Combined use of ion mobility and collision-induced dissociation to investigate the opening of disulfide bridges by electron-transfer dissociation in peptides bearing two disulfide bonds.
Disulfide bonds are post-translational modifications (PTMs) often found in peptides and proteins. They increase their stability toward enzymatic degradations and provide the structure and (consequently) the activity of such folded proteins. The characterization of disulfide patterns, i.e., the cysteine connectivity, is crucial to achieve a global picture of the active conformation of the protein of interest. Electron-transfer dissociation (ETD) constitutes a valuable tool to cleave the disulfide bonds in the gas phase, avoiding chemical reduction/alkylation in solution. To characterize the cysteine pairing, the present work proposes (i) to reduce by ETD one of the two disulfide bridges of model peptides, resulting in the opening of the cyclic structures, (ii) to separate the generated species by ion mobility, and (iii) to characterize the species using collision-induced dissociation (CID). Results of this strategy applied to several peptides show different behaviors depending on the connectivity. The loss of SH· radical species, observed for all the peptides, confirms the cleavage of the disulfides during the ETD process.
†Laboratory of Mass Spectrometry, Department of Chemistry, GIGA-R, University of Liege, Allée de la Chimie 3, B-4000 Liege, Belgium.
Philippe
P
Massonnet
‡Commissariat à l'Energie Atomique, DSV/iBiTec-S/SIMOPRO, F91191 Gif-sur-Yvette, France.
Gregory
G
Upert
†Laboratory of Mass Spectrometry, Department of Chemistry, GIGA-R, University of Liege, Allée de la Chimie 3, B-4000 Liege, Belgium.
Nicolas
N
Smargiasso
‡Commissariat à l'Energie Atomique, DSV/iBiTec-S/SIMOPRO, F91191 Gif-sur-Yvette, France.
Nicolas
N
Gilles
†Laboratory of Mass Spectrometry, Department of Chemistry, GIGA-R, University of Liege, Allée de la Chimie 3, B-4000 Liege, Belgium.
Loïc
L
Quinton
†Laboratory of Mass Spectrometry, Department of Chemistry, GIGA-R, University of Liege, Allée de la Chimie 3, B-4000 Liege, Belgium.
Edwin
E
De Pauw
D000595:Amino Acid Sequence; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry*; D015195:Drug Design; D004579:Electron Transport; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D053719:Tandem Mass Spectrometry
D004220:Disulfides; D010455:Peptides; D003545:Cysteine