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Protein–Protein Interactions: Pull-Down Assays

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Bacterial Protein Secretion Systems

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1615))

Abstract

Determining protein partners is an essential step toward understanding protein function and identifying relevant biological pathways. Many methods exist for investigating protein–protein interactions. The pull-down assay is an in vitro technique used to detect physical interactions between two or more proteins and an invaluable tool for confirming a predicted protein–protein interaction or identifying novel interacting partners. This method typically involves the use of affinity purification with various wash and elution steps. In this chapter, we describe how an interaction between two purified bacterial proteins or between bacterial and eukaryotic proteins can be detected by pull-down experiments.

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References

  1. Costa TRD, Felisberto-Rodrigues C, Meir A, Prevost MS, Redzej A, Trokter M, Waksman G (2015) Secretion systems in Gram-negative bacteria: structural and mechanistic insights. Nat Rev Microbiol 13:343–359

    Article  CAS  PubMed  Google Scholar 

  2. McBride MJ, Nakane D (2015) Flavobacterium gliding motility and the type IX secretion system. Curr Opin Microbiol 28:72–77

    Article  CAS  PubMed  Google Scholar 

  3. Rodríguez-Negrete E, Bejarano ER, Castillo AG (2014) Using the yeast two-hybrid system to identify protein-protein interactions. Methods Mol Biol 1072:241–258

    Article  CAS  PubMed  Google Scholar 

  4. Zoued A, Brunet YR, Durand E, Aschtgen M-S, Logger L, Douzi B, Journet L, Cambillau C, Cascales E (2014) Architecture and assembly of the Type VI secretion system. Biochim Biophys Acta 1843:1664–1673

    Article  CAS  PubMed  Google Scholar 

  5. Boucrot E, Henry T, Borg J-P, Gorvel J-P, Méresse S (2005) The intracellular fate of Salmonella depends on the recruitment of kinesin. Science 308:1174–1178

    Article  CAS  PubMed  Google Scholar 

  6. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  CAS  Google Scholar 

  7. Di Guan C, Li P, Riggs PD, Inouye H (1988) Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. Gene 67:21–30

    Article  PubMed  Google Scholar 

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Correspondence to Sarah Bigot .

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Louche, A., Salcedo, S.P., Bigot, S. (2017). Protein–Protein Interactions: Pull-Down Assays. In: Journet, L., Cascales, E. (eds) Bacterial Protein Secretion Systems. Methods in Molecular Biology, vol 1615. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7033-9_20

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  • DOI: https://doi.org/10.1007/978-1-4939-7033-9_20

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7031-5

  • Online ISBN: 978-1-4939-7033-9

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