Quality control by the ribosome following peptide bond formation

Abstract

The overall fidelity of protein synthesis has been thought to rely on the combined accuracy of two basic processes: the aminoacylation of transfer RNAs with their cognate amino acid by the aminoacyl-tRNA synthetases, and the selection of cognate aminoacyl-tRNAs by the ribosome in cooperation with the GTPase elongation factor EF-Tu. These two processes, which together ensure the specific acceptance of a correctly charged cognate tRNA into the aminoacyl (A) site, operate before peptide bond formation. Here we report the identification of an additional mechanism that contributes to high fidelity protein synthesis after peptidyl transfer, using a well-defined in vitro bacterial translation system. In this retrospective quality control step, the incorporation of an amino acid from a non-cognate tRNA into the growing polypeptide chain leads to a general loss of specificity in the A site of the ribosome, and thus to a propagation of errors that results in abortive termination of protein synthesis. In vitro translation studies have not been able to fully account for the reported fidelity of protein synthesis in the cell. The DNA and RNA polymerase complexes, which comprise the cell's other major macromolecular synthetic machineries, possess proofreading functions but until now no such activity has been found in the ribosome itself. Hani Zaher and Rachel Green now report the discovery of a previously uncharacterized system for ensuring the fidelity of protein synthesis that acts after peptide bond formation. If an incorrect amino acid is incorporated into a growing polypeptide chain, there is a general loss of specificity in the ribosome's active site, leading to a build-up of errors that triggers premature termination of peptide synthesis. This post-peptidyl transfer editing goes some way towards explaining the impressive in vivo accuracy achieved during protein synthesis. The DNA and RNA polymerases encode proofreading activities that help to ensure the fidelity of the readout. The ribosome, which synthesizes protein, was believed to lack a similar activity that would recognize errors after a peptide bond had been formed. This study provides evidence that the ribosome does have such a quality control mechanism. It is shown that after an incorrect amino acid has been incorporated into the growing polypeptide chain, there is a general loss of specificity in the ribosome's active site, leading to an accumulation of errors that triggers premature termination of peptide synthesis.