Peptidoglycan-Mimetic Inhibitors of D,D- and L,D-Transpeptidases: Mechanistic Design Principles and Translational Barriers in the Post-β-Lactam Era

Abstract

Background: Peptidoglycan cross-linking is essential for bacterial cell-wall integrity and is catalyzed by D,D-transpeptidases (penicillin-binding proteins, PBPs) and L,D-transpeptidases (Ldts). Escalating β-lactam resistance driven by β-lactamases, altered PBPs, and compensatory reliance on Ldt-mediated 3→3 cross-links has intensified interest in non-β-lactam strategies that directly inhibit transpeptidases. Objective: This narrative review critically synthesizes mechanistic, structural, and medicinal chemistry evidence supporting peptidoglycan-mimetic inhibition of PBPs and Ldts, with emphasis on covalent warhead design, structural validation, and translational barriers. Methods: A structured narrative literature search of PubMed/MEDLINE, Scopus, and Web of Science was conducted for January 2010–March 2025, supplemented by reference list screening. Eligible articles reported transpeptidase inhibitor design with biochemical potency metrics, structural evidence of binding, and/or microbiological activity. Results: Substrate-mimetic and conformationally constrained scaffolds can reproduce key recognition motifs, including m-DAP pocket interactions, while boronic acids/cyclic boronates enable reversible covalent inhibition of serine-dependent PBPs via tetrahedral transition-state mimicry, and electrophilic warheads such as nitriles/cyanamides can covalently modify catalytic cysteine residues in Ldts. Structural studies validate active-site engagement across representative PBPs and Ldts, but whole-cell translation is frequently limited by Gram-negative outer membrane permeability, efflux susceptibility, and potential off-target reactivity of electrophiles. Conclusion: Peptidoglycan-mimetic transpeptidase inhibition is mechanistically compelling, yet clinically meaningful antibacterial development will require integrated optimization of target engagement, selectivity, and bacterial accumulation alongside pharmacokinetic feasibility