Mode of Action of the Monobactam LYS228 and Mechanisms Decreasing In Vitro Susceptibility in Escherichia coli and Klebsiella pneumoniae

The monobactam scaffold is attractive for the development of new agents to treat infections caused by drug-resistant Gram-negative bacteria because it is stable to metallo-β-lactamases (MBLs). However, the clinically used monobactam aztreonam lacks stability to serine β-lactamases (SBLs) that are often coexpressed with MBLs. LYS228 is stable to MBLs and most SBLs. LYS228 bound purified penicillin binding protein 3 (PBP3) similarly to aztreonam (derived acylation rate/equilibrium dissociation constant [ / ] of 367,504 s M and 409,229 s M , respectively) according to stopped-flow fluorimetry. A gel-based assay showed that LYS228 bound mainly to PBP3, with weaker binding to PBP1a and PBP1b. Exposing cells to LYS228 caused filamentation consistent with impaired cell division. No single-step mutants were selected from 12 strains expressing different classes of β-lactamases at 8× the MIC of LYS228 (frequency,