In-gel proteasome assay to determine the activity, amount, and composition of proteasome complexes from mammalian cells or tissues

Ayse Seda Yazgili; Thomas Meul; Vanessa Welk; Nora Semren; Ilona E Kammerl; Silke Meiners

doi:10.1016/j.xpro.2021.100526

In-gel proteasome assay to determine the activity, amount, and composition of proteasome complexes from mammalian cells or tissues

STAR Protoc. 2021 May 4;2(2):100526. doi: 10.1016/j.xpro.2021.100526. eCollection 2021 Jun 18.

Authors

Ayse Seda Yazgili¹, Thomas Meul¹, Vanessa Welk¹, Nora Semren¹, Ilona E Kammerl¹, Silke Meiners¹

Affiliation

¹ Comprehensive Pneumology Center (CPC), University Hospital of the Ludwig-Maximilians-University (LMU) and Helmholtz Zentrum München, Member of the German Center for Lung Research (DZL), Max-Lebsche Platz 31, 81377 Munich, Germany.

Abstract

This protocol describes an easy and reliable in-gel proteasome assay to quantify the activity and composition of different proteasome complexes in cells and tissues. The assay works well with limited amounts of total cell protein lysates. Although this assay is optimized specifically for the proteasome chymotrypsin-like activity, it can be expanded to other proteasome activities as well. Using antibodies that detect distinct proteasome subunits or regulators, we can determine the composition and relative quantity of active proteasome complexes. For complete details on the use and execution of this protocol, please refer to Meul et al. (2020).

Keywords: Antibody; Cell Biology; Molecular Biology; Protein Biochemistry.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

A549 Cells
Blotting, Western
Cells, Cultured
Cytological Techniques / methods*
Humans
Native Polyacrylamide Gel Electrophoresis
Proteasome Endopeptidase Complex* / analysis
Proteasome Endopeptidase Complex* / chemistry
Proteasome Endopeptidase Complex* / metabolism

Substances

Proteasome Endopeptidase Complex