Volume 22, Issue 11 p. 1961-1965
Full Paper

Approaches to Evaluate the Impact of a Small-Molecule Binder to a Noncatalytic Site of the Proteasome

Wenzhi Tian

Wenzhi Tian

Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana, 47907 USA

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Marianne E. Maresh

Marianne E. Maresh

Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana, 47907 USA

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Prof. Darci J. Trader

Corresponding Author

Prof. Darci J. Trader

Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana, 47907 USA

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First published: 22 February 2021
Citations: 1

Graphical Abstract

Investigating the impact of small molecules: There are a number of essential protein–protein interactions between the 19S regulatory particle and 20S core particle to form the full 26S proteasome. We investigated whether a small-molecule binder to Rpn-6, a 19S subunit, could disrupt 26S proteasome activity.

Abstract

Proteasome activity is crucial for cell survival and proliferation. In recent years, small molecules have been discovered that can affect the catalytic activity of the proteasome. Rather than targeting the active sites of the proteasome, it might be possible to affect ubiquitin-dependent degradation of proteins by limiting the association of the 19S regulatory particle (19S RP) with the 20S core particle (20S CP) of the proteasome. We recently described the discovery of TXS-8, a peptoid that binds to Rpn-6. Rpn-6 is a proteasome-associated protein that makes critical contacts with the 19S RP and the 20S CP. Herein, we present a general workflow to evaluate the impact of a small-molecule binder on proteasome activity by using TXS-8 as an example. This workflow contains three steps in which specific probes or overexpressed proteins in cells are used to determine whether the hydrolysis activity of the proteasome is affected. Although, in our case, TXS-8 did not affect proteasome activity, our workflow is highly amenable to studying a variety of small-molecule–proteasome subunit interactions.