Characterization of the dehydratase WcbK and the reductase WcaG involved in GDP-6-deoxy-<i>manno</i>-heptose biosynthesis in <i>Campylobacter jejuni</i>
Bibliographic record
Abstract
The capsule of Campylobacter jejuni strain 81-176 comprises the unusual 6-deoxy-α-D-altro-heptose, whose biosynthesis and function are not known. In the present study, we characterized enzymes of the capsular cluster, WcbK and WcaG, to determine their role in 6-deoxy-altro-heptose synthesis. These enzymes are similar to the Yersinia pseudotuberculosis GDP-manno-heptose dehydratase/reductase DmhA/DmhB that we characterized previously. Capillary electrophoresis and MS analyses showed that WcbK is a GDP-manno-heptose dehydratase whose product can be reduced by WcaG, and that WcbK/WcaG can use the substrate GDP-mannose, although with lower efficiency than heptose. Comparison of kinetic parameters for WcbK and DmhA indicated that the relaxed substrate specificity of WcbK comes at the expense of catalytic performance on GDP-manno-heptose. Moreover, although WcbK/WcaG and DmhA/DmhB are involved in altro- versus manno-heptose synthesis respectively, the enzymes can be used interchangeably in mixed reactions. NMR spectroscopy analyses indicated conservation of the sugar manno configuration during catalysis by WcbK/WcaG. Therefore additional capsular enzymes may perform the C3 epimerization necessary to generate 6-deoxy-altro-heptose. Finally, a conserved residue (Thr(187) in WcbK) potentially involved in substrate specificity was identified by structural modelling of mannose and heptose dehydratases. Site-directed mutagenesis and kinetic analyses demonstrated its importance for enzymatic activity on heptose and mannose substrates.
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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) | 0.000 | 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 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".