Here, we describe the design, synthesis, and validation experiments performed for a 3.7 million-member DEL, generated using diverse skeleton architectures with varying exit vectors and derived from DOS, to achieve structural diversity beyond what is possible by varying appendages alone. We also show screening results for three diverse protein targets. We will make this DEL available to the academic scientific community to increase access to novel structural features and accelerate early-phase drug discovery