Small Molecule Screen Identifies Non-catalytic USP3 Chemical Handle

Mandeep K Mann; Esther Wolf; Madhushika Silva; Haejin Angela Kwak; Brian Wilson; Albina Bolotokova; Derek J Wilson; Rachel J Harding; Matthieu Schapira

doi:10.1021/acsomega.3c07070

Small Molecule Screen Identifies Non-catalytic USP3 Chemical Handle

ACS Omega. 2023 Dec 15;9(1):917-924. doi: 10.1021/acsomega.3c07070. eCollection 2024 Jan 9.

Authors

Mandeep K Mann^{1

2}, Esther Wolf³, Madhushika Silva¹, Haejin Angela Kwak^{1

2}, Brian Wilson⁴, Albina Bolotokova¹, Derek J Wilson³, Rachel J Harding^{1

2}, Matthieu Schapira^{1

2}

Affiliations

¹ Structural Genomics Consortium, University of Toronto, 101 College Street, MaRS South Tower, Suite 700, Toronto, Ontario M5G 1L7, Canada.
² Department of Pharmacology and Toxicology, University of Toronto, 1 King's College Circle, Toronto, Ontario M5S 1A8, Canada.
³ Department of Chemistry, York University, Toronto, Ontario M3J 1P3, Canada.
⁴ Drug Discovery Program, Ontario Institute for Cancer Research, Toronto, Ontario M5G 0A3, Canada.

Abstract

Zinc-finger ubiquitin-binding domains (ZnF-UBDs) are noncatalytic domains mostly found in deubiquitylases (DUBs) such as USP3. They represent an underexplored opportunity for the development of deubiquitylase-targeting chimeras (DUBTACs) to pharmacologically induce the deubiquitylation of target proteins. We previously showed that ZnF-UBDs are ligandable domains. Here, a focused small molecule library screen against a panel of 11 ZnF-UBDs led to the identification of compound 59, a ligand engaging the ZnF-UBD of USP3 with a K_D of 14 μM. The compound binds the expected C-terminal ubiquitin binding pocket of USP3 as shown by hydrogen-deuterium exchange mass spectrometry experiments and does not inhibit the cleavage of K48-linked diubiquitin by USP3. As such, this molecule is a chemical starting point toward chemical tools that could be used to interrogate the function of the USP3 Znf-UBD and the consequences of recruiting USP3 to ubiquitylated proteins.