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Multicomponent Yeast Two-Hybrid System: Applications to Study Protein–Protein Interactions in SMC Complexes

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SMC Complexes

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2004))

Abstract

Analysis of protein–protein interactions (PPI) is key for the understanding of most protein assemblies including structural maintenance of chromosomes (SMC) complexes. SMC complexes are composed of SMC proteins, kleisin, and kleisin-interacting subunits. These subunits interact in specific ways to constitute and regulate the closed structure of the complexes. Specifically, kleisin molecules bridge the SMC dimers and the kleisin-interacting subunits modulate stability of the bridge. Here we describe a multicomponent version of a yeast two-hybrid (Y2H) method and its application for analysis of the bridging role of the Nse4 kleisin in the SMC5/6 complex. Using this technique, we also show a stabilizing effect of KITE (kleisin-interacting tandem winged-helix element) proteins on SMC5/6.

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Acknowledgments

Czech Science Foundation grant GA18-02067S and the Ministry of Education, Youth and Sports of the Czech Republic project CEITEC 2020 (LQ1601) are acknowledged for their financial support. This chapter reflects only the author’s view, and the Research Executive Agency is not responsible for any use that may be made of the information it contains.

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Authors and Affiliations

  1. National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Brno, Czech Republic

    Jan Josef Paleček, Lucie Vondrová & Jakub Otočka

  2. Mendel Centre for Plant Genomics and Proteomics, Central European Institute of Technology, Masaryk University, Brno, Czech Republic

    Jan Josef Paleček

  3. Genome Damage and Stability Centre, University of Sussex, Brighton, UK

    Kateřina Zábrady

Authors
  1. Jan Josef Paleček
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  2. Lucie Vondrová
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  3. Kateřina Zábrady
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  4. Jakub Otočka
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Corresponding author

Correspondence to Jan Josef Paleček .

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Editors and Affiliations

  1. National Centre for Biological Sciences, Tata Institute of Fundamental Research (TIFR), Bangalore, India

    Anjana Badrinarayanan

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Paleček, J.J., Vondrová, L., Zábrady, K., Otočka, J. (2019). Multicomponent Yeast Two-Hybrid System: Applications to Study Protein–Protein Interactions in SMC Complexes. In: Badrinarayanan, A. (eds) SMC Complexes. Methods in Molecular Biology, vol 2004. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9520-2_7

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  • DOI: https://doi.org/10.1007/978-1-4939-9520-2_7

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9519-6

  • Online ISBN: 978-1-4939-9520-2

  • eBook Packages: Springer Protocols

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