Top-Down Characterization of Denatured Proteins and Native Protein Complexes Using Electron Capture Dissociation Implemented within a Modified Ion Mobility-Mass Spectrometer

Electron-based fragmentation methods have revolutionized biomolecular mass spectrometry, in particular native and top-down protein analysis. Here, we report the use of a new electromagnetostatic cell to perform electron capture dissociation (ECD) within a quadrupole/ion mobility/time-of-flight mass...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 2020-03, Vol.92 (5), p.3674-3681
Main Authors: Williams, Jonathan P, Morrison, Lindsay J, Brown, Jeffery M, Beckman, Joseph S, Voinov, Valery G, Lermyte, Frederik
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Beta decay
Cattle
Chemistry
Electron capture
Electron transfer
Electrons
Fragmentation
Hemoglobin
Humans
Ionic mobility
Ions
Mass spectrometry
Mass Spectrometry - methods
Mass spectroscopy
Mobility
Models, Molecular
Peptides
Protein Denaturation
Protein Multimerization
Protein Structure, Quaternary
Proteins
Proteins - chemistry
Proteomics
Quadrupoles
Scientific imaging
Spectroscopy
Vapor phases
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Summary:Electron-based fragmentation methods have revolutionized biomolecular mass spectrometry, in particular native and top-down protein analysis. Here, we report the use of a new electromagnetostatic cell to perform electron capture dissociation (ECD) within a quadrupole/ion mobility/time-of-flight mass spectrometer. This cell was installed between the ion mobility and time-of-flight regions of the instrument, and fragmentation was fast enough to be compatible with mobility separation. The instrument was already fitted with electron transfer dissociation (ETD) between the quadrupole and mobility regions prior to modification. We show excellent fragmentation efficiency for denatured peptides and proteins without the need to trap ions in the gas phase. Additionally, we demonstrate native top-down backbone fragmentation of noncovalent protein complexes, leading to comparable sequence coverage to what was achieved using the instrument’s existing ETD capabilities. Limited collisional ion activation of the hemoglobin tetramer before ECD was reflected in the observed fragmentation pattern, and complementary ion mobility measurements prior to ECD provided orthogonal evidence of monomer unfolding within this complex. The approach demonstrated here provides a powerful platform for both top-down proteomics and mass spectrometry-based structural biology studies.
ISSN:0003-2700
1520-6882
DOI:10.1021/acs.analchem.9b04763