PXD028150-1

PXD028150 is an original dataset announced via ProteomeXchange.

Title	Interrogating kinase-substrate relationships with proximity labeling and phosphorylation enrichment
Description	Kinases govern many cellular responses through the reversible transfer of a phosphate moiety to their substrates. However, pairing a substrate with a kinase is challenging. In proximity labeling experiments, proteins proximal to a target protein are marked by biotinylation, and mass spectrometry can be used for their identification. Here, we combine ascorbate peroxidase (APEX) proximity labeling and a phosphorylation enrichment-based workflow, Phospho-APEX (pAPEX), to rapidly identify phosphorylated and biotinylated neighbor proteins which can be considered for candidate substrates. The pAPEX strategy enriches and quantifies differences in proximity for proteins and phosphorylation sites proximal to an APEX2-tagged kinase under the kinase “ON” and kinase “OFF” conditions. As a proof of concept, we identified candidate substrates of MAPK1 in HEK293T and HCT116 cells and candidate substrates of PKA in HEK293T cells. In addition to many known substrates, C15orf39 was identified and confirmed as a novel MAPK1 substrate. In all, we adapted the proximity labeling-based platform to accommodate phosphorylation analysis for kinase substrate identification. TMT1: pAPEX-MAPK1 experiment in HEK293T cells TMT2: pAPEX-MAPK1 experiment in HCT116 cells TMT3: pAPEX-PKA experiment in HEK293T cells
HostingRepository	PRIDE
AnnounceDate	2022-03-11
AnnouncementXML	Submission_2022-03-11_11:09:35.453.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Tian Zhang
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	phosphorylated residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2021-08-26 04:06:39	ID requested
⏵ 1	2022-03-11 11:09:35	announced
2	2023-11-14 08:54:18	announced	2023-11-14: Updated project metadata.

10.1021/ACS.JPROTEOME.1C00865;

submitter keyword: Proximity labeling, APEX, Kinases, substrates, MAPK1, PKA

Steven Gygi
contact affiliation	Cell Biology Department, Harvard Medical School
contact email	steven_gygi@hms.harvard.edu
lab head
Tian Zhang
contact affiliation	Harvard Medical School
contact email	tian_zhang@hms.harvard.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD028150
     1. Label: PRIDE project
     2. Name: Interrogating kinase-substrate relationships with proximity labeling and phosphorylation enrichment