C–H Bond Cleavage Is Rate-Limiting for Oxidative C–P Bond Cleavage by the Mixed Valence Diiron-Dependent Oxygenase PhnZ
Bibliographic record
Abstract
PhnZ utilizes a mixed valence diiron(II/III) cofactor and O2 to oxidatively cleave the carbon–phosphorus bond of (R)-2-amino-1-hydroxyethylphosphonic acid to form glycine and orthophosphate. The active site residues Y24 and E27 are proposed to mediate induced-fit recognition of the substrate and access of O2 to one of the active site Fe ions. H62 is proposed to deprotonate the C1-hydroxyl of the substrate during catalysis. Kinetic isotope effects (KIEs), pH–rate dependence, and site-directed mutagenesis were used to probe the rate-determining transition state and the roles of these three active site residues. Primary deuterium KIE values of 5.5 ± 0.3 for D(V) and 2.2 ± 0.4 for D(V/K) were measured with (R)-2-amino[1-2H1]-1-hydroxyethylphosphonic acid, indicating that cleavage of the C1–H bond of the substrate is rate-limiting. This step is also rate-limiting for PhnZ Y24F, as shown by a significant deuterium KIE value of 2.3 ± 0.1 for D(V). In contrast, a different reaction step appears to be rate-limiting for the PhnZ E27A and H62A variants, which exhibited D(V) values near unity. A solvent KIE of 2.2 ± 0.3 for D2O(V) is observed for PhnZ. Significant solvent KIE values are also observed for the PhnZ Y24F and E27A variants. In contrast, the PhnZ H62A variant does not show a significant solvent KIE, suggesting that H62 is mediating proton transfer in the transition state. A proton inventory study with PhnZ indicates that 1.5 ± 0.6 protons are in flight in the rate-determining step. Overall, the rate-determining transition state for oxidative C–P bond cleavage by PhnZ is proposed to involve C–H bond cleavage that is coupled to deprotonation of the substrate C1-hydroxyl by H62.
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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 | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.001 |
| 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) | 0.001 | 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 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".