Bacterial ribosome conformational dynamics during translation termination and rescue
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Hydrolysis of polypeptide from the ribosome is a critical step that must occur prior to the ribosome recycling phase of translation. Inability of cells to do so can result in translation inhibition and eventually leading to cell death. In bacteria, class one release factors bind to the ribosome to catalyze the release of the mature polypeptide during translation termination. However, in the event of ribosome stalling as a result of mRNA truncation, ribosome rescue factors bind to the ribosome to catalyze the release of the growing polypeptide from the stalled complex. This rescue process is then followed by ribosome recycling. Here we employ smFRET to study the effects of the class I release factors, RF1 and RF2, and an alternative release factor known as YaeJ on the conformational dynamics of the ribosome following hydrolysis of peptidyl tRNAs. Further, we investigated the role of A-site mRNA on the global conformation of the ribosome. Our results demonstrate that upon binding to their cognate stop codon, the class I release factors stabilize ribosome complexes in the non-rotated state. Similarly, binding of YaeJ to complexes that are assembled on truncated mRNAs resulted in ribosomes that occupy primarily the non-rotated state. We also observe that absence of mRNA in the A-site induces a hyper-rotated conformation between the two subunits. Together, these findings further characterize the interactions between these different ligands and the bacterial ribosome. In addition, these results suggest that stabilization of the ribosome in the non-rotated state is critical for priming the ribosome for the recycling phase of translation.
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