Table 3. Kinetic constants for the wild-type and mutant NK reactions with SUB1, and the results of three different types of FEP calculation.
Another substrate, SUB2, was used in FEP calculations. −∞, the results of FEP calculations trend to negative infinity; NA, not available.
| Enzyme | kcat (s−1) | Km (μM) | kcat/Km (s−1·mM−1) | ΔΔGT (kcal/mol) | ΔGsolv (kcal/mol) | ΔGbind (kcal/mol) | ΔGcat (kcal/mol) | ΔΔGbind (kcal/mol) | ΔΔGcat (kcal/mol) | ΔΔGb+c (kcal/mol) |
|---|---|---|---|---|---|---|---|---|---|---|
| Wild-type | 52.1 | 196 | 265.3 | NA | NA | NA | NA | NA | NA | NA |
| Mutant | ||||||||||
| S33A | 16.5 | 358 | 46.1 | 1.1 | 7.1 | 7.4 | 8.7 | 0.3 | 1.3 | 1.6 |
| D60A | 3.7 | 527 | 7.0 | 2.2 | −∞ | −∞ | −∞ | NA | NA | NA |
| S62A | 8.2 | 386 | 21.2 | 1.6 | 6.4 | 7.1 | 8.1 | 0.7 | 1.0 | 1.7 |
| T220A | 6.5 | 432 | 15.0 | 1.8 | 9.1 | 9.8 | 10.9 | 0.7 | 1.1 | 1.8 |