Anionic copper complex fragmentations from enkephalins under low‐energy collision‐induced dissociation in an ion trap mass spectrometer

Abstract

Peptide metallation with Cu2+ was explored in the negative ESI mode using an ion trap mass spectrometer. Under these conditions, the [(M-3H) + CuII]- species formed were investigated under low-energy collision-induced dissociation conditions. MS2 experiments indicate a very different behavior of CuII metallated complexes compared with [M-H]- species. CuII induces an easy loss of CO2 and specific side-chain cleavages (by radical losses) at the C-terminal residue, as observed previously by prompt 'in source' dissociation experiments. The loss of CO2 yields an unstable carbylide that leads to further dissociations involving the migration of a proton or a hydrogen radical (through the reduction of CuII). Multistage MS3 experiments were carried out to rationalize this behavior. Fragmentation pathways are proposed in order to explain the product ions observed. The side-chain radical loss at the C-terminus was demonstrated to be a consecutive process. Finally, evidence is provided that the specific side-chain cleavages can be used for the differentiation of Leu/Ile and Gln/Lys residues when they are located at the C-terminus. The existence of a zwitterionic form in the case of the anionic YGGFK-CuII complex is proposed.

Negative‐charge driven fragmentations for evidencing zwitterionic forms from doubly charged coppered peptides

“…However, the absence of a CO 2 loss, indicating the absence of a free carboxylate group, (Kjeldsen F, Silivra OA, Zubarev RA, Zwitterionic States in Gas‐Phase Poly‐Peptide ions Revealed by 157 nm Ultra‐Violet Photo‐Dissociation , Personal Communication. )38–40, 68, 69 suggests the attribution of the negative charge to the phenol group and/or to one of the amide groups rather than to the C ‐terminal carboxylic acid. The protonation of the C ‐terminal carboxylic acid was also confirmed by the observation of a formic acid loss during the MS 3 analysis of the product m / z 300.6 ion (data not reported).…”

“…The studied enkephalin analogs (YGGFL, YGGFL(NH 2 ), YGGFK, YGGFQ, KYGGF and QYGGF) were synthesized as described elsewhere39 and dissolved in deionized water at 1 mg.ml −1 . A solution of CuCl 2 .6H 2 O (Sigma, St. Quentin Fallavier, France) containing 500 pmol.µl −1 was also prepared in deionized water.…”

“…For peptides without a coordinating side‐chain, the electron donor atoms, which preferentially coordinate Cu II , are thus the amide groups of the peptide backbone and the N ‐terminal amine. Moreover, the metallation significantly increases the acidity of these nitrogen‐containing electron donors and thus enhances their deprotonation 35–42…”

“…The dissociation of ternary Cu II − 2, 2′‐bipyridyl (bpy) complexes [(AA‐H)bpy + Cu II ] + was used to distinguish and quantify the isomeric I/L and the isobaric K/Q AA 43. Moreover, the analysis of the [(M − 3H) + Cu II ] − ions of pentapeptides in the negative ESI mode was used to differentiate the isomeric L/I38–40 and the isobaric K/Q38, 40 residues when they are located at the C ‐terminus. Binary singly charged copper complexes [(M − H) + Cu II ] + were also analyzed using the ESI positive mode to differentiate diastereomeric peptides 37.…”

Formation and Fragmentation of Unsaturated Fatty Acid [M – 2H + Na]– Ions: Stabilized Carbanions for Charge-Directed Fragmentation

“…Unlike fatty acids, however, anionic peptide-metal ion complexes have also been investigated with alkali [22,23], alkaline earth [24][25][26][27] and transition metals [28][29][30].…”