Mechanistic Evidence for a Radical-Radical Recombination Pathway of Flavin-based Photocatalytic Tyrosine Labeling
Bibliographic record
Abstract
We recently introduced flavin-based photocatalysts such as riboflavin tetraacetate (RFT) as a robust platform for light- mediated protein labeling via phenoxyl radical-mediated tyrosine-biotin phenol coupling on live cells. To gain insight into this coupling reaction, we conducted detailed mechanistic analysis for RFT-photomediated activation of phenols for tyrosine labeling. Contrary to previously proposed mechanisms, we find that the initial covalent binding step between the tag and tyrosine is not radical addition, but rather radical-radical recombination. Radical addition to a phenol, while often proposed in the literature, is prohibitively unfavorable according to DFT calculations. Radical-radical recombination, followed by rearomatization, is the preferred pathway for the phenol-phenol coupling presented herein, but may also explain the mechanism of other tyrosine-tagging approaches described in the literature. Competitive kinetics experiments show that phenoxyl radicals are generated by the reaction of phenols with several reactive intermediates in the proposed mechanism: primarily with the riboflavin-photocatalyst excited state or singlet oxygen, but also possibly with the semi-reduced photocatalyst or hydroperoxyl radicals produced at key steps in the mechanism.
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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 | 0.001 |
| 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 | 0.001 | 0.001 |
| Research integrity | 0.001 | 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".