Bis-Catechol Siderophores Modulate β-Lactam Acylation at the Active Site of Pseudomonas aeruginosa Penicillin-Binding Protein 3

Siderophore−β-lactam conjugates represent a promising strategy to enhance antibiotic efficacy, yet their molecular basis remains incompletely understood. While improved permeability through TonB-dependent transporters is invoked to explain their activity, our results highlight increased penicillin-binding protein (PBP) affinity. Using Pseudomonas aeruginosa penicillin-binding protein 3 as a model, we compared the binding efficiency constant k2/Ks of unconjugated ampicillin, loracarbef, and their siderophore-based conjugate. Siderophore conjugation induced a modest (∼2-fold) increase in potency for ampicillin, but a much larger (∼50-fold) enhancement for loracarbef. Crystallographic analyses revealed a conserved β-lactam binding mode, while showing that siderophore extension at R1 modifies active-site interactions, notably recruiting residues such as Arg489 and modifying H-bonding interactions. Furthermore, in silico modeling suggests possible interactions between the siderophore and the β3−β4 loop residues. These findings support siderophores as both delivery vectors and structural elements able to increase β-lactam inhibitory potency against PBP.