Structure of a hibernating 100S ribosome reveals an inactive conformation of the ribosomal protein S1.

To survive under conditions of stress, such as nutrient deprivation, bacterial 70S ribosomes dimerize to form hibernating 100S particles1. In γ-proteobacteria, such as Escherichia coli, 100S formation requires the ribosome modulation factor (RMF) and the hibernation promoting factor (HPF)2-4. Here we present single-particle cryo-electron microscopy structures of hibernating 70S and 100S particles isolated from stationary-phase E. coli cells at 3.0 Å and 7.9 Å resolution, respectively. The structures reveal the binding sites for HPF and RMF as well as the unexpected presence of deacylated E-site transfer RNA and ribosomal protein bS1. HPF interacts with the anticodon-stem-loop of the E-tRNA and occludes the binding site for the messenger RNA as well as A- and P-site tRNAs. RMF facilitates stabilization of a compact conformation of bS1, which together sequester the anti-Shine-Dalgarno sequence of the 16S ribosomal RNA (rRNA), thereby inhibiting translation initiation. At the dimerization interface, the C-terminus of uS2 probes the mRNA entrance channel of the symmetry-related particle, thus suggesting that dimerization inactivates ribosomes by blocking the binding of mRNA within the channel. The back-to-back E. coli 100S arrangement is distinct from 100S particles observed previously in Gram-positive bacteria5-8, and reveals a unique role for bS1 in translation regulation.