PXD000464

PXD000464 is an original dataset announced via ProteomeXchange.

Title	LC-MS data of peptides from p53 isolated by custom immunoaffinity chromatography
Description	We exploited a primary human fibroblast system in which the p53 tumor suppressor protein accumulates to high concentrations due to infection by an adenovirus type 5 mutant that lacks the E1B 55kDa E3 ubiquitin ligase, which targets p53 for degradation, to examine select post-translational modifications (PTMs) present on this transcriptionally inactivated endogenous human p53, as well as on p53 activated in response to etoposide treatment. These forms of p53 were isolated from whole cell lysates by immunoaffinity chromatography using a cocktail of monoclonal antibodies, followed by SDS-PAGE, and peptides derived from these species were subjected to nano-ultra-high performance liquid chromatography-MS and MS/MS analyses on high resolution Orbitrap platforms. Proteomics informatics: Raw LC-MS/MS data were processed into mgf-format peaklist files and searched using Mascot (v. 2.2.7) against the human p53 sequence determined by direct sequencing of the p53 gene from cDNA derived from the specific batch of HFFs used in this study (identical to the consensus sequence for UniProt P04637), employing a window of 8 ppm for the precursor and 1.2 Da for the fragment ion species and allowing for <= 3 missed trypsin and semi-trypsin cleavages, carbamidomethylation of cysteines as a fixed modification, with methionine oxidation, N-terminal protein acetylation, phosphorylation of Ser and Thr, acetylation of Lys, methylation, dimethylation, and trimethylation of Lys and Arg, diglycine modification of Lys (ubiquitinylation), and deamidation of Asn and Gln, iteratively as variable modifications. Aggregate search results for each sample were collated and consolidated using Scaffold (vs. 4.0.7) according to the PeptideProphet and ProteinProphet parsimony algorithms implemented therein, and filtered to the 90% peptide confidence level and to a precursor mass accuracy within 2-3 ppm.
HostingRepository	PRIDE
AnnounceDate	2014-07-28
AnnouncementXML	Submission_2014-07-28_00:48:23.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD000464
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	David Perlman
SpeciesList	scientific name: Cercopithecus aethiops (Green monkey) (Grivet); NCBI TaxID: 9534;
ModificationList	monohydroxylated residue; acetylated residue; dehydrated residue; phosphorylated residue; iodoacetamide derivatized residue; ubiquitination signature dipeptidyl lysine; deaminated residue; monomethylated residue; formylated residue
Instrument	LTQ Orbitrap; instrument model: LTQ Orbitrap

Revision	Datetime	Status	ChangeLog Entry
0	2013-09-13 07:55:23	ID requested
1	2013-10-17 05:51:50	announced
1	2013-10-17 06:17:51	announced
⏵ 2	2014-07-28 00:48:23	announced	Updated project metadata.

Dataset with its publication pending

submitter keyword: p53 Post-translational Modification

David Perlman
contact affiliation	Princeton University Molecular Biology
contact email	PerlmanD@Princeton.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD000464
     1. Label: PRIDE project
     2. Name: LC-MS data of peptides from p53 isolated by custom immunoaffinity chromatography