Structural Basis for the Altered Activity of Gly794 Variants of <i>Escherichia coli</i> β-Galactosidase<sup>†</sup>
Bibliographic record
Abstract
The open-closed conformational switch in the active site of Escherichia coli beta-galactosidase was studied by X-ray crystallography and enzyme kinetics. Replacement of Gly794 by alanine causes the apoenzyme to adopt the closed rather than the open conformation. Binding of the competitive inhibitor isopropyl thio-beta-D-galactoside (IPTG) requires the mutant enzyme to adopt its less favored open conformation, weakening affinity relative to wild type. In contrast, transition-state inhibitors bind to the enzyme in the closed conformation, which is favored for the mutant, and display increased affinity relative to wild type. Changes in affinity suggest that the free energy difference between the closed and open forms is 1-2 kcal/mol. By favoring the closed conformation, the substitution moves the resting state of the enzyme along the reaction coordinate relative to the native enzyme and destabilizes the ground state relative to the first transition state. The result is that the rate constant for galactosylation is increased but degalactosylation is slower. The covalent intermediate may be better stabilized than the second transition state. The substitution also results in better binding of glucose to both the free and the galactosylated enzyme. However, transgalactosylation with glucose to produce allolactose (the inducer of the lac operon) is slower with the mutant than with the native enzyme. This suggests either that the glucose is misaligned for the reaction or that the galactosylated enzyme with glucose bound is stabilized relative to the transition state for transgalactosylation.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 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 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
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 source (direct Gemma or distilled Codex), 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".