Nitrogen-centered and β-carbon-centered hydrogen-deficient peptide radicals are considered to be intermediates in the matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD)-induced C-C bond cleavage of peptide backbones when using an oxidizing matrix. To understand the general mechanism of C-C bond cleavage by MALDI-ISD， I study the fragmentation of model peptides and investigate the fragment formation pathways using calculations with density functional theory and transition state theory. The calculations indicate that the nitrogen-centered radical immediately undergoes C-C bond cleavage， leading to the formation of an a•/x fragment pair. In contrast， the dissociation of the β-carbon-... More
Nitrogen-centered and β-carbon-centered hydrogen-deficient peptide radicals are considered to be intermediates in the matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD)-induced C-C bond cleavage of peptide backbones when using an oxidizing matrix. To understand the general mechanism of C-C bond cleavage by MALDI-ISD， I study the fragmentation of model peptides and investigate the fragment formation pathways using calculations with density functional theory and transition state theory. The calculations indicate that the nitrogen-centered radical immediately undergoes C-C bond cleavage， leading to the formation of an a•/x fragment pair. In contrast， the dissociation of the β-carbon-centered radical is kinetically feasible under MALDI-ISD conditions， leading to the formation of an a/x• fragment pair. To discriminate these processes， I focus on the yield of d fragments， which originate from a• radicals through radical-induced side-chain loss， not from a fragments. The C-C bond cleavage on the C-terminal side of the carbamidomethylated cysteine residue is found to produce d fragments instead of a fragments. According to the calculation of the rate constant， the corresponding fragmentation occurs within 1 ns. The intense signal arising from d fragments and the lack of or weak signal from a fragments strongly suggest that the C-C bond cleavage occurs through a nitrogen-centered radical intermediate. In addition to the side-chain loss， the resulting a• radical undergoes hydrogen atom abstraction by the matrix. The results for a deuterium-labeled peptide indicate that the matrix abstracts a hydrogen atom from either the amide nitrogen or the β-carbon.
