PXD038473

PXD038473 is an original dataset announced via ProteomeXchange.

Title	In-vivo crosslinking based affinity purification-mass spectrometry for targeting intracellular protein-protein interactions
Description	Comprehensive interactome analysis of targeted proteins is of great significance to understand how pro-teins work together for functional regulation. Until now, affinity purification strategy with mass spec-trometry (AP-MS) has been recognized as the most widely used technique for studying protein-protein interactions (PPIs). However, some proteins with weak interactions, responsible for key roles in pathway regulation, are easily broken during cell lysis by AP approach. To advance the identification depth of protein interactome, herein we developed an approach named in vivo crosslinking based affinity purifi-cation-mass spectrometry (ICAP). By this method, in vivo crosslinking was introduced for covalently fixing the intracellular PPIs at their functional states to assure all the PPIs could be integrally maintained during cell disruption. Contributed by this advantage, the transient/weak PPIs missed in the AP-MS method could be captured in the consecutive biotin-streptavidin purification. With ICAP approach, the interactome of ERK1 from 293A cells was profiled with 6.15-fold improvement in identification number than those by conventional AP-MS. Contributed by the superior performance, ICAP was further extend-ed to the temporal profiling of ERK1 interacting proteins under activation by cAMP-mediated pathway. The reasonable regulatory manner of MAPK pathways was explored by depicting the quantitative changes of ERK1 and its interacting proteins at different time points after activation. Furthermore, owing to chemical cleavable crosslinkers-DSEP, we can choose to perform CXMS analysis for identification of specific protein-protein interactions and sites within interacting proteins in the network. Therefore, all these results demonstrated that our approach can provide comprehensive information on interactome of target protein for molecular biologists in a variety of diverse applications.
HostingRepository	iProX
AnnounceDate	2022-12-01
AnnouncementXML	Submission_2023-12-05_23:41:16.613.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Bowen Zhong
SpeciesList	scientific name: Homo sapiens; NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2022-12-01 01:20:42	ID requested
1	2022-12-01 01:22:33	announced
⏵ 2	2023-12-05 23:41:17	announced	2023-12-06: Update information.

Zhong B, An Y, Gao H, Zhao L, Li X, Liang Z, Zhang Y, Zhao Q, Zhang L, In vivo cross-linking-based affinity purification and mass spectrometry for targeting intracellular protein-protein interactions. Anal Chim Acta, 1265():341273(2023) [pubmed]

submitter keyword: in vivo cross-linking, affinity purification, DSEP, ERK1

Lihua Zhang
contact affiliation	Dalian Institute of Chemical Physics, Chinese Academy of Science
contact email	lihuazhang@dicp.ac.cn
lab head
Bowen Zhong
contact affiliation	Dalian Institute of Chemical Physics, Chinese Academy of Science
contact email	zhongbowen1810@dicp.ac.cn
dataset submitter

iProX dataset URI