Presentation of a homobifunctional azo-reagent for protein structure analysis by collision-induced dissociative chemical cross-linking: Proof-of-principle

摘要: Abstract 4,4′-Azobis[4-cyanopentanoic] acid, a symmetrical free radical initiator, is transformed in the bis-N-succinimidyl-active ester (azoXL) and utilized for peptide cross-linking (XL). The azoXL-reagent reacts with amine functionalities performs upon collision induced dissociation (CID) exclusively loss of N2 leading to formation two sites. (+)ESI-MSn product-ion spectra protonated molecular ion model peptides (MRFA RKDVY) interconnected N-terminally by 4,4′-azobis[4-cyanopentanoic] acid show characteristically mass-shifted product ions neutral losses (N2 = 28 Da; 2-methylacrylonitrile = 67 Da), potentially useful identification XL derivatized peptides. Additionally, CID fragmentation reactions lead sequence-specific fragment ions, allowing primary structure elucidation. presented results demonstrate potential azo-compounds as CID-labile XL-reagents tandem MS analysis. behaviour (MRFA, RKDVY Substance P) azoXL reagent studied MSn, exact mass measurements ion-molecule (IMRs) allyliodide dimethyl disulfide.