Structural basis of ClpXP recognition and unfolding of ssrA-tagged substrates

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Abstract

When ribosomes fail to complete normal translation, all cells have mechanisms to ensure degradation of the resulting partial proteins to safeguard proteome integrity. In E. coli and other eubacteria, the tmRNA system rescues stalled ribosomes and adds an ssrA tag or degron to the C-terminus of the incomplete protein, which directs degradation by the AAA+ ClpXP protease. Here, we present cryo-EM structures of ClpXP bound to the ssrA degron. C-terminal residues of the ssrA degron initially bind in the top of an otherwise closed ClpX axial channel and subsequently move deeper into an open channel. For short-degron protein substrates, we show that unfolding can occur directly from the initial closed-channel complex. For longer-degron substrates, our studies illuminate how ClpXP transitions from specific recognition into a nonspecific unfolding and translocation machine. Many AAA+ proteases and protein-remodeling motors are likely to employ similar multistep recognition and engagement strategies.

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Structures have been deposited in PDB under the accession codes 6WSG, 6WR2, and 6WRF.

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Xue Fei

Department of Biology, Massachusetts Institute of Technology, Cambridge, United States

Competing interests
The authors declare that no competing interests exist.
Tristan A Bell

Department of Biology, Massachusetts Institute of Technology, Cambridge, United States

Competing interests
The authors declare that no competing interests exist.
Sarah R Barkow

Department of Chemistry, Massachusetts Institute of Technology, Cambridge, United States

Competing interests
The authors declare that no competing interests exist.
Tania A Baker

Department of Biology, Massachusetts Institute of Technology, Cambridge, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-0737-3411
Robert T Sauer

Department of Biology, Massachusetts Institute of Technology, Cambridge, United States

For correspondence
bobsauer@mit.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-1719-5399

Funding

National Institute of General Medical Sciences (GM-101988)

Robert T Sauer

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

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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.