Protocol for the analysis of proteasome activity, assembly state, and composition in fission yeast extracts using native gels

The proteasome is a macromolecular complex responsible for degrading short-lived or damaged proteins. Proteasome analysis presents challenges due to its high molecular weight and low stability. Here, we present a protocol to assess proteasome activity, assembly, and composition in Schizosaccharomyce...

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Bibliographic Details
Authors: Ruiz Romero, Gabriel, Daga, Rafael R., Salas-Pino, Silvia
Format: article
Status:Published version
Publication Date:2025
Country:España
Institution:Consejo Superior de Investigaciones Científicas (CSIC)
Repository:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:dnet:digitalcsic_::10f1de0de24f02f230ec81f4bd276b08
Online Access:http://hdl.handle.net/10261/431647
Access Level:Open access
Description
Summary:The proteasome is a macromolecular complex responsible for degrading short-lived or damaged proteins. Proteasome analysis presents challenges due to its high molecular weight and low stability. Here, we present a protocol to assess proteasome activity, assembly, and composition in Schizosaccharomyces pombe. We describe steps for lysate preparation and native electrophoresis to separate proteasome complexes. We then detail procedures for detecting proteasome activity using fluorescent substrates, fluorescently-tagged native proteins, and non-tagged proteasome components by immunodetection in a single gel.

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