Hydrogen bonds or salt bridges are usually formed to stabilize the buried ionizable residues. However, such interactions do not exist for two buried residues D271 and E305 of Trichoderma reesei Cel5A, an endoglucanase. Mutating D271 to alanine or leucine improves the enzyme thermostability quantified by the temperature T-50 due to the elimination of the desolvation penalty of the aspartic acid. However, the same mutations for E305 decrease the enzyme thermostability. Free energy calculations based on the molecular dynamics simulation predict the thermostability of D271A, D271L, and E305A (compared to WT) in line with the experimental observation but overestimate the thermostability of E305L. Quantum mechanical calculations suggest that the carboxylpeptide plane stacking interactions occurring to E305 but not D271 are important for the carboxyl group stabilization. For the protonated carboxyl group, the interaction energy can be as much as about -4 kcal/mol for parallel stacking and about -7 kcal/mol for T-shaped stacking. For the deprotonated carboxyl group, the largest interaction energies for parallel stacking and T-shaped stacking are comparable, about -7 kcal/mol. The solvation effect generally weakens the interaction, especially for the charged system. A search of the carboxylpeptide plane stacking in the PDB databank indicates that parallel stacking but not T-shaped stacking is quite common, and the most probable distance between the two stacking fragments is close to the value predicted by the QM calculations. This work highlights the potential role of carboxyl amide pi-pi stacking in the stabilization of aspartic acid and glutamic acid in proteins.