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Detection of Protein–Protein Interactions Utilizing the Split-Ubiquitin Membrane-Based Yeast Two-Hybrid System

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Protein-Protein Interactions

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2690))

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Abstract

Identifying the interactors of a protein is a key step in understanding its possible cellular function(s). Among the various methods that can be used to study protein-protein interactions (PPIs), the yeast two-hybrid (Y2H) assay is one of the most standardized, sensitive, and cost-effective in vivo methods available. The most commonly used GAL4-based Y2H system utilizes the yeast transcription factor GAL4 to detect interactions between soluble proteins. By virtue of involving a transcription factor, the protein–protein interactions occur in the nucleus. The split-ubiquitin Y2H system offers an alternative to the traditional GAL4-based Y2H system and takes advantage of the reconstitution of split-ubiquitin in the cytosol to identify interactions between two proteins. Moreover, new membranous and soluble interacting partner(s) can be identified by screening a target protein against proteins produced from a cDNA library using this system.

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Author information

Authors and Affiliations

  1. Department of Microbiology and Biotechnology, Sister Nivedita University, Kolkata, West Bengal, India

    Siddhartha Dutta

  2. Department of Biology, Wilfrid Laurier University, Waterloo, ON, Canada

    Matthew D. Smith

Authors
  1. Siddhartha Dutta
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  2. Matthew D. Smith
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Corresponding author

Correspondence to Matthew D. Smith .

Editor information

Editors and Affiliations

  1. Department of Biology, University of Alabama at Birmingham, Birmingham, AL, USA

    Shahid Mukhtar

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Dutta, S., Smith, M.D. (2023). Detection of Protein–Protein Interactions Utilizing the Split-Ubiquitin Membrane-Based Yeast Two-Hybrid System. In: Mukhtar, S. (eds) Protein-Protein Interactions. Methods in Molecular Biology, vol 2690. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3327-4_4

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  • DOI: https://doi.org/10.1007/978-1-0716-3327-4_4

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3326-7

  • Online ISBN: 978-1-0716-3327-4

  • eBook Packages: Springer Protocols

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