PXD047469

PXD047469 is an original dataset announced via ProteomeXchange.

Title	It’s a trap: ClpP substrates caught in action
Description	Protein homeostasis in bacteria is regulated by proteases such as the tetradecameric caseinolytic protease P (ClpP). Although substrates of ClpP have been successfully deciphered in genetically engineered cells, methods which directly trap processed proteins within native cells are elusive. Here we introduce an in situ trapping strategy which utilizes trifunctional probes that bind to the active site serine of ClpP and capture adjacent substrates with an attached photocrosslinking moiety. Substrate deconvolution by mass spectrometry (MS) is accomplished after enrichment using an alkyne handle. We show that our two traps bind substoichiometrically to ClpP, retain protease activity, exhibit unprecedented selectivity for Staphylococcus aureus ClpP in living cells and capture numerous known and novel substrates. The exemplary validation of trapped hits using a targeted proteomics approach confirmed the fidelity of this technology. In conclusion, we provide a novel chemical platform suited to discover serine protease substrates beyond genetic engineering.
HostingRepository	PanoramaPublic
AnnounceDate	2024-10-03
AnnouncementXML	Submission_2024-10-03_10:56:44.662.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Laura Eck
SpeciesList	scientific name: Staphylococcus aureus; NCBI TaxID: 1280;
ModificationList	No PTMs are included in the dataset
Instrument	Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2023-12-03 00:11:35	ID requested
⏵ 1	2024-10-03 10:56:45	announced

Angew. Chem. Int. Ed. 2024, e202409220

submitter keyword: S. aureus, ClpP, protease substrate trapping, ClpP substrates, parallel reaction monitoring

Stephan Sieber
contact affiliation	TUM
contact email	stephan.sieber@tum.de
lab head
Laura Eck
contact affiliation	TUM
contact email	laura.eck@tum.de
dataset submitter

Panorama Public dataset URI