Characterization of PROTAC specificity and endogenous protein interactomes using ProtacID

Suman Shrestha, Matthew E. R. Maitland, Laili Jing, Shili Duan, David Y. Nie, Jonathan St-Germain, Michael Kanaris, Dalia Barsyte-Lovejoy, Cheryl H. Arrowsmith () and Brian Raught ()
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Suman Shrestha: University Health Network
Matthew E. R. Maitland: University Health Network
Laili Jing: University Health Network
Shili Duan: University Health Network
David Y. Nie: University Health Network
Jonathan St-Germain: University Health Network
Michael Kanaris: University of Toronto
Dalia Barsyte-Lovejoy: University of Toronto
Cheryl H. Arrowsmith: University Health Network
Brian Raught: University Health Network

Nature Communications, 2025, vol. 16, issue 1, 1-11

Abstract: Abstract Here we describe ProtacID, a flexible BioID (proximity-dependent biotinylation)-based approach to identify PROTAC-proximal proteins in living cells. ProtacID analysis of VHL- and CRBN-recruiting PROTACs targeting a number of different proteins (localized to chromatin or cellular membranes, and tested across six different human cell lines) demonstrates how this technique can be used to validate PROTAC degradation targets and identify non-productive (i.e. non-degraded) PROTAC-interacting proteins, addressing a critical need in the field of PROTAC development. We also demonstrate that ProtacID can be used to characterize native, endogenous multiprotein complexes without the use of antibodies, or modification of the protein of interest with epitope tags or biotin ligase tagging.

Date: 2025
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DOI: 10.1038/s41467-025-63357-7

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