Protein and Peptide Alkoxyl Radicals Can Give Rise to C-Terminal Decarboxylation and Backbone Cleavage

Previous studies have demonstrated that γ-irradiation of some free amino acids in the presence of oxygen gives high yields of side-chain hydroperoxides. It is shown in the present study thatN-acetyl amino acids and peptides also give high levels of hydroperoxides on γ-irradiation, even when the free amino acid does not, and that hydroperoxides can be formed on both the backbone (at α-carbon positions) and the side chain. Decomposition of α-carbon hydroperoxides by Fe(II)–EDTA gives initially an alkoxyl radical via a pseudo-Fenton reaction; these radicals fragment rapidly withkestimated as ≥107s−1. WithN-acetyl amino acids and dipeptides β-scission of an alkoxyl radical at the C-terminal α-carbon results in C-terminal decarboxylation, with release of CO−2; the corresponding amides undergo deamidation with release of C(O)NH2. Cyclic dipeptides undergo analogous reactions with cleavage of the α-carbon to carbonyl–carbon bond and formation of °C(O)NHR radicals. With substrates with large aliphatic side chains, radicals from side-chain hydroperoxides are also observed. C-terminal decarboxylation and backbone fragmentation are also observed with larger peptides, amino acid homopolymers, and proteins. These observations suggest that α-carbon alkoxyl radicals may be key intermediates in the fragmentation of proteins in the presence of oxygen. The radicals released in these processes may react further to form O−2, or redox cycle metal ions. These reactions may be propagating processes during protein chain oxidation.