Bibliographic record
Abstract
The AAA+ chaperone Hsp104 mediates the reactivation of aggregated proteins in Saccharomyces cerevisiae and is crucial for cell survival after exposure to stress. Protein disaggregation depends on cooperation between Hsp104 and a cognate Hsp70 chaperone system. Hsp104 forms a hexameric ring with a narrow axial channel penetrating the centre of the complex. In Chapter 2, I show that conserved loops in each AAA+ module that line this channel are required for disaggregation and that the position of these loops is likely determined by the nucleotide bound state of Hsp104. This evidence supports a common protein remodeling mechanism among Hsp100 members in which proteins are unfolded and threaded along the axial channel. In Chapter 3, I use a peptide-based substrate mimetic to reveal other novel features of Hsp104’s disaggregation mechanism. An Hsp104-binding peptide selected from solid phase arrays recapitulated several properties of an authentic Hsp104 substrate. Inactivation of the pore loops in either AAA+ module prevented stable peptide or protein binding. However, when the loop in the first AAA+ was inactivated, stimulation of ATPase turnover in the second AAA+ module of this mutant was abolished. Drawing on these data, I propose a detailed mechanistic model of protein unfolding by Hsp104 in which an initial unstable interaction involving the loop in the first AAA+ module simultaneously promotes penetration of the substrate into the second axial channel binding site and activates ATP turnover in the second AAA+ module. In Chapter 4, I explore the recognition elements within a model Hsp104-binding peptide that are required for rapid binding to Hsp104. Removal of bulky hydrophobic residues and lysines abrogated the ability of this peptide to function as a peptide-based substrate mimetic for Hsp104. Furthermore, rapid binding of a model unfolded protein to Hsp104 required an intact N-terminal domain and ATP binding at the first AAA+ module. Taken together, I have defined numerous structural features within Hsp104 and its model substrates that are crucial for substrate binding and processing by Hsp104. This work provides a theoretical framework that will encourage research in other protein remodeling AAA+ ATPases.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.001 |
| Meta-epidemiology (broad) | 0.001 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.001 | 0.001 |
| Scholarly communication | 0.002 | 0.003 |
| Open science | 0.002 | 0.001 |
| Research integrity | 0.002 | 0.004 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".