General Mechanism of C_α–C Peptide Backbone Bond Cleavage in Matrix-Assisted Laser Desorption/Ionization In-Source Decay Mediated by Hydrogen Abstraction

Cooperative dissociation of peptide backbones and side‐chains during matrix‐assisted laser desorption/ionization in‐source decay mediated by hydrogen abstraction

“…Finally, [a (N═C) − H], [a (C═C) − H], and [d − H] fragments are generated from the [a]• radicals. 18 The hydrogen abstraction from [a]• radicals preferentially occurs for the majority of amino acid residues, and the protonated form of [a − H], that is, [a] + fragments, can be observed as intense signals in the corresponding MALDI-ISD mass spectrum. Consequently, peptide and protein sequencing by MALDI-ISD with an oxidizing matrix are generally conducted by interpreting the mass differences between series of consecutive [a] + fragments.…”

“…In contrast, the cleavage of a C α -C bond at the C-terminal side of cysteine (Cys) and carbamidomethylated Cys residues was reported to produce intense signals due to [d] + fragment rather than [a] + fragment. 18 Such signal originating from the bond cleavage at the C-terminal side of Cys residues can be explained by the high dissociation rate constant of the corresponding [a]• radicals. In addition, the yield of [d] + fragments observed in a MALDI-ISD mass spectrum is dependent on the dissociation rate constant of the corresponding [a]• radicals.…”

“…In addition, the yield of [d] + fragments observed in a MALDI-ISD mass spectrum is dependent on the dissociation rate constant of the corresponding [a]• radicals. 18 Therefore, [a]• radicals are recognized as intermediates in the MALDI-ISD process. In contrast, the C α -C bonds located on the C-terminal side of the radical site in nitrogen-centered radicals produce [a (N═C) − H] fragment, which does not undergo side-chain loss (Scheme 1B).…”

“…In addition, the density functional theory (DFT) calculation also suggests that the C α -C bonds located on the N-terminal side of the radical site in nitrogen-centered radicals are preferentially cleaved. 18 Therefore, MALDI-ISD using an oxidizing matrix is concluded to produce [a]•/[x − H] fragment pair through a nitrogen-centered radical intermediate (Scheme 1A). 2,18,19 In contrast, Takayama and coworkers proposed a C α -C bond cleavage involving a β-carbon-centered peptide radical intermediate as the MALDI-ISD process.…”

“…18 Therefore, MALDI-ISD using an oxidizing matrix is concluded to produce [a]•/[x − H] fragment pair through a nitrogen-centered radical intermediate (Scheme 1A). 2,18,19 In contrast, Takayama and coworkers proposed a C α -C bond cleavage involving a β-carbon-centered peptide radical intermediate as the MALDI-ISD process. 20 The dissociation process of a β-carbon-centered peptide radical is shown in Scheme 1C.…”