Identifying small-molecule modulators of protein-protein interactions.

Alexander R. Horswill, Stephen Benkovic

Research output: Contribution to journalArticle

2 Citations (Scopus)

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.

Original languageEnglish (US)
JournalCurrent protocols in protein science / editorial board, John E. Coligan ... [et al.]
VolumeChapter 19
StatePublished - Jan 1 2006

Fingerprint

Cyclic Peptides
Modulators
Molecules
Peptide Library
Proteins
Plasmids
Fusion reactions
Inteins
Two-Hybrid System Techniques
Genetic Selection
Cell growth
Hybrid systems
Polymerase Chain Reaction
Growth

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Biochemistry

Cite this

@article{f6cd62fffd614c6fb42cf49063370594,
title = "Identifying small-molecule modulators of protein-protein interactions.",
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.",
author = "Horswill, {Alexander R.} and Stephen Benkovic",
year = "2006",
month = "1",
day = "1",
language = "English (US)",
volume = "Chapter 19",
journal = "Current Protocols in Protein Science",
issn = "1934-3655",
publisher = "John Wiley and Sons Inc.",

}

Identifying small-molecule modulators of protein-protein interactions. / Horswill, Alexander R.; Benkovic, Stephen.

In: Current protocols in protein science / editorial board, John E. Coligan ... [et al.], Vol. Chapter 19, 01.01.2006.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Identifying small-molecule modulators of protein-protein interactions.

AU - Horswill, Alexander R.

AU - Benkovic, Stephen

PY - 2006/1/1

Y1 - 2006/1/1

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=48849094688&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=48849094688&partnerID=8YFLogxK

M3 - Article

C2 - 18429304

AN - SCOPUS:48849094688

VL - Chapter 19

JO - Current Protocols in Protein Science

JF - Current Protocols in Protein Science

SN - 1934-3655

ER -