The l,d-Transpeptidase Ldt_Ab from Acinetobacter baumannii Is Poorly Inhibited by Carbapenems and Has a Unique...

l,d-Transpeptidases (LDTs) are enzymes that catalyze reactions essential for biogenesis of the bacterial cell wall, including formation of 3-3 cross-linked peptidoglycan. Unlike the historically well-known bacterial transpeptidases, the penicillin-binding proteins (PBPs), LDTs are resistant to inhibition by the majority of β-lactam antibiotics, with the exception of carbapenems and penems, allowing bacteria to survive in the presence of these drugs. Here we report characterization of Ldt_Ab from the clinically important pathogen, Acinetobacter baumannii. We show that A. baumannii survives inactivation of Ldt_Ab alone or in combination with PBP1b or PBP2, while simultaneous inactivation of Ldt_Ab and PBP1a is lethal. Minimal inhibitory concentrations (MICs) of all 13 β-lactam antibiotics tested decreased 2- to 8-fold for the Ldt_Ab deletion mutant, while further decreases were seen for both double mutants, with the largest, synergistic effect observed for the Ldt_Ab + PBP2 deletion mutant. Mass spectrometry experiments showed that Ldt_Ab forms complexes in vitro only with carbapenems. However, the acylation rate of these antibiotics is very slow, with the reaction taking longer than four hours to complete. Our X-ray crystallographic studies revealed that Ldt_Ab has a unique structural architecture and is the only known LDT to have two different peptidoglycan-binding domains.