PXD038295

PXD038295 is an original dataset announced via ProteomeXchange.

Title	Pyrophosphorylation sites of Flag tagged IRF3 in IRF3 KO 293T cells infected by SeV
Description	Total lysates were prepared from IRF3 KO 293T cells transfected with Flag-IRF3 and followed by immunoprecipitation with α-FLAG M2 beads. Immunoprecipitated proteins were separated by SDS-PAGE, and then stained with Coomassie Blue. The entire lane was excised, digested with trypsin, and analyzed with LC-MS/MS. LC-MS/MS identification of peptide mixtures was performed once with two replicates per condition at BIOPROFILE TECHNOLOGY (Shanghai, China). Briefly, peptides were chromatographed through Easy-nLC1200 (Thermo Fisher, California, USA). Peptide samples were loaded by Trap column (reverse-phase), (100 μm*2 cm, 5μm, C18, Dr. Maisch GmbH) and separated by Thermo scientific EASY column (75 μm*15cm, 3 μm, C18) at 300 nL min−1 for 60 min using a four-step acetonitrile (0.1% formic acid in 80% acetonitrile) gradient: 2–5% over the first 2 min and 5–28% for 2–44 min and 28–40% for 44–51 min and 40–100% for 51–53 min. The tandem mass spectrometry was performed by Q Exactive mass spectrometer (Thermo Fisher, California, USA). The MS1 survey scan (350–1800 m/z) was at a resolution of 60,000 at 200 m/z with automatic gain control (AGC) target of 3e6 and a maximum injection time of 50 ms. Dynamic exclusion was 60.0 s. Each full scan takes 20 MS2 scans. MS2 activation type was HCD model. Isolation window was 2 m/z. The MS2 survey was at a resolution of 15,000 at 200 m/z with automatic gain control (AGC) target of 1e5 and a maximum injection time of 50 ms. RAW files generated by spectrometer was subjected to Proteome Discoverer (version 1.4) software with searching the library of Uniprot homo sapiens (uniprot-Homo sapiens (Human) [9606]-204995-20220606.fasta) for protein identification. Trypsin was specified as the proteolytic enzyme, with up to two missed cleavage sites allowed. Carbamidomethylation was set as the fixed modification. Oxidation (M) and ubiquitination [GlyGly (K)] were set as the variable modifications. The precursor mass tolerance was set to 20 ppm and the fragment ion tolerance at 0.1 Da. Proteins were identified based on at least one unique peptide. Protein and peptide identification confidence threshold were set to an FDR of 1%. The tandem mass spectra of matched peptides were checked manually for their validity.
HostingRepository	iProX
AnnounceDate	2022-11-22
AnnouncementXML	Submission_2022-11-22_19:15:09.878.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Shuai Yang
SpeciesList	scientific name: Homo sapiens; NCBI TaxID: 9606;
ModificationList	phosphorylated residue
Instrument	Q Exactive Plus; Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2022-11-22 19:14:10	ID requested
⏵ 1	2022-11-22 19:15:10	announced

Dataset with its publication pending

submitter keyword: IRF3, pyrophosphorylation

Jun Cui
contact affiliation	Sun Yat-sen University
contact email	cuij5@mail.sysu.edu.cn
lab head
Shuai Yang
contact affiliation	Sun Yat-sen University
contact email	yangsh225@mail.sysu.edu.cn
dataset submitter

iProX dataset URI