PXD069830-2

PXD069830 is an original dataset announced via ProteomeXchange.

Title	Paradoxical non-catalytic kinase functions are driven by inhibitor-induced displacement of autoinhibitory domains
Description	Protein kinases are crucial regulators of cellular processes, and their dysregulation is frequently associated with human disorders. ATP-competitive type I inhibitors are widely used to block aberrant kinase activity. However, their effect on non-catalytic kinase functions such as conformation and protein-protein interactions (PPIs) remain largely uncharacterized. Here we present a multi-proteomics strategy to systematically measure these changes. First, we developed affinity purification (AP) and limited proteolysis coupled to mass spectrometry (AP-LiP-MS) to measure structural changes at high sequence coverage. We benchmarked AP-LiP-MS using DCLK1 before analyzing structural changes upon probe binding to CAMKK2, CHEK1, and PRKCA. All kinases underwent structural changes consistent with dissociation of domain-domain interactions (DDIs) between the kinase domain (KD) and the autoinhibitory domain (AID). Next, we applied AP and in vivo proximity labeling to analyze changes in kinase PPI networks. This revealed extensive and characteristic rewiring of functionally important PPIs, independent of catalytic activity. For instance, SCG-CAMKK2-1 binding to CAMKK2 sequestered PRKAA1, inhibiting its regulation by other kinases, while rabusertib caused CLPB-CHEK1 dissociation and mitochondrial fragmentation, and Gö 6983 resulted in PRKCA recruitment to cellular junctions. We propose that these unexpected phenotypes originate from on-target, off-mechanism effects, driven by structural changes and PPI rewiring. We advocate for the use of our workflow to systematically characterize overlooked consequences of small-molecule binding during drug development.
HostingRepository	PRIDE
AnnounceDate	2026-07-07
AnnouncementXML	Submission_2026-07-07_07:44:24.026.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Viviane Reber
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	carbamoylated residue; biotinylated residue; phosphorylated residue; acetylated residue; monohydroxylated residue
Instrument	Orbitrap Fusion Lumos; Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2025-10-23 08:26:33	ID requested
1	2026-06-18 15:38:52	announced
⏵ 2	2026-07-07 07:44:24	announced	2026-07-07: Updated project metadata.

10.1038/s44320-026-00229-2;

Reber V, Keller S, Loosli SA, Arima Y, Kleele T, Picotti P, Gstaiger M, Paradoxical non-catalytic kinase functions are driven by inhibitor-induced displacement of autoinhibitory domains. Mol Syst Biol, ():(2026) [pubmed]

submitter keyword: LiP-MS, proximity labeling, structural proteomics,AP-MS, BioID

Matthias Gstaiger
contact affiliation	Institute of Molecular Systems Biology, Department of Biology, ETH Zurich, Zurich, Switzerland
contact email	matthias.gstaiger@imsb.biol.ethz.ch
lab head
Viviane Reber
contact affiliation	IMSB, ETH Zurich
contact email	reberv@ethz.ch
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD069830
     1. Label: PRIDE project
     2. Name: Paradoxical non-catalytic kinase functions are driven by inhibitor-induced displacement of autoinhibitory domains