PXD061766

PXD061766 is an original dataset announced via ProteomeXchange.

Title	cross linking mass spectrometry of Exo84 pulldown in Sec9 and Sec18 depletion mutants
Description	Co-Immunoprecipitation with Crosslinking Using DSS for Exo84 Subunit Sample Preparation: Yeast strains expressing Exo84-3xGFP and FRB-tagged Sec18 or Sec9 were cultured in YPD at 30°C until OD600 0.8-1. Cells were harvested, washed with resuspension buffer (50 mM HEPES, 150 mM NaCl, protease inhibitors), and frozen in liquid nitrogen. Lysis was performed using a Freezer/Mill, followed by clarification via centrifugation. Crosslinking with DSS: Lysates were crosslinked with DSS (0.5 mM, 1h, RT) and quenched with glycine (100 mM, 15 min). Supernatants were clarified by centrifugation. Immunoprecipitation: GFP-Trap beads were used to capture Exo84 complexes. Lysates were pre-cleared with blocking beads (1h, 4°C) before incubation with GFP-Trap beads (overnight, 4°C). Beads were washed with a stringent buffer (2M NaCl, 8M urea, 1% Triton X-100) and PBS. Mass Spectrometry Sample Preparation: Immunoprecipitated complexes were reduced (DTT), alkylated (IAA), digested with trypsin (37°C, 8h), and desalted before LC-MS/MS. LC-MS/MS Analysis: Samples were analyzed on an Orbitrap Fusion Lumos coupled to an EASY-nLC 1200 with a 50 cm C18 column. Peptides were separated with a gradient from 5% to 40% acetonitrile over 60 min. MS1 scans were acquired at 120,000 resolution, and the Top Speed method was used for data-dependent MS2 acquisition with HCD fragmentation.
HostingRepository	PRIDE
AnnounceDate	2026-01-21
AnnouncementXML	Submission_2026-01-21_05:15:48.353.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Oriol Gallego
SpeciesList	scientific name: Saccharomyces cerevisiae (Baker's yeast); NCBI TaxID: NEWT:4932; scientific name: Saccharomyces cerevisiae; NCBI TaxID: NCBITaxon:4932;
ModificationList	acetylated residue; monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2025-03-12 10:52:37	ID requested
⏵ 1	2026-01-21 05:15:48	announced

10.1016/J.CELL.2025.11.038;

submitter keyword: Crosslinking,saccharomyces cerevisiae, LC-MS/MS

Oriol Gallego Moli
contact affiliation	Department of Medicine and Life Sciences, Biophysics in Cell Biology Group, Universitat Pompeu Fabra, Spain
contact email	oriol.gallego@upf.edu
lab head
Oriol Gallego
contact affiliation	Principal Investigator & Professor at the Universitat Pompeu Fabra
contact email	gallegolab@upf.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD061766
     1. Label: PRIDE project
     2. Name: cross linking mass spectrometry of Exo84 pulldown in Sec9 and Sec18 depletion mutants