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GTP-dependent structural rearrangement of the eRF1:eRF3 complex and eRF3 sequence motifs essential for PABP binding

Kononenko, Artem V. ; Mitkevich, Vladimir A. ; Atkinson, Gemma C. LU orcid ; Tenson, Tanel ; Dubovaya, Vera I. ; Frolova, Ludmila Yu ; Makarov, Alexander A. and Hauryliuk, Vasili LU orcid (2009) In Nucleic Acids Research 38(2). p.548-558
Abstract

Translation termination in eukaryotes is governed by the concerted action of eRF1 and eRF3 factors. eRF1 recognizes the stop codon in the A site of the ribosome and promotes nascent peptide chain release, and the GTPase eRF3 facilitates this peptide release via its interaction with eRF1. In addition to its role in termination, eRF3 is involved in normal and nonsense-mediated mRNA decay through its association with cytoplasmic poly(A)-binding protein (PABP) via PAM2-1 and PAM2-2 motifs in the N-terminal domain of eRF3. We have studied complex formation between full-length eRF3 and its ligands (GDP, GTP, eRF1 and PABP) using isothermal titration calorimetry, demonstrating formation of the eRF1:eRF3:PABP:GTP complex. Analysis of the... (More)

Translation termination in eukaryotes is governed by the concerted action of eRF1 and eRF3 factors. eRF1 recognizes the stop codon in the A site of the ribosome and promotes nascent peptide chain release, and the GTPase eRF3 facilitates this peptide release via its interaction with eRF1. In addition to its role in termination, eRF3 is involved in normal and nonsense-mediated mRNA decay through its association with cytoplasmic poly(A)-binding protein (PABP) via PAM2-1 and PAM2-2 motifs in the N-terminal domain of eRF3. We have studied complex formation between full-length eRF3 and its ligands (GDP, GTP, eRF1 and PABP) using isothermal titration calorimetry, demonstrating formation of the eRF1:eRF3:PABP:GTP complex. Analysis of the temperature dependence of eRF3 interactions with G nucleotides reveals major structural rearrangements accompanying formation of the eRF1:eRF3:GTP complex. This is in contrast to eRF1:eRF3:GDP complex formation, where no such rearrangements were detected. Thus, our results agree with the established active role of GTP in promoting translation termination. Through point mutagenesis of PAM2-1 and PAM2-2 motifs in eRF3, we demonstrate that PAM2-2, but not PAM2-1 is indispensible for eRF3:PABP complex formation.

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author
; ; ; ; ; ; and
publishing date
type
Contribution to journal
publication status
published
in
Nucleic Acids Research
volume
38
issue
2
article number
gkp908
pages
548 - 558
publisher
Oxford University Press
external identifiers
  • scopus:77449158439
  • pmid:19906736
ISSN
0305-1048
DOI
10.1093/nar/gkp908
language
English
LU publication?
no
additional info
Funding Information: Presidium of the Russian Academy of Sciences (Program Molecular and Cell Biology to A.A.M. and L.Yu.F.); the Russian Foundation for Basic Research (08-04-00375a to L.Yu.F.); Estonian Science Foundation grants (6768 to T.T. and 7616 to V.H.); Grant of the President of the Russian Federation for young scientists (MK-162.2009.4 to V.A.M.); and European Regional Development Fund through the Center of Excellence in Chemical Biology (to V.H. and T.T.). Funding for open access charge: Personal grant from European Regional Development Fund through the Center of Excellence in Chemical Biology (to V.H.). Copyright: Copyright 2010 Elsevier B.V., All rights reserved.
id
c73e46c4-62a0-4d01-bdd6-a5017e17497b
date added to LUP
2021-09-24 20:49:28
date last changed
2025-01-25 03:16:05
@article{c73e46c4-62a0-4d01-bdd6-a5017e17497b,
  abstract     = {{<p>Translation termination in eukaryotes is governed by the concerted action of eRF1 and eRF3 factors. eRF1 recognizes the stop codon in the A site of the ribosome and promotes nascent peptide chain release, and the GTPase eRF3 facilitates this peptide release via its interaction with eRF1. In addition to its role in termination, eRF3 is involved in normal and nonsense-mediated mRNA decay through its association with cytoplasmic poly(A)-binding protein (PABP) via PAM2-1 and PAM2-2 motifs in the N-terminal domain of eRF3. We have studied complex formation between full-length eRF3 and its ligands (GDP, GTP, eRF1 and PABP) using isothermal titration calorimetry, demonstrating formation of the eRF1:eRF3:PABP:GTP complex. Analysis of the temperature dependence of eRF3 interactions with G nucleotides reveals major structural rearrangements accompanying formation of the eRF1:eRF3:GTP complex. This is in contrast to eRF1:eRF3:GDP complex formation, where no such rearrangements were detected. Thus, our results agree with the established active role of GTP in promoting translation termination. Through point mutagenesis of PAM2-1 and PAM2-2 motifs in eRF3, we demonstrate that PAM2-2, but not PAM2-1 is indispensible for eRF3:PABP complex formation.</p>}},
  author       = {{Kononenko, Artem V. and Mitkevich, Vladimir A. and Atkinson, Gemma C. and Tenson, Tanel and Dubovaya, Vera I. and Frolova, Ludmila Yu and Makarov, Alexander A. and Hauryliuk, Vasili}},
  issn         = {{0305-1048}},
  language     = {{eng}},
  number       = {{2}},
  pages        = {{548--558}},
  publisher    = {{Oxford University Press}},
  series       = {{Nucleic Acids Research}},
  title        = {{GTP-dependent structural rearrangement of the eRF1:eRF3 complex and eRF3 sequence motifs essential for PABP binding}},
  url          = {{http://dx.doi.org/10.1093/nar/gkp908}},
  doi          = {{10.1093/nar/gkp908}},
  volume       = {{38}},
  year         = {{2009}},
}