Technique / Proteomics and protein biochemistry / Protein-protein interactions / Yeast two-hybrid system

Yeast two-hybrid system protocols
()

1: Curr Protoc Mol Biol. 2008 Apr;Chapter 20:Unit 20.1.

Interaction trap/two-hybrid system to identify interacting proteins.
Golemis EA, Serebriiskii I, Finley RL Jr, Kolonin MG, Gyuris J, Brent R.
Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

The yeast two-hybrid method (or interaction trap) is a powerful technique for
detecting protein interactions. The procedure is performed using transcriptional
activation of a dual reporter system in yeast to identify interactions between a
protein of interest (the bait protein) and the candidate proteins for
interaction. The method can be used to screen a protein library for interactions
with a bait protein or to test for association between proteins that are expected
to interact based on prior evidence. Interaction mating facilitates the screening
of a library with multiple bait proteins. (c) 2008 by John Wiley & Sons, Inc.

2: Curr Protoc Mol Biol. 2004 Feb;Chapter 20:Unit 20.8.

Interaction trap/two-hybrid system to identify loss-of-interaction mutant
proteins.
Mendelsohn AR.
The Molecular Sciences Institute, Berkeley, California, USA.

Protein-protein interactions play a critical role in biology. Disrupting a
specific protein-protein interaction and studying the resulting phenotype can
help elucidate the function of the interaction. A method to rapidly make and
identify mutant proteins that no longer bind to a specific partner protein is
described. The method takes advantage of the ease of the interaction
trap/two-hybrid system and PCR mutagenesis. PCR fusion of the target protein to
GFP is used to ensure the protein's open reading frame is not disrupted by
mutagenesis. The resulting noninteracting mutant proteins usually result from a
single missense mutation, which can easily be identified.

3: Curr Protoc Mol Biol. 2002 Nov;Chapter 20:Unit 20.7.

Two-hybrid dual bait system.

Kotova E, Serebriiskii I, Coleman T.

Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

The yeast two-hybrid system, or interaction trap, is one of the most versatile
methods available with which to identify and establish protein-protein
interactions. The dual bait system is one adaptation of the classic approach.
This system facilitates the simultaneous comparison of two distinct baits with
one prey. One protein of interest is expressed as a fusion to the DNA binding
protein LexA (bait 1), while a second protein of interest is expressed as a
fusion to the DNA binding protein cI (bait 2). Strains of yeast engineered for
screening of these dual baits possess four separate reporter genes. A plasmid
expressing an activation domain-fused protein (prey), which can be either a
defined protein interactor or a cDNA library, is also expressed to allow dual
hybrid-mediated transcriptional activation.

(Selected reviews on yeast two-hybrid systems)
1: Parrish JR, Gulyas KD, Finley RL Jr.
Yeast two-hybrid contributions to interactome mapping.
Curr Opin Biotechnol. 2006 Aug;17(4):387-93. Epub 2006 Jun 27. Review.

2: Causier B.
Studying the interactome with the yeast two-hybrid system and mass
spectrometry.
Mass Spectrom Rev. 2004 Sep-Oct;23(5):350-67. Review.

3: Thaminy S, Miller J, Stagljar I.
The split-ubiquitin membrane-based yeast two-hybrid system.
Methods Mol Biol. 2004;261:297-312. Review.

4: Serebriiskii IG, Kotova E.
Analysis of protein-protein interactions utilizing dual bait yeast two-hybrid
system.
Methods Mol Biol. 2004;261:263-96. Review.

5: Miller J, Stagljar I.
Using the yeast two-hybrid system to identify interacting proteins.
Methods Mol Biol. 2004;261:247-62. Review.

6: Causier B, Davies B.
Analysing protein-protein interactions with the yeast two-hybrid system.
Plant Mol Biol. 2002 Dec;50(6):855-70. Review.

7: Coates PJ, Hall PA.
The yeast two-hybrid system for identifying protein-protein interactions.
J Pathol. 2003 Jan;199(1):4-7. Review.

8: Gietz RD, Woods RA.
Screening for protein-protein interactions in the yeast two-hybrid system.
Methods Mol Biol. 2002;185:471-86. Review.

9: Serebriiskii IG, Toby GG, Finley RL Jr, Golemis EA.
Genomic analysis utilizing the yeast two-hybrid system.
Methods Mol Biol. 2001;175:415-54. Review.

10: Serebriiskii IG, Khazak V, Golemis EA.
Redefinition of the yeast two-hybrid system in dialogue with changing
priorities in biological research.
Biotechniques. 2001 Mar;30(3):634-6, 638, 640 passim. Review.

11: Stephens DJ, Banting G.
The use of yeast two-hybrid screens in studies of protein:protein interactions
involved in trafficking.
Traffic. 2000 Oct;1(10):763-8. Review.

12: Topcu Z, Borden KL.
The yeast two-hybrid system and its pharmaceutical significance.
Pharm Res. 2000 Sep;17(9):1049-55. Review.

13: Serebriiskii IG, Golemis EA.
Uses of lacZ to study gene function: evaluation of beta-galactosidase assays
employed in the yeast two-hybrid system.
Anal Biochem. 2000 Oct 1;285(1):1-15. Review.

14: Biffo S, Marchisio PC.
The identification of signaling molecules by the yeast two-hybrid system.
Methods Mol Biol. 1999;96:211-20. Review.

15: Wallach D, Boldin MP, Kovalenko AV, Malinin NL, Mett IL, Camonis JH.
The yeast two-hybrid screening technique and its use in the study of
protein-protein interactions in apoptosis.
Curr Opin Immunol. 1998 Apr;10(2):131-6. Review.

16: White MA.
The yeast two-hybrid system: forward and reverse.
Proc Natl Acad Sci U S A. 1996 Sep 17;93(19):10001-3. Review. No abstract
available.

17: Bai C, Elledge SJ.
Gene identification using the yeast two-hybrid system.
Methods Enzymol. 1996;273:331-47. Review.

18: Luban J, Goff SP.
The yeast two-hybrid system for studying protein-protein interactions.
Curr Opin Biotechnol. 1995 Feb;6(1):59-64. Review.

19: Bemis LT, Geske FJ, Strange R.
Use of the yeast two-hybrid system for identifying the cascade of protein
interactions resulting in apoptotic cell death.
Methods Cell Biol. 1995;46:139-51. Review.