Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine
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 dissociat...
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| Format: | Article |
| Language: | en_US |
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Elsevier
2016
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| Online Access: | http://hdl.handle.net/1721.1/101700 https://orcid.org/0000-0003-1710-9582 https://orcid.org/0000-0002-1719-5399 |
| _version_ | 1826217400147640320 |
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| author | Iosefson, Ohad Olivares, Adrian O. Baker, Tania Sauer, Robert T |
| author2 | Massachusetts Institute of Technology. Department of Biology |
| author_facet | Massachusetts Institute of Technology. Department of Biology Iosefson, Ohad Olivares, Adrian O. Baker, Tania Sauer, Robert T |
| author_sort | Iosefson, Ohad |
| collection | MIT |
| description | 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. |
| first_indexed | 2024-09-23T17:03:02Z |
| format | Article |
| id | mit-1721.1/101700 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T17:03:02Z |
| publishDate | 2016 |
| publisher | Elsevier |
| record_format | dspace |
| spelling | mit-1721.1/1017002022-10-03T10:03:15Z Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine Iosefson, Ohad Olivares, Adrian O. Baker, Tania Sauer, Robert T Massachusetts Institute of Technology. Department of Biology Iosefson, Ohad Olivares, Adrian O. Baker, Tania Sauer, Robert T. 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. National Institutes of Health (U.S.) (Grant GM-101988) National Institutes of Health (U.S.) (Grant GM-049224) 2016-03-14T17:59:01Z 2016-03-14T17:59:01Z 2015-07 2015-06 Article http://purl.org/eprint/type/JournalArticle 22111247 http://hdl.handle.net/1721.1/101700 Iosefson, Ohad, Adrian O. Olivares, Tania A. Baker, and Robert T. Sauer. “Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine.” Cell Reports 12, no. 6 (August 2015): 1032–41. https://orcid.org/0000-0003-1710-9582 https://orcid.org/0000-0002-1719-5399 en_US http://dx.doi.org/10.1016/j.celrep.2015.07.007 Cell Reports Creative Commons Attribution http://creativecommons.org/licenses/by/4.0/ application/pdf Elsevier Elsevier |
| spellingShingle | Iosefson, Ohad Olivares, Adrian O. Baker, Tania Sauer, Robert T Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine |
| title | Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine |
| title_full | Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine |
| title_fullStr | Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine |
| title_full_unstemmed | Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine |
| title_short | Dissection of Axial-Pore Loop Function during Unfolding and Translocation by a AAA+ Proteolytic Machine |
| title_sort | dissection of axial pore loop function during unfolding and translocation by a aaa proteolytic machine |
| url | http://hdl.handle.net/1721.1/101700 https://orcid.org/0000-0003-1710-9582 https://orcid.org/0000-0002-1719-5399 |
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