Analysis of Membrane Protein Interactions with a Bacterial Adenylate Cyclase-Based Two-Hybrid (BACTH) Technique

Autor: Daniel Ladant, Marilyne Davi, Gouzel Karimova, Scot P. Ouellette
Přispěvatelé: Biochimie des Interactions Macromoléculaires / Biochemistry of Macromolecular Interactions, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Sanford School of Medicine [Vermillion], University of South Dakota [Vermillion] (USD), This work was supported by Institut Pasteur and the Centre National de la Recherche Scientifique (CNRS UMR 3528, Biologie Structurale et Agents Infectieux)., Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)
Jazyk: angličtina
Rok vydání: 2017
Předmět:
0301 basic medicine
MESH: Signal Transduction
Operon
[SDV]Life Sciences [q-bio]
lac operon
MESH: Recombinant Fusion Proteins / metabolism
MESH: Escherichia coli / genetics
Bordetella pertussis
Protein Interaction Mapping
Cyclic AMP
membrane protein
MESH: Animals
Protein Interaction Maps
Cloning
Molecular
MESH: Protein Interaction Maps
MESH: Adenylyl Cyclases / metabolism
MESH: Membrane Proteins / genetics
Chemistry
two-hybrid technique
protein interaction assay
General Medicine
Biochemistry
Second messenger system
MESH: Escherichia coli / metabolism
Adenylyl Cyclases
Signal Transduction
MESH: Cloning
Molecular / methods
MESH: Recombinant Fusion Proteins / genetics
library screening
MESH: Protein Interaction Mapping / methods
Recombinant Fusion Proteins
030106 microbiology
MESH: Bordetella pertussis / metabolism
Adenylate kinase
MESH: Adenylyl Cyclases / genetics
Cyclase
03 medical and health sciences
MESH: Escherichia coli / enzymology
MESH: Two-Hybrid System Techniques
Two-Hybrid System Techniques
MESH: Membrane Proteins / metabolism
Escherichia coli
Animals
Humans
MESH: Bordetella pertussis / genetics
MESH: Humans
Membrane Proteins
[SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry
Molecular Biology/Molecular biology
MESH: Bordetella pertussis / enzymology
Fusion protein
[SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology
cAMP signaling
030104 developmental biology
Regulon
MESH: Cyclic AMP / metabolism
Membrane protein
Zdroj: Current protocols in molecular biology
Current protocols in molecular biology, Wiley, 2017, 118, pp.20.12.1-20.12.24. ⟨10.1002/cpmb.36⟩
Current Protocols in Molecular Biology
Current Protocols in Molecular Biology, 2017, 118, pp.20.12.1-20.12.24. ⟨10.1002/cpmb.36⟩
ISSN: 1934-3647
DOI: 10.1002/cpmb.36⟩
Popis: The bacterial two-hybrid (BACTH, for "Bacterial Adenylate Cyclase-based Two-Hybrid") technique is a simple and fast genetic approach to analyze protein-protein interactions in vivo. In this system, the proteins of interest are genetically fused to two complementary fragments from the catalytic domain of Bordetella pertussis adenylate cyclase and co-expressed in strains of Escherichia coli deficient in adenylate cyclase. Association of the hybrid proteins restores synthesis of cyclic AMP (cAMP), which then triggers the expression of catabolic operons such as the lactose operon or the maltose regulon. As BACTH uses a cAMP second messenger, the association between the chimeric proteins can take place at a distance from the transcription machinery. This technique is therefore particularly appropriate for studying interactions involving integral-membrane or membrane-associated proteins that may not be soluble in the cytoplasm, and/or that may only associate in the plane of the membrane. This unit describes the basic procedures to characterize protein-protein interactions with the BACTH genetic system and to search for potential partners of known proteins. © 2017 by John Wiley & Sons, Inc.
Databáze: OpenAIRE
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