UNIT 19.15 Identifying Small-Molecule Modulators of Protein-Protein Interactions

  1. Alexander R. Horswill1,
  2. Stephen J. Benkovic2

Published Online: 1 DEC 2006

DOI: 10.1002/0471140864.ps1915s46

Current Protocols in Protein Science

Current Protocols in Protein Science

How to Cite

Horswill, A. R. and Benkovic, S. J. 2006. Identifying Small-Molecule Modulators of Protein-Protein Interactions. Current Protocols in Protein Science. 46:19.15:19.15.1–19.15.19.

Author Information

  1. 1

    University of Iowa, Iowa City, Iowa

  2. 2

    The Pennsylvania State University, University Park, Pennsylvania

Publication History

  1. Published Online: 1 DEC 2006
  2. Published Print: NOV 2006


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.


  • small-molecule;
  • protein-protein;
  • cyclic peptide;
  • two-hybrid;
  • inhibitor