PXD070779

PXD070779 is an original dataset announced via ProteomeXchange.

Title	Identification of Transglutaminase 2 associated proteins in HUVEC
Description	TG2 is physiologically expressed in HUVEC cells, contributing to the maintenance of cardiovascular homeostasis, but the mechanistic details remain unclear. Identification of TG2-associated proteins can provide a deeper view of TG2's role in endothelial cells. We have applied two approaches: First, a site-specifically N-terminally biotinylated recombinant TG2 was produced in the Rosetta 2 strain, and the Neutravidin-agarose resin was used to isolate TG2-associated proteins from the immortalised HUVEC cell extract. Second, we have developed an endogenous TG2-silenced, N-terminally triple-Flag-tagged transgenic TG2-expressing HUVEC cell line, and intracellularly assembled TG2-associated protein complexes were isolated using anti-Flag agarose resin. To reveal the conformation-dependent interactions of TG2, NC9 (24 hours treatment before lysis) and GTPgammaS (during CoIP) were applied to stabilise open or closed TG2, respectively. (One acidic and a denaturing elution were applied in the case of each sample from the appropriate resin. The eluted sample was run into an SDS-polyacrylamide gel till the top of the separation gel, and the protein-containing part of the gel was cut based on Coomassie (Page-Blue) staining. The proteins in the submitted gel pieces were digested with trypsin overnight (37 C degree). After digestion, the samples were dried to a volume of 0.01 ml. The protein samples were prepared for MS analysis using the Easy nLC1200 (Thermo Scientific) nanouplc-Orbitrap Fusion (Thermo Scientific) MS/MS system. The results were imported into the Scaffold 5.0.1 software. In the Scaffold software, the Exclusive Unique Peptide Count display option and a protein threshold of 1.0% false discovery rate (FDR), along with a peptide threshold of 0.1% FDR, were applied, requiring a minimum of one identified peptide for each protein. The fragmentation table of each detected unique peptide was meticulously checked to verify that the detected peptide is a genuine hit. For this verification, the fragmentation table must include at least four B or Y ion series. The experiments were repeated at least 3 times, and the hits obtained from each repetition were aggregated, after which the control non-specifically bound protein hits were deducted.)
HostingRepository	MassIVE
AnnounceDate	2025-12-22
AnnouncementXML	Submission_2025-12-22_05:50:02.073.xml
DigitalObjectIdentifier
ReviewLevel	Non peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Robert Kiraly
SpeciesList	scientific name: Homo sapiens; common name: human; NCBI TaxID: 9606;
ModificationList	Acetyl; Carbamidomethyl; Oxidation; Deamidated
Instrument	Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2025-11-15 13:29:39	ID requested
⏵ 1	2025-12-22 05:50:02	announced

no publication

submitter keyword: transglutaminase 2, HUVEC, CoIP, protein-protein interaction, conformation dependence, DatasetType:Proteomics

Robert Kiraly
contact affiliation	University of Debrecen
contact email	kiralyr@med.unideb.hu
lab head
Robert Kiraly
contact affiliation	University of Debrecen
contact email	kiralyr@med.unideb.hu
dataset submitter

MassIVE dataset URI

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