Rett syndrome causing missense mutations in the methyl-CpG-binding domain (MBD) of methyl CpG-binding protein 2 (MeCP2) were investigated both in silico and in vitro to reveal their effect on protein stability. It is demonstrated that the vast majority of frequently occurring mutations in the human population indeed alter the MBD folding free energy by a fraction of a kcal/mol up to more than 1 kcal/mol. While the absolute magnitude of the change of the free energy is small, the effect on the MBD functionality may be substantial since the folding free energy of MBD is about 2 kcal/mol only. Thus, it is emphasized that the effect of mutations on protein integrity should be evaluated with respect to the wild-type folding free energy but not with the absolute value of the folding free energy change. Furthermore, it was observed that the magnitude of the effect is correlated neither with the burial of the mutation sites nor with the basic amino acid physicochemical property change. Mutations that strongly perturb the immediate structural features were found to have little effect on folding free energy, while very conservative mutations resulted in large changes of the MBD stability. This observation was attributed to the protein's ability to structurally relax and reorganize to reduce the effect of mutation. Comparison between in silico and in vitro results indicated that some Web servers perform relatively well, while the free energy perturbation approach frequently overpredicts the magnitude of the free energy change especially when a charged amino acid is involved.