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A Combined Yeast/Bacteria Two-hybrid System: Development and Evaluation
Serebriiskii, Ilya G. Fang, Rui Latypova, Ekaterina Hopkins, Richard Vinson, Charles Joung, J. Keith
Serebriiskii, Ilya G.
Fang, Rui
Latypova, Ekaterina
Hopkins, Richard
Vinson, Charles
Joung, J. Keith
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Journal article
Date
2005-03-20
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Cancer and Cellular Biology
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http://dx.doi.org/10.1074/mcp.t500005-mcp200
Abstract
Two-hybrid screening is a standard method used to identify and characterize protein-protein interactions and has become an integral component of many proteomic investigations. The two-hybrid system was initially developed using yeast as a host organism. However, bacterial two-hybrid systems have also become common laboratory tools and are preferred in some circumstances, although yeast and bacterial two-hybrid systems have never been directly compared. We describe here the development of a unified yeast and bacterial two-hybrid system in which a single bait expression plasmid is used in both organismal milieus. We use a series of leucine zipper fusion proteins of known affinities to compare interaction detection using both systems. Although both two-hybrid systems detected interactions within a comparable range of interaction affinities, each demonstrated unique advantages. The yeast system produced quantitative readout over a greater dynamic range than that observed with bacteria. However, the phenomenon of “autoactivation” by baits was less of a problem in the bacterial system than in the yeast. Both systems identified physiological interactors for a library screen with a cI-Ras test bait; however, non-identical interactors were obtained in yeast and bacterial screens. The ability to rapidly shift between yeast and bacterial systems provided by these new reagents should provide a marked advantage for two-hybrid investigations. In addition, the modified expression vectors we describe in this report should be useful for any application requiring facile expression of a protein of interest in both yeast and bacteria.
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Citation
Serebriiskii IG, Fang R, Latypova E, Hopkins R, Vinson C, Joung JK, Golemis EA. A Combined Yeast/Bacteria Two-hybrid System: Development and Evaluation. Mol Cell Proteomics. 2005 Mar 20;4(6):819-826. doi:10.1074/mcp.T500005-MCP200.
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Elsevier
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Molecular & Cellular Proteomics, Vol. 4, Iss. 6
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