含芳香氨基酸三肽及其衍生物自由基离子的形成和解离机理研究

Structure and gas-phase fragmentation of tyrosyl-glycyl-tryptophan [YGW]^•+ and its derivative [Ac-YGW-OMe]^•+ have been studied using electrospray tandem mass spectrometry (ESI-MS/MS) combined with density functional theory (DFT). These peptide radical ions were generated via multistage collision-induced dissociation (CID) of transition metal-ligand-peptide tertiary complexes, [Cu(L)M]²⁺ (L=4'-chloro-2,2':6',2"-terpyridine (4Cl-tpy); M=YGW or Ac-YGW-OMe). Low-energy CID experiments revealed that the fragmentation mechanisms of [YGW]^•+ and [Ac-YGW-OMe]^•+ are dramatically different. [YGW]^•+ dissociate mainly through the loss of CO₂ and subsequent loss of indole radical to produce [M-CO₂]^•+(m/z 380.05) and [M-CO₂-116]⁺ (m/z 264.05) fragment ions. Other minor fragments include [c₂+2H]⁺ (m/z 238.04), [G^•GW]⁺ (m/z 318.05) and [1H-indole]^•+ (m/z 117.23). In contrast, the main fragmentation of [Ac-YGW-OMe]^•+ is the loss of CH₃OH to give rise to [M-CH₃OH]^•+ (m/z 448.14) product ion. Other minor fragments are [z₁-H]^•+ (m/z 200.93), [c₂+2H]⁺ (m/z 279.99) and [M-CH₃COO•]⁺ (m/z 421.16). The gas-phase fragmentation mechanisms of [YGW]^•+ and [Ac-YGW-OMe]^•+ are proposed based on the aforementioned CID results. For [YGW]^•+, the major fragment ions [M-CO₂]^•+ (m/z 380.05) and [M-CO₂-116]⁺ (m/z 264.05) are generated through proton transfer from the carboxylic OH group to the amide oxygen to form carboxyl radical which undergoes cleavages of C_α-C and C_β-C_γ bonds, respectively. [G^•GW]⁺ (m/z 318.05) fragment ion is formed by C_α-C_β bond cleavage with the loss of p-quinomethide, and [c₂+2H]⁺ (m/z 238.04) fragment ion is produced by proton transfer from the C_β-H of tryptophan to the amide oxygen to produce the β radical followed by N-C_α bond cleavage. For [Ac-YGW-OMe]^•+, the main product ion [M-CH₃OH]^•+ (m/z 448.14) is generated through proton transfer from three possible sources (C_α-H from glycine, C_α-H from tryptophan, C_β-H from side chain of tryptophan) to methyl ester oxygen followed by C-O bond cleavage, [z₁-H]^•+ (m/z 200.93) fragment ion is produced through proton transfer from the C_β-H of tryptophan to the amide oxygen to form β radical which undergoes N-C_α bond cleavage. The DFT calculation results suggest that the optimized geometries of [YGW]^•+ and [Ac-YGW-OMe]^•+ are non-zwitterionic structures with a weak hydrogen bond between the amino nitrogen and the indole nitrogen ([N₁-H⁺...N₄] (bond length ca. 2.043 Å) in [YGW]^•+ and a strong hydrogen bond between the acetyl oxygen and the indole nitrogen ([N₄-H⁺...O₁] (bond length ca.1.666 Å) in [Ac-YGW-OMe]^•+, respectively.