Table 5.
Apparent dissociation constants (Kdapp), Hill coefficients and variations in binding free energy (ΔΔG) for wild-type and mutant p53 N-D-T binding to fluorescently labeled double stranded oligonucleotides*
| p53 N-D-T mutant | Wild Type | K120E | H178D | E180K | D186A | R248Q | C277W | H178D E180K | Bac-p53 |
|---|---|---|---|---|---|---|---|---|---|
| p21 promoter | |||||||||
| Kdapp. (nM) | ≤6.4 | ≤6.4 | 18±8 | 13±3 | 6±3 | 110±10 | ≤6.4 | 11±10 | 6.2±0.6 |
| Hill Coefficient | 2.05 | 2.04 | 1.05 | 1.01 | 1.49 | 0.59 | 1.65 | 1.14 | 1.59 |
| ΔΔG Wild Type** | 0 | N.D. | +0.61 | +0.42 | N.D. | +1.68 | N.D. | +0.30 | n.d. |
|
| |||||||||
| puma promoter | |||||||||
| Kdapp. (nM) | ≤6.4 | 150±20 | 15±6 | 19±5 | 7±3 | 140±40 | 130±10 | 10±4 | 7.0±0.4 |
| Hill Coefficient | 1.78 | 0.69 | 1.13 | 1.08 | 1.55 | 0.66 | 0.63 | 1.27 | 1.50 |
| ΔΔG Wild Type** | 0 | +1.86 | +0.50 | +0.63 | +0.05 | +1.81 | +1.80 | +0.25 | n.d. |
*
Error ranges indicate uncertainties of fit for curves obtained from the average of three independent titrations.
**
Determined as follows: ΔΔG = −RT*ln(Kdapp.mut/Kdapp.wild type); values in Kcal/mol