PXD009227

PXD009227 is an original dataset announced via ProteomeXchange.

Title	High-throughput and high-sensitivity phosphoproteomics with the EasyPhos platform
Description	Mass spectrometry has transformed the field of cell signalling by enabling global studies of dynamic protein phosphorylation (‘phosphoproteomics’). Recent developments are enabling increasingly sophisticated phosphoproteomics studies, but practical challenges remain. The EasyPhos workflow addresses these, and is sufficiently streamlined to enable analysis of hundreds of phosphoproteomes at a depth of >10,000 quantified phosphorylation sites. Here we present a detailed and updated protocol that further ensures high performance in sample-limited conditions, while also reducing sample preparation time. By eliminating protein precipitation steps and performing the entire protocol including digestion in a single 96-well plate, we now greatly minimize opportunities for sample loss and variability. This results in very high reproducibility and low sample requirements of 200 μg of protein starting material or less. After cell culture or tissue collection, the protocol takes 1 d, whereas mass spectrometry measurements require 1 h per sample. Applied to glioblastoma cells acutely treated with EGF, EasyPhos quantified 20,132 distinct phosphopeptides from 200 μg protein in less than one day of measurement time, revealing thousands of EGF-regulated phosphorylation events.
HostingRepository	PRIDE
AnnounceDate	2018-09-07
AnnouncementXML	Submission_2018-09-12_23:58:15.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Mario Oroshi
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	phosphorylated residue
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2018-03-16 00:52:55	ID requested
1	2018-09-07 02:21:35	announced
⏵ 2	2018-09-12 23:58:16	announced	Updated publication reference for PubMed record(s): 30190555.

Humphrey SJ, Karayel O, James DE, Mann M, High-throughput and high-sensitivity phosphoproteomics with the EasyPhos platform. Nat Protoc, 13(9):1897-1916(2018) [pubmed]

curator keyword: Technical

submitter keyword: mass spectrometry, proteomics, phosphoproteomics, phosphorylation, signal transduction, post-translational modification, EGF, cancer, glioblastoma

Matthias Mann
contact affiliation	Department of Proteomics and Signal Transduction Max Planck Institute of Biochemistry
contact email	mmann@biochem.mpg.de
lab head
Mario Oroshi
contact affiliation	Proteomics
contact email	oroshi@biochem.mpg.de
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2018/09/PXD009227

PRIDE project URI

• PRIDE
  1. PXD009227
     1. Label: PRIDE project
     2. Name: High-throughput and high-sensitivity phosphoproteomics with the EasyPhos platform