Switching cell-penetrating peptides from "deliverer" to "enforcer" by hydrophobic amino acid end-tagging to combat intracellular bacterial infections.

The presence of recalcitrant intracellular bacteria (ICB) and antimicrobial resistance poses a significant challenge to global public health. Antimicrobial peptides (AMPs) and cell-penetrating peptides (CPPs) play irreplaceable roles in combating multidrug-resistant (MDR) bacteria and drug delivery, respectively, but how to combine the advantages of both to design peptide-based biomaterials to fight ICB is still a formidable challenge. Here we constructed a miniature peptide library containing thirty AMPs with dual functions of antimicrobial and cell-penetrating properties based on CPP transactivator of transcription (Tat) by combining the end-tagging and structure-activity relationship of AMPs. The lead peptide, 5VT, identified by screening of a peptide library, exhibited potent activity, including against clinically isolated drug-resistant strains, while also demonstrating excellent biocompatibility. Strikingly, 5VT efficiently penetrates cells primarily through clathrin-mediated endocytosis and macropinocytosis, and is capable of eradicating intracellular Salmonella typhimurium and alleviating inflammatory responses. Additionally, 5VT kills bacteria primarily through employing a dual mechanism that involves attacking bacterial cell membranes and triggering abnormal production of reactive oxygen species. Further in vivo analysis showed that 5VT was non-systemically toxic and exhibited substantial therapeutic capacity in a peritonitis-sepsis model. We provide a simple and efficient strategy for transforming cationic CPPs from "deliverers" to "all-purpose warriors" to fight ICB, which provides new insights into the design and development of peptide-based antimicrobial materials to deal with the increasing prevalence of intracellular bacterial and MDR bacteria.