Rationally Designed Novel Phenyloxazoline Synthase Inhibitors: Chemical Synthesis and Biological Evaluation to Accelerate the Discovery of New Antimycobacterial Antibiotics.

The uncontrolled spread of drug-resistant tuberculosis (DR-TB) clinical cases necessitates the urgent discovery of newer chemotypes with novel mechanisms of action. Here, we report the chemical synthesis of rationally designed novel transition-state analogues (TSAs) by targeting the cyclization (Cy) domain of phenyloxazoline synthase (MbtB), a key enzyme of the conditionally essential siderophore biosynthesis pathway. Following bio-assay-guided evaluation of TSA analogues preferentially in iron-deprived and iron-rich media to understand target preferentiality against a panel of pathogenic and non-pathogenic mycobacteria strains, we identified a hit, i.e., . Molecular docking, dynamics, and MMPBSA calculations enabled us to comprehend 's stable binding at the active site pocket of and the results imply that the binding pocket has a strong affinity for electron-withdrawing functional groups and contributes to stable polar interactions between enzyme and ligand. Furthermore, enhanced intracellular killing efficacy (8 μg/mL) of against in infected macrophages is noted in comparison to moderate in vitro antimycobacterial efficacy (64 μg/mL) against . also demonstrates whole-cell efflux pump inhibitory activity against . Identification of by focusing on the modular domain paves the way for accelerating novel anti-TB antibiotic discoveries.