Circular permutation of a bacterial tyrosinase enables efficient polyphenol‐specific oxidation and quantitative preparation of orobol
Bibliographic record
Abstract
Tyrosinase is a type 3 copper oxygenase that catalyzes a phenol moiety into ortho-diphenol, and subsequently to ortho-quinone. Diverse tyrosinases have been observed across the kingdom including Animalia, Bacteria, Plantae, and Fungi. Among the tyrosinases, bacterial, and mushroom tyrosinases have been extensively exploited to prepare melanin, ortho-hydroxy-polyphenols, or novel plant secondary metabolites during the past decade. And their use as a biocatalyst to prepare various functional biocompounds have drawn great attention worldwide. Herein, we tailored a bacterial tyrosinase from Bacillus megaterium (BmTy) using circular permutation (CP) engineering technique which is a novel enzyme engineering technique to covalently link original N and C termini and create new termini on the middle of its polypeptide. To construct a smart rationally-designed CP library, we introduced 18 new termini at the edge of each nine loops that link α-helical secondary structure in BmTy. Among the small library, seven functional CP variants were successfully identified and they represented dramatic change in their enzyme characteristics including kinetic properties and substrate specificity. Especially, cp48, 102, and 245 showed dramatically decreased tyrosine hydroxylase activity, behaving like a catechol oxidase. Exploiting the dramatic increased polyphenol oxidation activity of cp48, orobol (3'-hydroxy-genistein) was quantitatively synthesized with 1.48 g/L, which was a 6-fold higher yield of truncated wild-type. We examined their kinetic characters through structural speculation, and suggest a strategy to solubilize the insoluble artificial variants effectively.
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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".