Accessory active site residues of <i>Streptomyces</i> sp. N174 chitosanase
Bibliographic record
Abstract
The chitosanase from Streptomyces sp. N174 (CsnN174) is an inverting glycoside hydrolase belonging to family 46. Previous studies identified Asp40 as the general base residue. Mutation of Asp40 into glycine revealed an unexpectedly high residual activity. D40G mutation did not affect the stereochemical mechanism of catalysis or the mode of interaction with substrate. To explain the D40G residual activity, putative accessory catalytic residues were examined. Mutation of Glu36 was highly deleterious in a D40G background. Possibly, the D40G mutation reconfigured the catalytic center in a way that allowed Glu36 to be positioned favorably to perform catalysis. Thr45 was also found to be essential. Thr45 is thought to orientate the nucleophilic water molecule in a position to attack the glycosidic link. The finding that expression of heterologous CsnN174 in Escherichia coli protects cells against the antimicrobial effect of chitosan, allowed the selection of active chitosanase variants after saturation mutagenesis. Thr45 could be replaced only by serine, indicating the importance of the hydroxyl group. The newly identified accessory catalytic residues, Glu36 and Thr45 are located on a three-strand beta sheet highly conserved in GH19, 22, 23, 24 and 46, all members of the 'lysozyme superfamily'. Structural comparisons reveal that each family has its catalytic residues located among a small number of critical positions in this beta sheet. The position of Glu36 in CsnN174 is equivalent to general base residue in GH19 chitinases, whereas Thr45 is located similarly to the catalytic residue Asp52 of GH22 lysozyme. These examples reinforce the evolutionary link among these five GH families.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| 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) | 0.000 | 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 teacher head, 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".