Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1997 May 15;25(10):2035–2036. doi: 10.1093/nar/25.10.2035

Getting more from the two-hybrid system: N-terminal fusions to LexA are efficient and sensitive baits for two-hybrid studies.

F Béranger 1, S Aresta 1, J de Gunzburg 1, J Camonis 1
PMCID: PMC146676  PMID: 9115375

Abstract

Two-hybrid methods detect interactions between two proteins fused at the C-termini of, respectively, a DNA-binding domain and the activation domain of a transcriptional activator. Thus the N-terminus of none of these proteins is available for interaction. We have tested whether a bait protein with a reverted polarity (i.e. N-bait-LexA-C) is suitable for two-hybrid interaction. We show that such constructs give a specific interaction signal, and document two cases where the sensitivity is dramatically increased. Such constructs might lead to the identification of partners missed during classical two-hybrid screens.

Full Text

The Full Text of this article is available as a PDF (39.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cantor S. B., Urano T., Feig L. A. Identification and characterization of Ral-binding protein 1, a potential downstream target of Ral GTPases. Mol Cell Biol. 1995 Aug;15(8):4578–4584. doi: 10.1128/mcb.15.8.4578. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Fields S., Song O. A novel genetic system to detect protein-protein interactions. Nature. 1989 Jul 20;340(6230):245–246. doi: 10.1038/340245a0. [DOI] [PubMed] [Google Scholar]
  3. Gietz R. D., Sugino A. New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene. 1988 Dec 30;74(2):527–534. doi: 10.1016/0378-1119(88)90185-0. [DOI] [PubMed] [Google Scholar]
  4. Gyuris J., Golemis E., Chertkov H., Brent R. Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell. 1993 Nov 19;75(4):791–803. doi: 10.1016/0092-8674(93)90498-f. [DOI] [PubMed] [Google Scholar]
  5. Hofer F., Fields S., Schneider C., Martin G. S. Activated Ras interacts with the Ral guanine nucleotide dissociation stimulator. Proc Natl Acad Sci U S A. 1994 Nov 8;91(23):11089–11093. doi: 10.1073/pnas.91.23.11089. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Jullien-Flores V., Dorseuil O., Romero F., Letourneur F., Saragosti S., Berger R., Tavitian A., Gacon G., Camonis J. H. Bridging Ral GTPase to Rho pathways. RLIP76, a Ral effector with CDC42/Rac GTPase-activating protein activity. J Biol Chem. 1995 Sep 22;270(38):22473–22477. doi: 10.1074/jbc.270.38.22473. [DOI] [PubMed] [Google Scholar]
  7. Jung V., Wei W., Ballester R., Camonis J., Mi S., Van Aelst L., Wigler M., Broek D. Two types of RAS mutants that dominantly interfere with activators of RAS. Mol Cell Biol. 1994 Jun;14(6):3707–3718. doi: 10.1128/mcb.14.6.3707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Spaargaren M., Bischoff J. R. Identification of the guanine nucleotide dissociation stimulator for Ral as a putative effector molecule of R-ras, H-ras, K-ras, and Rap. Proc Natl Acad Sci U S A. 1994 Dec 20;91(26):12609–12613. doi: 10.1073/pnas.91.26.12609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Vojtek A. B., Hollenberg S. M., Cooper J. A. Mammalian Ras interacts directly with the serine/threonine kinase Raf. Cell. 1993 Jul 16;74(1):205–214. doi: 10.1016/0092-8674(93)90307-c. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES