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26S and PA28-20S Proteasome Activity in Cytosolic Extracts from Embryonic Stem Cells

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Embryonic Stem Cell Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1341))

Abstract

The proteasome is a complex multisubunit protease that plays a major role in the degradation of proteins in eukaryotic cells. Proteasome function is one of the key players regulating the proteome and it is vital for many cellular processes. The method described here makes it possible to assay the proteolytic capacities of proteasome complexes separately in crude cytosolic extracts from ES cells. The method is based on hydrolysis of a fluorogenic peptide substrate in lysates prepared under conditions that favor the interactions of the 20S proteasomal catalytical core with either the 19S or the PA28αβ proteasome regulator.

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Acknowledgement

Thanks are due to Prof. Thomas Nyström, Prof. Jennifer Rivett, and Dr. Fiona Stratford for their valuable intellectual and methodological input in the development in this protocol. In addition, Prof. Thomas Nystr­m and Dr. Madina Karimova kindly reviewed the manuscript.

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Authors and Affiliations

  1. Discovery Sciences, AstraZeneca R&D, SE-431 83, Mölndal, Sweden

    Malin Hernebring

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  1. Malin Hernebring
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Correspondence to Malin Hernebring .

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Editors and Affiliations

  1. Ottawa Hospital Research Institute, Ottawa, Ontario, Canada

    Kursad Turksen

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Hernebring, M. (2015). 26S and PA28-20S Proteasome Activity in Cytosolic Extracts from Embryonic Stem Cells. In: Turksen, K. (eds) Embryonic Stem Cell Protocols. Methods in Molecular Biology, vol 1341. Humana Press, New York, NY. https://doi.org/10.1007/7651_2015_216

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  • DOI: https://doi.org/10.1007/7651_2015_216

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2953-5

  • Online ISBN: 978-1-4939-2954-2

  • eBook Packages: Springer Protocols

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