Protein–protein interaction: Bacterial two-hybrid

Gouzel Karimova, Emilie Gauliard, Marilyne Davi, Scot P. Ouellette, Daniel Ladant

Research output: Chapter in Book/Report/Conference proceedingChapter

12 Scopus citations


The bacterial two-hybrid (BACTH, for “Bacterial Adenylate Cyclase-Based Two-Hybrid”) system is a simple and fast genetic approach to detecting and characterizing protein–protein interactions in vivo. This system is based on the interaction-mediated reconstitution of a cyclic adenosine monophosphate (cAMP) signaling cascade in Escherichia coli. As BACTH uses a diffusible cAMP messenger molecule, the physical association between the two interacting chimeric proteins can be spatially separated from the transcription activation readout, and therefore it is possible to analyze protein–protein interactions that occur either in the cytosol or at the inner membrane level as well as those that involve DNA-binding proteins. Moreover, proteins of bacterial origin can be studied in an environment similar (or identical) to their native one. The BACTH system may thus permit a simultaneous functional analysis of proteins of interest—provided the hybrid proteins retain their activity and their association state. This chapter describes the principle of the BACTH genetic system and the general procedures to study protein–protein interactions in vivo in E. coli.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Number of pages18
StatePublished - 2017

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745


  • Chimeric proteins
  • Membrane protein
  • Protein interaction assay
  • Two-hybrid system
  • cAMP signaling

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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    Karimova, G., Gauliard, E., Davi, M., Ouellette, S. P., & Ladant, D. (2017). Protein–protein interaction: Bacterial two-hybrid. In Methods in Molecular Biology (pp. 159-176). (Methods in Molecular Biology; Vol. 1615). Humana Press Inc..