Recombinant Nepenthesin II for Hydrogen/Deuterium Exchange Mass Spectrometry
Bibliographic record
Abstract
The pitcher secretions of the Nepenthes genus of carnivorous plants contain a proteolytic activity that is very useful for hydrogen/deuterium exchange mass spectrometry (HX-MS). Our efforts to reconstitute pitcher fluid activity using recombinant nepenthesin I (one of two known aspartic proteases in the fluid) revealed a partial cleavage profile and reduced enzymatic stability in certain HX-MS applications. We produced and characterized recombinant nepenthesin II to determine if it complemented nepenthesin I in HX-MS applications. Nepenthesin II shares many properties with nepenthesin I, such as fast digestion at reduced temperature and pH, and broad cleavage specificity, but in addition, it cleaves C-terminal to tryptophan. Neither enzyme reproduces the C-terminal proline cleavage we observed in the natural extract. Nepenthesin II is considerably more resistant to chemical denaturants and reducing agents than nepenthesin I, and it possesses a stability profile that is similar to that of pepsin. Higher stability combined with the slightly broader cleavage specificity makes nepenthesin II a useful alternative to pepsin and a more complete replacement for pitcher fluid in HX-MS applications.
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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 | 0.000 |
| 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 | 0.001 |
| Insufficient payload (model declined to judge) | 0.002 | 0.002 |
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".