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Studying protein complexes by the yeast two-hybrid system.

ABSTRACT: Protein complexes are typically analyzed by affinity purification and subsequent mass spectrometric analysis. However, in most cases the structure and topology of the complexes remains elusive from such studies. Here we investigate how the yeast two-hybrid system can be used to analyze direct interactions among proteins in a complex. First we tested all pairwise interactions among the seven proteins of Escherichia coli DNA polymerase III as well as an uncharacterized complex that includes MntR and PerR. Four and seven interactions were identified in these two complexes, respectively. In addition, we review Y2H data for three other complexes of known structure which serve as "gold-standards", namely Varicella Zoster Virus (VZV) ribonucleotide reductase (RNR), the yeast proteasome, and bacteriophage lambda. Finally, we review an Y2H analysis of the human spliceosome which may serve as an example for a dynamic mega-complex.

SUBMITTER: Rajagopala SV

PROVIDER: S-EPMC3517932 | biostudies-literature | 2012 Dec

REPOSITORIES: biostudies-literature

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Studying protein complexes by the yeast two-hybrid system.

Rajagopala Seesandra V SV Sikorski Patricia P Caufield J Harry JH Tovchigrechko Andrey A Uetz Peter P

Methods (San Diego, Calif.) 20120724 4

Protein complexes are typically analyzed by affinity purification and subsequent mass spectrometric analysis. However, in most cases the structure and topology of the complexes remains elusive from such studies. Here we investigate how the yeast two-hybrid system can be used to analyze direct interactions among proteins in a complex. First we tested all pairwise interactions among the seven proteins of Escherichia coli DNA polymerase III as well as an uncharacterized complex that includes MntR a ...[more]

PMID: 22841565

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