Abstract
To characterize a protein’s function, it is often advantageous to identify other proteins with which it interacts. The yeast two-hybrid system is one of the most versatile methods available for detection and characterization of protein-protein interactions, and in the recent years it has become a mature and robust technology. A further improvement to this technique is the ability to examine and distinguish more than one interaction simultaneously. This is achieved in the Dual Bait, which has successfully been used to detect proteins and peptides that target specific motifs in larger proteins, to facilitate rapid identification of specific interactors from a pool of putative interacting proteins obtained in a library screen, and to score specific drug-mediated disruption of protein-protein interaction.
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References
Fields, S. and Song, O. (1989) A novel genetic system to detect protein-protein interaction. Nature 340, 245–246.
Ito, T. et al. (2001) A comprehensive two-hybrid analysis to explore the yeast protein interactome. Proc. Natl. Acad. Sci. USA 98, 4569–4574.
Schwikowski, B., Uetz, P., and Fields, S. (2000) A network of protein-protein interactions in yeast. Nat. Biotechnol. 18, 1257–1261.
Inouye, C., Dhillon, N., Durfee, T., Zambryski, P. C., and Thorner, J. (1997) Mutational analysis of STE5 in the yeast Saccharomyces cerevisiae: application of a differential interaction trap assay for examining protein-protein interactions. Genetics 147, 479–492
Xu, C. W., Mendelsohn, A. R., and Brent, R. (1997) Cells that register logical relationships among proteins. Proc. Natl. Acad. Sci. USA 94, 12,473–12,478
Serebriiskii, I., Khazak, V., and Golemis, E. A. (1999) A two-hybrid dual bait system to discriminate specificity of protein interactions. J. Biol. Chem. 274, 17,080–17,087.
Gyuris, J., Golemis, E. A., Chertkov, H., and Brent, R. (1993) Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell 75, 791–803.
Reeder, M. K., Serebriiskii, I. G., Golemis, E. A., and Chernoff, J. (2001) Analysis of small GTPase signaling pathways using Pak1 mutants that selectively couple to Cdc42. J. Biol. Chem. 276, 40,606–40,613.
Schiestl, R. H. and Gietz, R. D. (1989) High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier. Curr. Genet. 16, 339–346.
Duttweiler, H. M. (1996) A highly sensitive and non-lethal beta-galactosidase plate assay for yeast. TIG 12, 340–341.
Harlow, E. and Lane, D. (1988). Antibodies: a Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.
Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989). Molecular Cloning: a Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.
Finley, R. and Brent, R. (1994) Interaction mating reveals binary and ternary connections between Drosophila cell cycle regulators. Proc. Nat. Acad. Sci. USA 91, 12,980–12,984.
Petermann, R., Mossier, B. M., Aryee, D. N., and Kovar, H. (1998) A recombination based method to rapidly assess specificity of two-hybrid clones in yeast. Nucleic Acids Res. 26, 2252–2253.
Oldenburg, K. R., Vo, K. T., Michaelis, S., and Paddon, C. (1997) Recombination-mediated PCR-directed plasmid construction in vivo in yeast. Nucleic Acids Res. 25, 451–452.
DeMarini, D. J., Creasy, C. L., Lu, Q., et al. (2001) Oligonucleotide-mediated, PCR-independent cloning by homologous recombination. Biotechniques 30, 520–523.
Serebriiskii, I., Toby, G., Finley, R.L., and Golemis, E.A.. (2001). Genomic analysis utilizing the yeast two-hybrid system, in Genomic Protocols (Starkey, M. ed.), Humana, Totowa, NJ, pp. 415–454.
Serebriiskii, I. G., Mitina, O., Pugacheva, E. N., et al. (2002) Detection of peptides, proteins, and drugs that selectively interact with protein targets. Genome Res. 12, 1785–1791.
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Serebriiskii, I.G., Kotova, E. (2004). Analysis of Protein-Protein Interactions Utilizing Dual Bait Yeast Two-Hybrid System. In: Fu, H. (eds) Protein-Protein Interactions. Methods in Molecular Biology, vol 261. Humana Press. https://doi.org/10.1385/1-59259-762-9:263
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DOI: https://doi.org/10.1385/1-59259-762-9:263
Publisher Name: Humana Press
Print ISBN: 978-1-58829-120-2
Online ISBN: 978-1-59259-762-8
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