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Eukaryotic release factor 3 is required for multiple turnovers of peptide release catalysis by eukaryotic release factor 1.

Eyler DE, Wehner KA, Green R - J. Biol. Chem. (2013)

Bottom Line: This effect was generalizable across all stop codons and in a variety of contexts.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.

View Article: PubMed Central - PubMed

Affiliation: From the Howard Hughes Medical Institute and the Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.

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.

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Model of the roles of eRF1 and eRF3 in eukaryotic termination. A simplified model for eukaryotic termination, emphasizing the steps at which eRF1 may be influenced by eRF3. krelease and ksplitting refer to the processes of peptide release and subunit dissociation, respectively. These two steps could be directly measured in our kinetic analysis; binding steps (indicated with equilibria) have not been directly monitored here.
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Figure 5: Model of the roles of eRF1 and eRF3 in eukaryotic termination. A simplified model for eukaryotic termination, emphasizing the steps at which eRF1 may be influenced by eRF3. krelease and ksplitting refer to the processes of peptide release and subunit dissociation, respectively. These two steps could be directly measured in our kinetic analysis; binding steps (indicated with equilibria) have not been directly monitored here.

Mentions: Numerous hypotheses have been advanced regarding the essential function of eRF3 in eukaryotic cells (6, 15, 23, 26). To gain further insight into eRF3 function, we systematically characterized the contributions of eRF3 to the kinetics of eRF1-mediated peptide release and subunit dissociation. We have found that eRF3:GTP makes modest (5-fold) contributions to the rate constant for peptide release (krel) catalyzed by eRF1 (Fig. 5, krelease) and that this effect is broadly consistent across stop codons and different contexts. When eRF1 is limiting, however, the effect of eRF3:GTP is more substantial, with >20-fold increases in the initial rates (kobs) of the reaction and substantial increases in end point from <10% to 90%.


Eukaryotic release factor 3 is required for multiple turnovers of peptide release catalysis by eukaryotic release factor 1.

Eyler DE, Wehner KA, Green R - J. Biol. Chem. (2013)

Model of the roles of eRF1 and eRF3 in eukaryotic termination. A simplified model for eukaryotic termination, emphasizing the steps at which eRF1 may be influenced by eRF3. krelease and ksplitting refer to the processes of peptide release and subunit dissociation, respectively. These two steps could be directly measured in our kinetic analysis; binding steps (indicated with equilibria) have not been directly monitored here.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3795251&req=5

Figure 5: Model of the roles of eRF1 and eRF3 in eukaryotic termination. A simplified model for eukaryotic termination, emphasizing the steps at which eRF1 may be influenced by eRF3. krelease and ksplitting refer to the processes of peptide release and subunit dissociation, respectively. These two steps could be directly measured in our kinetic analysis; binding steps (indicated with equilibria) have not been directly monitored here.
Mentions: Numerous hypotheses have been advanced regarding the essential function of eRF3 in eukaryotic cells (6, 15, 23, 26). To gain further insight into eRF3 function, we systematically characterized the contributions of eRF3 to the kinetics of eRF1-mediated peptide release and subunit dissociation. We have found that eRF3:GTP makes modest (5-fold) contributions to the rate constant for peptide release (krel) catalyzed by eRF1 (Fig. 5, krelease) and that this effect is broadly consistent across stop codons and different contexts. When eRF1 is limiting, however, the effect of eRF3:GTP is more substantial, with >20-fold increases in the initial rates (kobs) of the reaction and substantial increases in end point from <10% to 90%.

Bottom Line: This effect was generalizable across all stop codons and in a variety of contexts.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.

View Article: PubMed Central - PubMed

Affiliation: From the Howard Hughes Medical Institute and the Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.

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.

Show MeSH
Related in: MedlinePlus