PXD057272

PXD057272 is an original dataset announced via ProteomeXchange.

Title	C-terminal glycosylation of type IX secretion system cargo proteins in Prevotella intermedia bearing both short and long secretion signals
Description	Prevotella intermedia is a Gram-negative bacterium that is notably linked to periodontitis and acute necrotizing ulcerative gingivitis. P. intermedia is known to utilise the T9SS to secrete and anchor virulence factors to the cell surface, presumably via C-terminal modification of the cargo protein with a cell surface polysaccharide. The identity of the linking sugar and the sites of modification on the cargo have yet to be determined. Here, we first employed Hidden Markov Models to predict cargo proteins in P. intermedia, and then conducted LC-MS/MS analyses of partially deglycosylated fractions to characterise the C-terminal glycosylation. A total of 80 cargo proteins were predicted based on the presence of a T9SS C-terminal domain signal (CTD), and these were divided into 48 short CTDs predicted by AlphaFold to adopt a single 3-stranded β-sheet and 32 long CTDs predicted to form the more typical larger β-sandwich structures. Cleavage sites for five short CTDs and four long CTDs were experimentally determined, and glycosylation was observed at the mature C-terminus of six cargo. Two glycans were identified of delta masses 419.198 and 433.185 Da, corresponding to novel linkages to N-alanyl dHex-HexNAc and N-alanyl (Me-dHex)-HexNAc, respectively, with the alanine amide-linked to the protein C-terminus. This indicated that both short and long CTDs were cleaved and glycosylated. AlphaFold multimer modelling predicted that both kinds of CTDs could bind to the PorV shuttle protein in the same manner, with the conserved CTD motifs interacting with the same sites in PorV.
HostingRepository	PRIDE
AnnounceDate	2025-05-07
AnnouncementXML	Submission_2025-05-07_04:04:19.765.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD057272
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Paul Veith
SpeciesList	scientific name: Prevotella intermedia ATCC 25611 = DSM 20706; NCBI TaxID: 1122984;
ModificationList	monohydroxylated residue; complex glycosylation; iodoacetamide derivatized residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2024-10-28 20:57:06	ID requested
⏵ 1	2025-05-07 04:04:20	announced

Ye X, Bin Rustam N, Gorasia D, Reynolds E, Ghosal D, Veith P, with both short and long secretion signals. Open Biol, 15(3):240335(2025) [pubmed]

10.6019/PXD057272;

10.1098/rsob.240335;

submitter keyword: glycosylation, secretion signal, surface proteins,Prevotella intermedia, alaninamide, mass spectrometry, protein secretion

Eric Charles Reynolds
contact affiliation	Bio21 Institute, Melbourne Dental School, University of Melbourne
contact email	ecr@unimelb.edu.au
lab head
Paul Veith
contact affiliation	Melbourne Dental School
contact email	pdv@unimelb.edu.au
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD057272
     1. Label: PRIDE project
     2. Name: C-terminal glycosylation of type IX secretion system cargo proteins in Prevotella intermedia bearing both short and long secretion signals