Structure of Ddi2, a highly inducible detoxifying metalloenzyme from Saccharomyces cerevisiae
Bibliographic record
Abstract
Cyanamide (H 2 N-CN) is used to break bud dormancy in woody plants and to deter alcohol use in humans. The biological effects of cyanamide in both these cases require the enzyme catalase. We previously demonstrated that Saccharomyces cerevisiae exposed to cyanamide resulted in strong induction of DDI2 gene expression. Ddi2 enzymatically hydrates cyanamide to urea and belongs to the family of HD-domain metalloenzymes (named after conserved active-site metal-binding His and Asp residues). Here, we report the X-ray structure of yeast Ddi2 to 2.6 Å resolution, revealing that Ddi2 is a dimeric zinc metalloenzyme. We also confirm that Ddi2 shares structural similarity with other known HD-domain proteins. HD residues His-55, His-88, and Asp-89 coordinate the active-site zinc, and the fourth zinc ligand is a water/hydroxide molecule. Other HD domain enzymes have a second aspartate metal ligand, but in Ddi2 this residue (Thr-157) does not interact with the zinc ion. Several Ddi2 active-site point mutations exhibited reduced catalytic activity. We kinetically and structurally characterized H137N and T157V mutants of Ddi2. A cyanamide soak of the Ddi2-T157V enzyme revealed cyanamide bound directly to the Zn 2+ ion, having displaced the zinc-bound water molecule. The mode of cyanamide binding to Ddi2 resembles cyanamide binding to the active-site zinc of carbonic anhydrase, a known cyanamide hydratase. Finally, we observed that the sensitivity of ddi2 Δ ddi3 Δ to cyanamide was not rescued by plasmids harboring ddi2-H137N or ddi2-TI57V variants, demonstrating that yeast cells require a functioning cyanamide hydratase to overcome cyanamide-induced growth defects.
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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 | 0.001 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| 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".