Table II. DNA binding and repair by R128 mutants.
| Protein | Binding to O6–methyl-containing oligonucleotide |
Rate of repair |
||
|---|---|---|---|---|
| Double stranded KD (μM) | Single stranded KD (μM) | O6–BG k2 (M–1 min–1 × 10–2) | Methylated DNA k2 (M–1 min–1 × 10–6) | |
| Wild type | NMa | NMa | 6.1 | 38 |
| C145A | 0.13 | 0.69 | inactive | inactive |
| R128K | NMa | NMa | 7.3 | 24 |
| R128L | 2.6 | 6.1 | 7.1 | 10 |
| R128E | 3.4 | 11 | 7.8 | 0.6 |
| R128D | 4.2 | 22 | 5.1 | 0.03 |
| R128A | 3.1 | 28 | 8.4 | 0.02 |
| R128G | ND | ND | 6.1 | <0.006 |
ND, not determined.
aRate of repair prohibits the use of a gel shift assay. The inactive C145A mutant is included as a surrogate of these values. There was no difference in the binding of wild type and C145A to oligo– nucleotides that do not contain O6–methylguanine (KD = 0.6 μM for double-stranded DNA, 1.0 μM for single-stranded DNA).