Accessibility of Carboxypeptidase A-bound Zinc to Chelation Correlates with an Intermediate State in the Protein’s Unfolding Pathway
Bibliographic record
Abstract
In view of the emerging role of metal ions in improper protein folding (a phenomenon associated with a variety of diseases), new tools to characterize structural changes that accompany folding transitions are highly sought. Using a combination of fluorescence spectroscopy and studies involving the chromophoric chelator 4-(2-pyridylazo)resorcinol (PAR), we here show that the prototypical zinc protease carboxypeptidase A (CPA) unfolds in the presence of guanidine hydrochloride via a previously unidentified folding intermediate that resembles a molten globular state and retains the zinc ion. The spontaneous dissociation of the metal ion from CPA was observed only upon transition of the intermediate to the fully unfolded state of the protein. Furthermore, an analysis of zinc ion binding during CPA unfolding using PAR revealed the intermediate state to directly correlate with the ability of the chelator to gain access to the active site and to associate with the protein-bound metal ion. This observation is indicative of CPA's active site being PAR-inaccessible in the native state but becoming PAR-accessible in the folding intermediate. Taken together, the current study demonstrates the usefulness of PAR as a simple spectrophotometric tool to assess structural changes during the unfolding of CPA and potentially other zinc proteins. Hence, zinc accessibility probes (ZAPs) such as PAR may find utility in gaining further insight into the mechanism(s) of metalloprotein folding.
Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.
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) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
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".