posted on 2021-06-03, 14:36authored byTakashi Baba, Pavel Ryumin, Eva Duchoslav, Keqin Chen, Anjali Chelur, Bill Loyd, Igor Chernushevich
We
report the progress
on an electron-activated dissociation (EAD)
device coupled to a quadrupole TOF mass spectrometer (QqTOF MS) developed
in our group. This device features a new electron beam optics design
allowing up to 100 times stronger electron currents in the reaction
cell. The electron beam current reached the space-charge limit of
0.5 μA at near-zero electron kinetic energies. These advances
enable fast and efficient dissociation of various analytes ranging
from singly charged small molecules to multiply protonated proteins.
Tunable electron energy provides access to different fragmentation
regimes: ECD, hot ECD, and electron-impact excitation of ions from
organics (EIEIO). The efficiency of the device was tested on a wide
range of precursor charge states. The EAD device was installed in
a QqTOF MS employing a novel trap-and-release strategy facilitating
spatial mass focusing of ions at the center of the TOF accelerator.
This technique increased the sensitivity 6–10 times and allows
for the first time comprehensive structural lipidomics on an LC time
scale. The system was evaluated for other compound classes such as
intact proteins and glycopeptides. Application of hot ECD for the
analysis of glycopeptides resulted in rich fragmentation with predominantly
peptide backbone fragments; however, glycan fragments attributed to
the ECD process were also observed. A standard small protein ubiquitin
(8.6 kDa) was sequenced with 90% cleavage coverage at spectrum accumulation
times of 100 ms and 98% at 800 ms. Comparable cleavage coverage for
a medium-size protein (carbonic anhydrase: 29 kDa) could be achieved,
albeit with longer accumulation times.