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Curr Protoc Protein Sci. 2006 Dec;Chapter 19:Unit 19.15. doi: 10.1002/0471140864.ps1915s46.

Identifying small-molecule modulators of protein-protein interactions.

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1
University of Iowa, Iowa City, Iowa, USA.

Abstract

This unit outlines methods for identifying cyclic peptides that inhibit protein-protein interactions. Proteins of interest are cloned into a two-hybrid system engineered to operate in reverse, allowing the disruption of a protein complex to be coupled to cell growth. Cyclic peptide libraries are generated using an intein-based plasmid construct, and the cyclized sequence is randomized using a PCR procedure. By transforming plasmid libraries into host cells containing the two-hybrid fusions, cyclic peptide inhibitors can be identified by growing the cells under the appropriate selective conditions. A detailed procedure for performing the genetic selection and identifying false positives is provided. Methods for building the two-hybrid protein fusions and optimizing media conditions, as well as an additional protocol for constructing cyclic peptide libraries are also provided.

PMID:
18429304
DOI:
10.1002/0471140864.ps1915s46
[Indexed for MEDLINE]
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