Eukaryotic Release Factor 3 Is Required for Multiple Turnovers of Peptide Release Catalysis by Eukaryotic Release Factor 1

Daniel E Eyler,Karen A Wehner,Rachel Green

doi:10.1074/jbc.m113.487090

Daniel E Eyler, Karen A Wehner + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m113.487090

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Abstract

Eukaryotic peptide release factor 3 (eRF3) is a conserved, essential gene in eukaryotes implicated in translation termination. We have systematically measured the contribution of eRF3 to the rates of peptide release with both saturating and limiting levels of eukaryotic release factor 1 (eRF1). Although eRF3 modestly stimulates the absolute rate of peptide release (∼5-fold), it strongly increases the rate of peptide release when eRF1 is limiting (>20-fold). This effect was generalizable across all stop codons and in a variety of contexts. Further investigation revealed that eRF1 remains associated with ribosomal complexes after peptide release and subunit dissociation and that eRF3 promotes the dissociation of eRF1 from these post-termination complexes. These data are consistent with models where eRF3 principally affects binding interactions between eRF1 and the ribosome, either prior to or subsequent to peptide release. A role for eRF3 as an escort for eRF1 into its fully accommodated state is easily reconciled with its close sequence similarity to the translational GTPase EFTu.

Highlights

ERF3 is an essential, conserved gene, whose essential function has remained obscure
The Rate Constant for Peptide Release by eukaryotic release factor 1 (eRF1) Is Modestly Stimulated by Eukaryotic peptide release factor 3 (eRF3) and Is Dependent on GTP Hydrolysis— We used a previously developed in vitro-reconstituted yeast translation system to evaluate the contributions of eRF3 to termination and recycling in eukaryotes [7]
Our kinetic measurements are consistent with previously published qualitative results [6, 18] and indicate that GTP hydrolysis by eRF3 plays a modest role in determining the rate of peptide release at saturating concentrations [7] of the release factors

Summary

Introduction

ERF3 is an essential, conserved gene, whose essential function has remained obscure. Results: eRF3 increases multiple turnover peptide release rates beyond the level expected from its stimulation of single turnover krel. The Rate Constant for Peptide Release (krel) by eRF1 Is Modestly Stimulated by eRF3 and Is Dependent on GTP Hydrolysis— We used a previously developed in vitro-reconstituted yeast translation system to evaluate the contributions of eRF3 to termination and recycling in eukaryotes [7].

Results

Conclusion