Mechanism of ribosome rescue by ArfA and RF2

Abstract
Data availability
Article and author information
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Abstract

ArfA rescues ribosomes stalled on truncated mRNAs by recruiting release factor RF2, which normally binds stop codons to catalyze peptide release. We report two 3.2-Å resolution cryo-EM structures – determined from a single sample – of the 70S ribosome with ArfA•RF2 in the A site. In both states, the ArfA C-terminus occupies the mRNA tunnel downstream of the A site. One state contains a compact inactive RF2 conformation. Ordering of the ArfA N-terminus in the second state rearranges RF2 into an extended conformation that docks the catalytic GGQ motif into the peptidyl-transferase center. Our work thus reveals the structural dynamics of ribosome rescue. The structures demonstrate how ArfA “senses” the vacant mRNA tunnel and activates RF2 to mediate peptide release without a stop codon, allowing stalled ribosomes to be recycled.

Data availability

The following data sets were generated

(2017) 3.2 A cryo-EM ArfA-RF2 ribosome rescue complex (Structure I)
Publicly available at the RCSB Protein Data Bank (accession no: 5U9G).

http://www.rcsb.org/pdb/explore/explore.do?structureId=5U9G
(2017) 3.2 A cryo-EM ArfA-RF2 ribosome rescue complex (Structure I)
Publicly available at EMDataBank (accession no. EMD-8522).

http://emsearch.rutgers.edu/atlas/8522_summary.html
(2017) 3.2 A cryo-EM ArfA-RF2 ribosome rescue complex (Structure II)
Publicly available at the RCSB Protein Data Bank (accession no: 5U9F).

http://www.rcsb.org/pdb/explore/explore.do?structureId=5U9F
1. Demo G
2. Svidritskiy E
(2017) 3.2 A cryo-EM ArfA-RF2 ribosome rescue complex (Structure II)
Publicly available at EMDataBank (accession no. EMD-8521).

http://emsearch.rutgers.edu/atlas/8521_summary.html

Article and author information

Author details

Gabriel Demo

RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States

Competing interests
No competing interests declared.
Egor Svidritskiy

RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States

Competing interests
No competing interests declared.
Rohini Madireddy

Medicago, Durham, United States

Competing interests
No competing interests declared.
Ruben Diaz-Avalos

Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States

Competing interests
No competing interests declared.
Timothy Grant

Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States

Competing interests
No competing interests declared.
Nikolaus Grigorieff

Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States

Competing interests
Nikolaus Grigorieff, Reviewing editor, eLife.

"This ORCID iD identifies the author of this article:" 0000-0002-1506-909X
Duncan Sousa

Department of Biological Science, Florida State University, Tallahassee, United States

Competing interests
No competing interests declared.
Andrei A Korostelev

RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States

For correspondence
andrei.korostelev@umassmed.edu

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0003-1588-717X

Funding

National Institutes of Health (GM106105)

Andrei A Korostelev

National Institutes of Health (GM107465)

Andrei A Korostelev

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.