A Novel Peptide Antibiotic Targeting Gram-Negative Infections Designed from <i>Mycobacterium tuberculosis</i> Adenylate Kinase

We explored the lipopolysaccharide-binding properties of adenylate kinase from Mycobacterium tuberculosis (MtAdk) to facilitate the design of novel peptide antibiotics. Notably, we de novo designed 11-mer peptides derived from the AMP-binding domain (Lys44 to Asp54) of MtAdk. Among 71 designed peptides, DD-S067 was the most effective, especially against carbapenem-resistant Acinetobacter baumannii (CRAB), with minimal development of drug resistance. DD-S067 exhibited multiple antibacterial mechanisms, including disrupting both the outer and inner bacterial membranes, and inducing reactive oxygen species that trigger lipid peroxidation. Transcriptome analysis revealed that DD-S067 disrupted key cellular pathways in CRAB by inhibiting the electron transport chain and triggering oxidative stress responses, ultimately suppressing CRAB virulence mechanisms. Furthermore, DD-S067 exhibited significant protective effects in a CRAB-induced septic shock mouse model, highlighting its potential as a novel peptide antibiotic for treating Gram-negative infections. These findings pave the way for innovative strategies in developing protein-based antibiotics.