PXD070486

PXD070486 is an original dataset announced via ProteomeXchange.

Title	Extensive backbone cleavage coverage of intact proteoforms in a mass range of 10-70 kDa by integrating electron, collision, and photon-based fragmentation techniques during an electrophoretic timescale
Description	Capillary zone electrophoresis (CZE) –-tandem mass spectrometry (MS/MS) has been documented as a useful tool for top-down proteomics (TDP). However, CZE-MS/MS-based TDP typically has limited backbone cleavage coverage for identified proteoforms due to the use of traditional collision-based fragmentation methods (i.e., higher-energy collisional dissociation, HCD). Here, for the first time, we coupled CZE to a high-end Orbitrap Ascend mass spectrometer to investigate the performance of collision, electron, and photon-based fragmentation methods and their combinations for boosting the backbone cleavage coverage of proteoforms during the electrophoretic timescale using a standard protein mixture covering a mass range of about 10-70 kDa. CZE-MS achieved reproducible measurement of six proteins, including three insulin-like growth factor (IGF) proteoforms with different modifications. Systematic investigations of HCD, electron-transfer dissociation (ETD), electron-transfer/HCD (EThcD), and ultraviolet photodissociation (UVPD) during CZE-MS/MS analysis revealed distinct yet complementary fragmentation characteristics. ETD, EThcD, and UVPD, in general, provided higher backbone cleavage coverage than HCD. The integration of HCD, ETD, EThcD, and UVPD data offered a 74% and 98% sequence coverage for carbonic anhydrase (a 30-kDa protein) and thioredoxin (a 12-kDa protein), which is 185% and 100% higher than that produced by HCD alone. Adding internal fragments further boosted the backbone cleavage coverage substantially, for example, from 74% to 94% for 30-kDa carbonic anhydrase, and from 35% to 94% for 50-kDa Protein AG. The results demonstrate the capability of CZE-MS/MS with the integration of various fragmentation techniques for comprehensive characterization of proteoforms in a wide mass range.
HostingRepository	PRIDE
AnnounceDate	2026-02-16
AnnouncementXML	Submission_2026-02-15_16:26:21.915.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Qianjie Wang
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	deamidated residue
Instrument	Orbitrap Ascend

Revision	Datetime	Status	ChangeLog Entry
0	2025-11-08 01:20:52	ID requested
⏵ 1	2026-02-15 16:26:22	announced

Wang Q, Wang Q, Melani RD, Liu Q, Nurmi P, Sun L, Extensive Backbone Cleavage Coverage of Intact Proteoforms in a Mass Range of 10-70 kDa by Integrating Electron, Collision, and Photon-Based Fragmentation Techniques during an Electrophoretic Time Scale. J Am Soc Mass Spectrom, 37(2):505-513(2026) [pubmed]

10.1021/jasms.5c00384;

submitter keyword: Capillary zone electrophoresis mass spectrometry

Top-down proteomics

Orbitrap Ascend Tribrid

ultraviolet photodissociation

higher-energy collisional dissociation

electron transfer dissociation

Qianjie Wang
contact affiliation	Michigan State University
contact email	wangqi50@msu.edu
lab head
Qianjie Wang
contact affiliation	Michigan State University
contact email	wangqi50@msu.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD070486
     1. Label: PRIDE project
     2. Name: Extensive backbone cleavage coverage of intact proteoforms in a mass range of 10-70 kDa by integrating electron, collision, and photon-based fragmentation techniques during an electrophoretic timescale