tmRNA-induced Release of Messenger RNA from Stalled Ribosomes

A ribosome stalled on a truncated mRNA in the eubacterial cell can be rescued by tmRNA via a process called trans-translation. We demonstrate here that release of truncated mRNAs from stalled ribosomes accelerates significantly already after trans-peptidation following tmRNA binding to the ribosome. However, rapid release of truncated mRNA requires EF-G-dependent translocation of peptidyl-tmRNA from the A to the P site of the ribosome. We show also that the rate of mRNA release before and after peptidyl-tmRNA translocation correlates well with the rate of dissociation of deacylated tRNA, indicating that mRNA is retained on the ribosome mainly through codon:anticodon interaction with tRNA. The rate of mRNA release is reduced for mRNAs with strong Shine–Dalgarno (SD)-like sequences in the vicinity of the truncation site as well as for mRNAs with long 3′ extensions downstream from the P-site codon. The reduced rate of release in the former case was due to a persisting SD–anti SD interaction between mRNA and the ribosome.