Table 1. Thermal stability of Atm1 decreases upon truncation of the C-terminal helix.
Thermal stability of wild-type Atm1 and the C-terminal L657 truncation mutant protein was measured by nano–differential scanning fluorimetry (nanoDSF) in DDM detergent solution. Measurements were performed in the absence (apo) or presence of AMP-PNP and Mg2+. The unfolding transition temperature was determined from the inflection point of the fluorescence signal (mean ± SD; n = 3). As a control, the specific ATPase activity of the proteins from above and the Walker B E598Q Atm1 mutant protein was determined by an NADH-coupled assay (mean ± SD; n ≥ 3).
| Thermal stability (Tm) (°C) | ||||
| apo | AMP-PNP-Mg2+ | |||
| Atm1 wild type | 42.5 | ±0.4 | 61.5 | ±2.7 |
| Atm1-L657 | 37.9 | ±1.4 | 56.1 | ±0.8 |
| Specific ATPase activity (nmol ATP min−1 mg−1 Atm1) | ||||
| Atm1 wild type | 175.9 | ±39.9 | ||
| Atm1 E598Q | 22.2 | ±10.0 | ||
| Atm1-L657 | 389.5 | ±39.9 | ||