Bacterial populations contain persisters, cells which survive exposure to bactericidal antibiotics and other lethal factors. Persisters do not have a genetic resistance mechanism, and their means to tolerate killing remain unknown. In exponentially growing populations of Escherichia coli the frequency of persister formation usually is 10 ⁻⁷ to 10 ⁻⁵ . It has been shown that cells overexpressing either of the toxic proteins HipA and RelE, both members of the bacterial toxin-antitoxin (TA) modules, have the ability to form more persisters, suggesting a specific role for these toxins in the mechanism of persistence. However, here we show that cells expressing proteins that are unrelated to TA modules but which become toxic when ectopically expressed, chaperone DnaJ and protein PmrC of Salmonella enterica , also form 100- to 1,000-fold more persisters. Thus, persistence is linked not only to toxicity caused by expression of HipA or dedicated toxins but also to expression of other unrelated proteins.

Antimicrobial peptide Api137 inhibits translation by trapping release factors 1 or 2 associated with ribosomes and arresting termination. Many antibiotics stop bacterial growth by inhibiting different steps of protein synthesis. However, no specific inhibitors of translation termination are known. Proline-rich antimicrobial peptides, a component of the antibacterial defense system of multicellular organisms, interfere with bacterial growth by inhibiting translation. Here we show that Api137, a derivative of the insect-produced antimicrobial peptide apidaecin, arrests terminating ribosomes using a unique mechanism of action. Api137 binds to the Escherichia coli ribosome and traps release factor (RF) RF1 or RF2 subsequent to the release of the nascent polypeptide chain. A high-resolution cryo-EM structure of the ribosome complexed with RF1 and Api137 reveals the molecular interactions that lead to RF trapping. Api137-mediated depletion of the cellular pool of free release factors causes the majority of ribosomes to stall at stop codons before polypeptide release, thereby resulting in a global shutdown of translation termination.

Significance: Chloramphenicol and linezolid interfere with translation by targeting the ribosomal catalytic center and are viewed as universal inhibitors of peptide bond formation. We show that, contrary to this view, the activity of these antibiotics critically depends on the nature of specific amino acids of the nascent chain carried by the ribosome and by the identity of the residue entering the A site. These findings indicate that the nascent protein modulates the properties of the ribosomal catalytic center and affects binding of its ligands. Understanding the principles of context specificity of ribosomal drugs may help develop better antibiotics.