Structural insights into context-dependent inhibitory mechanisms of chloramphenicol in cells

Abstract

Ribosome-targeting antibiotics represent an important class of antimicrobial drugs. Chloramphenicol (Cm) is a well-studied peptidyl transfer center (PTC) binder, and growing evidence suggests it inhibits translation in a nascent peptide sequence-dependent manner. How such inhibition on the molecular scale translates to action on the cellular level remains unclear. Here, we employ cryo-electron tomography to visualize the impact of Cm inside the bacterium Mycoplasma pneumoniae. By resolving cellular Cm-bound ribosomes to 3.0 [A], we provide atomic detail on Cms coordination and interaction with natural nascent peptides and tRNAs in the PTC. We find that Cm leads to accumulation of translation elongation states that indicate ongoing futile accommodation cycles, and to extensive ribosome collisions. We thus suggest that beyond its inhibition of protein synthesis, the action of Cm may involve activation of cellular stress responses. This work exemplifies how in-cell structural biology advances understanding of mechanisms of action for extensively-studied antibiotics.