PXD028138-1

PXD028138 is an original dataset announced via ProteomeXchange.

Title	A BRET Biosensor for Measuring Uncompetitive Engagement of PRMT5 Complexes in Cells
Description	Protein arginine methyl transferase 5 (PRMT5) plays a global role in cell physiology and is an established therapeutic target in cancer. In approximately 10-15% of human cancers, deletion of the methylthioadenosine phosphorylase (MTAP) gene results in accumulation of methylthioadenosine (MTA), exposing a synthetic lethality and opportunity for precision medicine by selective targeting of PRMT5 in this context. Reported small molecule PRMT5 inhibitors engage either cosubstrate S-adenosyl methionine (SAM) or peptide-substrate pockets through diverse mechanisms. A subset of chemotypes demonstrate uncompetitive engagement with SAM or its inhibitory metabolic precursor, MTA. Although uncompetitive engagement can be evaluated in cell-free systems, no methods exist to directly assess this in cells. Here, we describe the development of a fluorescent probe that acts as a dynamic BRET biosensor of the intracellular SAM/MTA pool that overcomes the current limitations of competitive binding analyses. Using this biosensor, we evaluate a range of diverse PRMT5 inhibitors to mechanistically characterize and quantify uncompetitive target engagement as well as ternary complex formation at PRMT5-SAM and PRMT5-MTA complexes in live cells, enabling direct insights into drug mechanism-of-action and metabolite-dependent responses of inhibitors.
HostingRepository	PRIDE
AnnounceDate	2025-12-08
AnnouncementXML	Submission_2025-12-07_18:38:35.925.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Elisabeth Mira Rothweiler
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2021-08-26 04:05:12	ID requested
⏵ 1	2025-12-07 18:38:36	announced

10.1038/s41467-025-65558-6;

Rothweiler EM, Michaud A, Stefaniak J, Singh U, Mikulsky BB, Vasta JD, Beck MT, Wilkinson J, Ward JA, Rogers CM, Bal, ı, k, ç, ı E, Tranberg-Jensen J, Hansen JS, Loppnau P, Whitty A, Brennan PE, Tonge PJ, Robers MB, Huber KVM, A BRET biosensor for measuring uncompetitive engagement of PRMT5 complexes in cells. Nat Commun, 16(1):10129(2025) [pubmed]

submitter keyword: NanoBRET - chemical proteomics – cell-based assay – SAM – target engagement – thermal profiling

Kilian Huber
contact affiliation	Nuffield Department of Medicine
contact email	kilian.huber@cmd.ox.ac.uk
lab head
Elisabeth Mira Rothweiler
contact affiliation	University of Oxford
contact email	elisabeth.rothweiler@wolfson.ox.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD028138
     1. Label: PRIDE project
     2. Name: A BRET Biosensor for Measuring Uncompetitive Engagement of PRMT5 Complexes in Cells