Enzymatic Post-Transfer Editing Mechanism of <i>E. coli</i> Threonyl-tRNA Synthetase (ThrRS): A Molecular Dynamics (MD) and Quantum Mechanics/Molecular Mechanics (QM/MM) Investigation
Bibliographic record
Abstract
Threonyl-tRNA synthetase (ThrRS) is a class II aminoacyl-tRNA synthetase (aaRS), which are a ubiquitous family of enzymes that have a vital role in protein biosynthesis. In particular, it catalyzes the activation and subsequent aminoacylation of its corresponding tRNA Thr . Because of the close structural and electronic similarity between its cognate substrate threonine and the noncognate serine, the catalytic aminoacylation site of ThrRS is not able to fully discriminate between them. In this study we have explored multiple possible post-transfer editing mechanisms for ThrRS from Escherichia coli . The editing site is known to contain two conserved histidyls (His73 and His186) and a cysteinyl (Cys182), all of which could act as the required mechanistic base. We have performed detailed molecular dynamics (MD) and quantum mechanics/molecular mechanics (QM/MM) studies in which the protonation states of each of these residues was varied. Furthermore, using the various substrate-bound active site models obtained, we have examined previously proposed and alternative possible mechanisms for deaminoacylation of Ser-tRNA Thr by ThrRS in which His73 or Cys182 acts as the base: 11 mechanisms in total. The present results suggest that the most feasible mechanism is obtained when both His73 and His186 are neutral, while the thiol of Cys182 is deprotonated and acts as a base. The resulting mechanism is found to occur in two steps. First, deprotonation of an active site water by the thiolate of Cys182 with its concomitant nucleophilic attack at the substrate’s C carb center occurs with a calculated free energy barrier of 9.9 kcal/mol. The subsequent, and overall rate-limiting step, is a water-meditated proton transfer from Lys156 onto the Ado76 3′-oxygen resulting in simultaneous cleavage of the Ado76 3′O–C carb bond with a free energy barrier of 20.8 kcal/mol.
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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) | 0.001 | 0.001 |
| Meta-epidemiology (broad) | 0.001 | 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 | 0.000 |
| 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".