PXD072417

PXD072417 is an original dataset announced via ProteomeXchange.

Title	Development of Substrate-Directed Activity-Based Probes via Disulfide-Trapping for Site-Specific Interrogation of Nucleosomal Deubiquitination
Description	Activity-based ubiquitin probes (Ub-ABPs) have been instrumental in interrogating deubiquitinases (DUBs), yet conventional Ub-ABPs lack the substrate-specific and site-specific contextual cues required to investigate nucleosomal DUBs, whose activity is tightly regulated by the chromatin environments. Here, we developed a substrate-directed nucleosomal DUB activity-based probe (SD-NucDUB-ABP) employing a disulfide trapping mechanism. This probe features a 2,2′-dithiodipyridine-activated 2-mercaptoethyl modification on ubiquitinated histone isopeptide amide, enabling selective covalent capture of nucleosome-interacting DUBs via their catalytic cysteines while remaining resistant to hydrolysis. Using this strategy, we synthesized the H2AK15UbAT nucleosome probe and applied it in activity-based profiling of DUBs in nuclear lysates of DNA-damaged HeLa cells. Subsequent biochemical analyses confirmed USP3 as a bona fide DUB for H2AK15Ub nucleosomes. Cross-linking mass spectrometry (XL-MS) further delineated the spatial interaction network among USP3, Ub and the nucleosome, providing mechanistic insights into H2AK15 deubiquitination. Furthermore, we extended this strategy to other nucleosomal ubiquitination sites (H2AK119Ub and H2BK120Ub), capturing their cognate DUBs (USP16 and Ubp10, respectively). The SD-NucDUB-ABP platform thus enables integrated proteomic discovery and mechanistic dissection of nucleosome-specific deubiquitination, providing a versatile chemical tool for epigenetic research at the intersection of chemical biology and chromatin biology.
HostingRepository	iProX
AnnounceDate	2025-12-25
AnnouncementXML	Submission_2025-12-24_22:59:22.467.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Yanling Zhang
SpeciesList	scientific name: Homo sapiens; NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Q Exactive Plus

Revision	Datetime	Status	ChangeLog Entry
0	2025-12-24 22:59:08	ID requested
⏵ 1	2025-12-24 22:59:23	announced

Dataset with its publication pending

submitter keyword: Activity-based ubiquitin probes, nucleosomal DUB, DNA damage, proteomics

Wei Qin
contact affiliation	Tsinghua University
contact email	weiqin@tsinghua.edu.cn
lab head
Yanling Zhang
contact affiliation	Tsinghua University
contact email	yanlingzhang@mail.tsinghua.edu.cn
dataset submitter

iProX dataset URI