Computational Analysis of the Mechanism of Nonenzymatic Peptide Bond Cleavage at the C-Terminal Side of an Asparagine Residue

[Image: see text] The nonenzymatic peptide bond cleavage at the C-terminal side of Asn residues is a protein post-translational modification that occurs under physiological conditions. This reaction proceeds much slower than the deamidation of the Asn side chain and causes denaturation and hypofunction of proteins. The peptide bond cleavage of Asn is detected primarily in crystallins and aquaporin 0 in the eye lens. Therefore, cleavage is thought to be involved in age-related cataracts. In this study, to clarify the mechanism underlying succinimide formation for the peptide bond cleavage of the Asn residue, we performed quantum chemical calculations on the model compound Ace-Asn-Gly-Nme (Ace = acetyl and Nme = methylamino). The density functional theory with the B3LYP/6-31+G(d,p) level of theory was used to obtain optimized geometries. The results suggested that the reaction proceeds through two steps, cyclization and C-terminal fragment release, and the required proton transfers can be mediated by H(2)PO(4)(–) and HCO(3)(–) ions. The conformational change of the main chain on the N-terminal side of Asn was needed for the C-terminal fragmentation step, and a separate conformational change at the C-terminal side was required for the cyclization step. Furthermore, the calculated activation barriers of the reactions catalyzed by the H(2)PO(4)(–) ion (130 kJ mol(–1)) and the HCO(3)(–) ion (123 kJ mol(–1)) were sufficiently low for the reactions to occur under normal physiological conditions.