Fundamentals of Electron Capture Dissociation

Since its discovery in 1998, electron capture dissociation (ECD) has been broadly applied to the structural characterization of a wide variety of biomolecules, including proteins/peptides, DNAs/RNAs, carbohydrates, lipids, and protein complexes. For proteins and peptides, ECD has the ability to indiscriminately cleave the backbone N-Cα bond, while leaving the more labile side-chain modifications intact. This unique characteristic of ECD was traditionally attributed to its “non-ergodic” nature, a premise that is the subject of an intense debate. Several primary ECD mechanisms have been proposed to explain the preferential formation of c- and z·-type ions, but they cannot account for the formation of other types of fragment ions, particularly those arising from multiple (backbone or side-chain) cleavages. It was proposed that the z·-ion formed upon primary N-Cα bond cleavage can undergo further radical-induced reactions, leading to secondary fragment ion formation, via a process known as the free radical cascade (FRC).

The original FRC paper can be downloaded here:

Leymarie, N.; Costello, C. E.; O’Connor, P. B. Electron capture dissociation initiates a free radical reaction cascade J. Am. Chem. Soc. 2003, 125, 8949-8958 Link.

In addition to secondary cleavages, a radical may also induce hydrogen migration within the nascently formed fragment ion complex, leading to the formation of c·- and z-type ions. These ions have a 1-Da shift from the corresponding c- and z·-type ions, which may lead to erroneous peak assignment or peak intensity calculation, if not properly considered. A series of experiments employing isotope-labeled peptides, peptide with fixed-charge tags or radical trap modifications, MS3, ion activation, and/or resonant ejection of the fragment ion complexes, were performed to investigate the diverse roles of radicals in the peptide ECD process. These findings have led to the development of novel methods for analysis of modified peptides (see also, the isoaspartome project), and have significant implications in automatic analysis of ECD spectra using bioinformatics tools. Some of the tools developed, such as the double resonance (DR)-ECD, have been utilized to study gas-phase protein/peptide folding kinetics and thermodynamics.

The following papers describe these studies in detail:

Sargaeva, N. P.; Lin, C.; O’Connor, P. B. Unusual Fragmentation of β-Linked Peptides by ExD Tandem Mass Spectrometry J. Am. Soc. Mass Spectrom. 2011, 22, 480. Link

Li, X.; Huang, Y.; O’Connor, P. B.; Lin, C. Structural Heterogeneity of Doubly-Charged Peptide b-Ions J. Am. Soc. Mass Spectrom. 2011, 22, 245. Link

Li, X.; Lin, C.; Han, L.; Costello, C. E.; O’Connor, P. B. Charge Remote Fragmentation in Electron Capture and Electron Transfer Dissociations J. Am. Soc. Mass Spectrom. 2010, 21, 646. Link

Li, X.; Cournoyer, J. J.; Lin, C.; O’Connor, P. B. The Effect of Fixed Charge Modifications on Electron Capture Dissociation J. Am. Soc. Mass Spectrom. 2008, 19, 1514. Link

Lin, C.; Cournoyer, J. J.; O’Connor, P. B. Probing the Gas Phase Folding Kinetics of Peptide Ions by IR Activated DR-ECD J. Am. Soc. Mass Spectrom. 2008, 19, 780. Link

Lin, C.; Cournoyer, J. J.; O’Connor, P. B. Use of a Double Resonance Electron Capture Dissociation Experiment to Probe Fragment Intermediate Lifetimes J. Am. Soc. Mass Spectrom., 2006, 17, 1605. Link

Belyayev, M. A.; Cournoyer, J. J.; Lin, C.; O’Connor, P. B. The Effect of Radical Trap Moieties on Electron Capture Dissociation Spectra of Substance P J. Am. Soc. Mass Spectrom. 2006, 17, 1428. Link

O’Connor, P. B.; Lin, C.; Cournoyer, J. J. Long-Lived Electron Capture Dissociation Product Ions Experience Radical Migration via Hydrogen Abstraction J. Am. Soc. Mass Spectrom. 2006, 17, 576. Link

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