Raw data_Submission ID: 248578315

The rise of multidrug-resistant (MDR) strains has been seen as an increasing threat to global public health. Among them, carbapenemases-producing strains, mainly Gram-negative bacteria, are particularly concerning the secretion of β-lactamases that inactivate carbapenem antibiotics, often regarded as the last line of defense against MDR infections. Through a high-throughput screen of the U.S. Food and Drug Administration (FDA)-approved compound library, we identified tavaborole (AN2690), as an ultra-broad-spectrum β-lactamase inhibitor with activity against class A, B, and D β-lactamases. Tavaborole exerts its inhibitory effect by forming a transition-state tetrahedral intermediate with the serine residue of β-lactamases through its boron atom electrophilic center. This interaction blocks the deacylation step crucial for β-lactamase activity, effectively neutralizing the enzyme. Transcriptomics analysis revealed that tavaborole acts not only as a reversible covalent inhibitor of serine β-lactamases but also disrupts normal energy metabolism in <i>Klebsiella pneumoniae </i>(<i>K. pneumoniae</i>) via metabolic reprogramming. Specifically, tavaborole inhibits leucyl-tRNA synthetase activity in <i>K. pneumoniae</i>, simulating amino acid starvation and triggering a stringent response. This leads to a transient increase in guanosine tetraphosphate (ppGpp) levels, redirecting the bacterial energy metabolism from the tricarboxylic acid (TCA) cycle to glycolysis. <i>In vivo</i> studies demonstrated the efficacy of combining tavaborole with meropenem, which successfully rescued mice infected intraperitoneally with KPC-2-positive <i>K. pneumoniae</i>, providing significant protection. This dual-function benzoxaborole compound integrates carbapenemase inhibition with antimicrobial activity, offering a promising strategy to combat carbapenem-resistant <i>Enterobacteriaceae</i> (CRE) pathogens and safeguard the efficacy of our current antibiotic arsenal.