PXD000459-1

PXD000459 is an original dataset announced via ProteomeXchange.

Title	Identification of SUMO sites by LC-MSMS
Description	We developed a novel method for the identification of SUMO sites by expression of His-tagged SUMO mutants and affinity purification of SUMOylated proteins, followed by trypsin digestion and immunocapture of peptides containing diglycine signature tags. SILAC labeling allowed to distinguish peptides reporting true SUMO sites from ubiquitin, NEDD8 or ISG15 sites. We identified a total of 331 SUMO1 sites of which the large majority was downregulated upon treatment with a bacterial toxin, listeriolysin O. LC-MS/MS analyses were performed on a Thermo LTQ Orbitrap Velos mass spectrometer operated in standard data-dependent mode. CID fragmentation spectra were recorded in the LTQ analyzer and used to create Mascot generic files (mgf) by the Mascot Distiller software (version 2.4.3.3). Generated peak lists were then searched with Mascot (version 2.3.0) against the human proteins in the Swiss-Prot database (database release version of March 6 2013). Three independent searches were performed to identify light, medium and heavy labeled SILAC peptides. For all three searches variable modifications were set to oxidation of methionine residues and pyroglutamate formation of N-terminal glutamine residues. Carbamidomethyl formation of cysteine residues was set as fixed modification. Depending on the search, additional variable modifications included di-glycine modification of lysine residues (+114.042927, light search), SILAC modification of lysine residues and di-glycine modification of SILAC labeled lysine residues (+4.025107 and +118.068034, medium search; +8.014199 and +122.057126, heavy search). SILAC modification of arginine residues was set as additional fixed modification for the medium (+6.020129) and heavy (+10.008269) search. For all searches mass tolerance of the precursor ions was set to 10 ppm and mass tolerance of the fragment ions was set to 0.5 Da. The peptide charge was set to 2+, 3+ and 4+, and up to three missed tryptic cleavage sites were allowed. Only peptides that were ranked first and scored above the threshold score set at 99% confidence were withheld. For processing of all MS data the ms_lims software platform was used (version 7.7.7).
HostingRepository	PRIDE
AnnounceDate	2014-07-29
AnnouncementXML	Submission_2014-07-29_02:59:24.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Impens Francis
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	monohydroxylated residue; iodoacetamide derivatized residue; ubiquitination signature dipeptidyl lysine; 2-pyrrolidone-5-carboxylic acid (Gln)
Instrument	LTQ Orbitrap Velos

Revision	Datetime	Status	ChangeLog Entry
0	2013-09-11 06:03:51	ID requested
⏵ 1	2014-07-29 02:59:25	announced
2	2014-07-30 01:14:54	announced	Updated project metadata.
3	2014-09-10 07:35:04	announced	Updated project metadata.
4	2024-10-22 04:21:16	announced	2024-10-22: Updated project metadata.

Dataset with its publication pending

curator keyword: Biomedical

submitter keyword: SUMO, HeLa cells, LC-MSMS, SILAC, immunocapture

Impens Francis
contact affiliation	Ghent University
contact email	francis.impens@vib-ugent.be
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD000459
     1. Label: PRIDE project
     2. Name: Identification of SUMO sites by LC-MSMS