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A systematic method for identifying small-molecule modulators of protein-protein interactions
Alexander R Horswill1, Sergey N Savinov, Stephen J Benkovic
1Department of Chemistry, Pennsylvania State University, 414 Wartik Laboratory, University Park, PA 16802, USA.
Summary
This study introduces a novel method to find small molecules that alter protein-protein interactions by linking complex disruption to cell survival. This approach successfully identified rare cyclic peptide inhibitors of ribonucleotide reductase.
Area of Science:
- Biochemistry
- Molecular Biology
- Drug Discovery
Background:
- Protein-protein interactions (PPIs) are crucial in cellular processes.
- Identifying small-molecule modulators of PPIs is challenging due to large, non-contiguous interfaces.
- Existing high-throughput methods for PPI inhibitor discovery are limited.
Purpose of the Study:
- To develop a robust methodology for discovering small-molecule modulators of protein-protein interactions.
- To demonstrate the feasibility of a functional approach coupling PPI disruption to host cell survival.
- To identify novel inhibitors of specific protein-protein interactions, such as ribonucleotide reductase.
Main Methods:
- A novel methodology coupling disruption of protein-protein complexes to host cell survival.
- Monitoring small-molecule-mediated protein-protein association (FKBP12-rapamycin-FRAP).
- Assessing dissociation of homodimeric HIV-1 protease and heterodimeric ribonucleotide reductase.
- Utilizing genetically encoded libraries for inhibitor screening.
- Developing solid-phase synthesis and peptide ELISAs for inhibitor characterization.
Main Results:
- Demonstrated feasibility using FKBP12-rapamycin-FRAP association and dissociation of HIV-1 protease and ribonucleotide reductase.
- Identified cyclic peptide inhibitors from genetically encoded libraries that dissociate ribonucleotide reductase subunits.
- Discovered cyclic peptides with an unprecedented mechanism of action.
- Successfully identified rare small-molecule PPI modulators at a frequency of less than one in 10 million.
Conclusions:
- The developed functional approach is effective for discovering small-molecule PPI modulators.
- Genetically encoded libraries coupled with a genetic selection strategy enable identification of rare inhibitors.
- This methodology offers a powerful tool for advancing drug discovery targeting PPIs.