Structural Superimposition

Abstact

During bacterial ribosome recycling, 70S ribosomes are split into subunits by ribosome recycling factor (RRF) and elongation factor G (EF-G). The antibiotic fusidic acid (FA) inhibits elongation and ribosome recycling by locking EF-G to the ribosome. Yet, no functional ribosome recycling FA complex has been successfully captured. Here, we used single-particle cryo-electron microscopy to resolve multiple FA-stalled intermediates of Staphylococcus aureus ribosomes, including a 70S intermediate with RRF and EF-G in a previously unobserved conformation. Our structures reveal how RRF and EF-G jointly disrupt inter-subunit bridges, promote back-rotation of the small subunit, and move the transfer RNA toward the E site to facilitate ribosome splitting. We further show that FA predominantly inhibits recycling by trapping EF-G on the post-termination complex in the absence of RRF, preventing formation of the active RRF*EF-G complex. These insights advance understanding of the molecular mechanism of bacterial ribosome recycling and the mode of action of FA as an antibiotic.