The energetic cost of domain reorientation in maltose-binding protein as studied by NMR and fluorescence spectroscopy
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Bibliographic record
Abstract
Maltose-binding protein (MBP) is a two-domain protein that undergoes a ligand-mediated conformational rearrangement from an "open" to a "closed" structure on binding to maltooligosaccharides. To characterize the energy landscape associated with this transition, we have generated five variants of MBP with mutations located in the hinge region of the molecule. Residual dipolar couplings, measured in the presence of a weak alignment medium, have been used to establish that the average structures of the mutant proteins are related to each other by domain rotation about an invariant axis, with the rotation angle varying from 5 degrees to 28 degrees. Additionally, the domain orientations observed in the wild-type apo and ligand-bound (maltose, maltotriose, etc.) structures are related through a rotation of 35 degrees about the same axis. Remarkably, the free energy of unfolding, measured by equilibrium denaturation experiments and monitored by fluorescence spectroscopy, shows a linear correlation with the rotation angle, with the stability of the (apo)protein decreasing with domain closure by 212 +/- 16 cal mol-1 per degree of rotation. The apparent binding energy for maltose also shows a similar correlation with the interdomain angle, suggesting that the mutations, as they relate to binding, affect predominantly the ligand-free structure. The linearity of the energy change is interpreted in terms of an increase in the extent of hydrophobic surface that becomes solvent accessible on closure. The combination of structural, stability, and binding data allows separation of the energetics of domain reorientation from ligand binding. This work presents a near quantitative structure-energy-binding relationship for a series of mutants of MBP, illustrating the power of combined studies involving protein engineering and solution NMR spectroscopy.
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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) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.001 |
| 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 it