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  A Fluorescent In Vitro Assay to Investigate Paralog-Specific SUMO Conjugation

Eisenhardt, N., Chaugule, V., & Pichler, A. (2016). A Fluorescent In Vitro Assay to Investigate Paralog-Specific SUMO Conjugation. In Methods in Molecular Biology (pp. 67-78). New York: Springer + Business.

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 Creators:
Eisenhardt, Nathalie1, Author
Chaugule, Vidith1, Author
Pichler, Andrea1, Author              
Affiliations:
1Department of Epigenetics, Max Planck Institute of Immunobiology and Epigenetics, Max Planck Society, ou_2243644              

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Free keywords: SUMO, SUMO chains, E3 ligase, RanBP2, PIAS1, ZNF451, SUMO protease, In vitro sumoylation, Sp100
 Abstract: Protein modification with the small ubiquitin-related modifier SUMO is a potent regulatory mechanism implicated in a variety of biological pathways. In vitro sumoylation reactions have emerged as a versatile tool to identify and characterize novel SUMO enzymes as well as their substrates. Here, we present detailed protocols for the purification and fluorescent labeling of mammalian SUMO paralogs for their application in sumoylation assays. These assays provide a fast readout for in vitro SUMO chain formation activity of E3 ligases in a paralog-specific manner. Finally, we critically analyze the application of fluorescent SUMO proteins to study substrate modification in vitro revealing also the drawbacks of the system.

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Language(s): eng - English
 Dates: 2016
 Publication Status: Published in print
 Pages: -
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 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.1007/978-1-4939-6358-4_5
 Degree: -

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Title: Methods in Molecular Biology
Source Genre: Series
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Publ. Info: New York : Springer + Business
Pages: - Volume / Issue: 1475 Sequence Number: - Start / End Page: 67 - 78 Identifier: -