Format

Send to

Choose Destination
Cell Rep. 2015 Aug 11;12(6):1032-41. doi: 10.1016/j.celrep.2015.07.007. Epub 2015 Jul 30.

Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine.

Author information

1
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
2
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
3
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: bobsauer@mit.edu.

Abstract

In the axial channels of ClpX and related hexameric AAA+ protein-remodeling rings, the pore-1 loops are thought to play important roles in engaging, mechanically unfolding, and translocating protein substrates. How these loops perform these functions and whether they also prevent substrate dissociation to ensure processive degradation by AAA+ proteases are open questions. Using ClpX pore-1-loop variants, single-molecule force spectroscopy, and ensemble assays, we find that the six pore-1 loops function synchronously to grip and unfold protein substrates during a power stroke but are not important in preventing substrate slipping between power strokes. The importance of grip strength is task dependent. ClpX variants with multiple mutant pore-1 loops translocate substrates as well as the wild-type enzyme against a resisting force but show unfolding defects and a higher frequency of substrate release. These problems are magnified for more mechanically stable target proteins, supporting a threshold model of substrate gripping.

PMID:
26235618
PMCID:
PMC4536184
DOI:
10.1016/j.celrep.2015.07.007
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center