PXD055856

PXD055856 is an original dataset announced via ProteomeXchange.

Title	PNGaseF-generated N-glycans adduct onto peptides in the gas phase
Description	Glycoproteomics has recently increased in popularity due to instrumental and methodological advances. That said, O-glycoproteomic analysis is still challenging for various reasons, including signal suppression and splitting, search algorithm limitations, and co-occupancy of N- and O-glycopeptides. To decrease sample complexity and simplify analysis, most O-glycoproteomic workflows include PNGaseF digestion, which is an endoglycosidase that removes mammalian N-glycans. Here, we report that N-glycans released from PNGaseF digestion, also referred to as ‘free glycans’, were identified during data acquisition and hampered detection of O-glycopeptides. Importantly, we noted instances where free glycans adducted to unmodified peptides in the gas phase and were misidentified by search algorithms as O-glycopeptides. We confirmed the presence of free glycans in other experiments performed in our laboratory, as well as from data generated by other groups. To overcome this limitation in common O-glycoproteomic workflows, we performed PNGaseF digestion following reduction and alkylation. Here, we demonstrated that released N-glycans can be removed using a molecular weight cut off (MWCO) filter, prior to (glyco)protease digestion, which improved O-glycoproteomic coverage.
HostingRepository	PRIDE
AnnounceDate	2026-06-29
AnnouncementXML	Submission_2026-06-28_16:19:46.862.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Valentina Rangel-Angarita
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	O-glycosylated residue
Instrument	Orbitrap Eclipse

Revision	Datetime	Status	ChangeLog Entry
0	2024-09-13 12:26:23	ID requested
⏵ 1	2026-06-28 16:19:47	announced

10.1021/jasms.4c00431;

Rangel-Angarita V, Chongsaritsinsuk J, Mahoney KE, Kim LM, Chen RJ, Appah-Sampong AA, Tran IP, Steigmeyer AD, Hollenhorst MA, Malaker SA, PNGaseF-Generated N-Glycans Adduct onto Peptides in the Gas Phase. J Am Soc Mass Spectrom, 36(3):542-552(2025) [pubmed]

submitter keyword: adducts,Glycoproteomics, mucinomics, PNGaseF, mass spectrometry, N-glycans

Stacy Malaker
contact affiliation	Department of Chemistry, Yale University, New Haven, CT
contact email	stacy.malaker@yale.edu
lab head
Valentina Rangel-Angarita
contact affiliation	Yale University Department of Chemistry
contact email	valentina.rangelangarita@yale.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD055856
     1. Label: PRIDE project
     2. Name: PNGaseF-generated N-glycans adduct onto peptides in the gas phase