The ionization effects on 28 conformations of N -glycylglycine are analyzed by means of the hybrid B3LYP and the hybrid meta-MPWB1K density functionals and by single-point calculations at the CCSD(T) level of theory. The most favorable process observed corresponds to the ionization of the only neutral conformation that presents a OHN H2 intramolecular hydrogen bond, which leads to C O2 elimination after a spontaneous proton transfer from -COOH to N H2. The remaining neutral structures evolve to 20 different conformations of N -glycylglycine radical cation, which lie about 25-40 kcalmol higher than the decarboxylated [N H3 C H2 CONHC H2] + [C O2] complex. Structural changes induced by ionization depend on the intramolecular hydrogen bonds of the initial conformation, since they determine the nature of the electron hole formed. In most cases, ionization takes place at the terminal - N H2 and -CO of the amide bond, which produces a strengthening of the peptide bond and the formation of new - N H2 O Camide and - N H2 OCOH hydrogen bonds. However, if - N H2 and - C Oamide simultaneously act as proton acceptor in the neutral conformation, ionization is mainly localized at the carboxylic group, which produces a strengthening of the - COOHO Camide bond. Both functionals lead to similar trends and compare well with CCSD(T) results except for a few cases for which B3LYP provides a too delocalized picture of the electron hole and consequently leads to artificial geometry reorganization. © 2006 American Institute of Physics.