Stability analysis for the cavity-filling mutations of the Myb DNA-binding domain utilizing free-energy calculations
✍ Scribed by Hidetoshi Kono; Minoru Saito; Akinori Sarai
- Publisher
- John Wiley and Sons
- Year
- 2000
- Tongue
- English
- Weight
- 281 KB
- Volume
- 38
- Category
- Article
- ISSN
- 0887-3585
No coin nor oath required. For personal study only.
✦ Synopsis
We have analyzed the effect of cavity-filling mutations on protein stability by means of free-energy calculations based on molecular dynamics simulations to identify the factors contributing to stability changes caused by the mutations. We have studied the DNA-binding domain of Myb, which has a cavity in one of three homologous repeat units, and analyzed a series of mutations with nonnatural and natural amino acids at a single site, which change the size of the cavity. We found that the calculated free-energy changes caused by the mutations are in excellent agreement with experimental data (correlation coefficient 0.98). The free-energy changes in the native and denatured states were independently compared with the unfolding freeenergy change (⌬⌬G) and cavity-volume changes (⌬V), and it was found that ⌬⌬G and ⌬V correlate with the native-state free-energy changes but not with the denatured-state free-energy changes. Further analyses in terms of enthalpy and entropy show that compensation between entropy and enthalpy occurs in the denatured state but not in the native state. The main contribution to the native-state free energy was found to be van der Waals interactions associated with the cavity. We estimate that the decrease in free energy per methylene group, which results from filling the cavity, is about 2 to 3 kcal/ mol. These results suggest that the stabilization of a protein by cavity-filling mutations be determined primarily by the free energy associated with the cavity volume in the native state.