PXD063177-1

PXD063177 is an original dataset announced via ProteomeXchange.

Title	Enhancing glycopeptide annotation and glycan localization using the EAD implementation of hot electron capture dissociation through a multiplexed parallel reaction monitoring design of experiment
Description	We present an optimized electron activated dissociation (EAD) methodology for liquid chromatography-tandem mass spectrometry (LC-MS/MS) based characterization of N- and O-glycopeptides. Recombinant human erythropoietin (rhEPO) was used as a model glycoprotein in this study. Applying a full factorial design of experiment (DoE) approach on the ZenoTOF 7600 instrument, we first optimized LC-MS parameters (i.e., ion spray voltage, ion source temperature, and active gradient time) to enhance glycopeptide ionization efficiency while reducing in-source fragmentation (ISF). Then, another DoE was performed for EAD parameter optimization. Multiplexed parallel reaction monitoring of one glycoform of each glycopeptide was performed to efficiently and comprehensively optimize the electron beam current, reaction time, and electron kinetic energy of the EAD set-up. Finally, the optimized EAD parameters were successfully applied in data-dependent acquisition (DDA) mode for the untargeted analysis of tryptic digests of rhEPO. Byonic and Mascot softwares were used to evaluate the potential of our optimized EAD setup against collision induced dissociation (CID), confirming that with EAD we improved glycan localization confidence while with CID a superior number of glycoforms were identified although with less confident glycan assignment.
HostingRepository	PRIDE
AnnounceDate	2025-10-23
AnnouncementXML	Submission_2025-10-23_03:47:59.408.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Maarten Dhaenens
SpeciesList	scientific name: Cricetulus griseus (Chinese hamster) (Cricetulus barabensis griseus); NCBI TaxID: NEWT:10029;
ModificationList	N-glycosylated residue; O-glycosylated residue; iodoacetamide derivatized residue
Instrument	ZenoTOF 7600

Revision	Datetime	Status	ChangeLog Entry
0	2025-04-22 00:54:29	ID requested
⏵ 1	2025-10-23 03:47:59	announced

10.1016/J.JPROT.2025.105546;

submitter keyword: Glycoproteomics, Parallel Reaction Monitoring, Electron Activated Dissociation, Design of Experiments, LC-MS/MS

Maarten Dhaenens
contact affiliation	ProGenTomics, Laboratory of Pharmaceutical Biotechnology, Faculty of Pharmaceutical Sciences, Ghent University, 9000 Ghent, Belgium
contact email	maarten.dhaenens@ugent.be
lab head
Maarten Dhaenens
contact affiliation	Faculity of Pharmaceutical Biotechnology
contact email	maarten.dhaenens@ugent.be
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD063177
     1. Label: PRIDE project
     2. Name: Enhancing glycopeptide annotation and glycan localization using the EAD implementation of hot electron capture dissociation through a multiplexed parallel reaction monitoring design of experiment