In order to elucidate the contribution of charged residues to protein stabilization at temperatures of over 100 °C, we constructed many mutants of the CutA1 protein ( EcCutA1) from Escherichia coli. The goal was to see if one can achieve the same stability as for a CutA1 from hyperthermophile Pyrococcus horikoshii that has the denaturation temperature near 150 °C. The hydrophobic mutant of EcCutA1 ( Ec0VV) with denaturation temperature ( Td) of 113.2 °C was used as a template for mutations. The highest Td of Ec0VV mutants substituted by a single charged residue was 118.4 °C. Multiple ion mutants were also constructed by combination of single mutants and found to have an increased thermostability. The highest stability of multiple mutants was a mutant substituted by nine charged residues that had a Td of 142.2 °C. To evaluate the energy of ion-ion interactions of mutant proteins, we used the structural ensemble obtained by a molecular dynamics simulation at 300 K. The Td of ionic mutants linearly increases with the increments of the computed energy of ion-ion interactions for ionic mutant proteins even up to the temperatures near 140 °C, suggesting that ion-ion interactions cumulatively contribute to the stabilization of a protein at high temperatures.