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A split intein and split luciferase-coupled system for detecting protein-protein interactions.

Zhong Yao1, Jiyoon Kim2, Betty Geng2

  • 1Donnelly Centre, University of Toronto, Toronto, ON, Canada. zhong.yao@utoronto.ca.

Molecular Systems Biology
|December 12, 2024
PubMed
Summary

A new Split Intein-Mediated Protein Ligation (SIMPL2) assay uses nanoluciferase for sensitive detection of protein-protein interactions (PPIs). This cost-effective tool enhances high-throughput screening for drug discovery and PPI mapping.

Keywords:
Drug DiscoveryInhibitorProtein-protein InteractionSplit InteinTri-part Nanoluciferase

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Drug Discovery

Background:

  • Protein-protein interactions (PPIs) are crucial in biomedical research.
  • Exploiting PPIs for drug discovery is a growing area of interest.
  • Previous Split Intein-Mediated Protein Ligation (SIMPL) method detects binary PPIs.

Purpose of the Study:

  • To enhance the SIMPL assay for improved quantifiability and operability.
  • To develop a sensitive and specific platform for PPI detection.
  • To create a modular system for screening PPI modulators.

Main Methods:

  • Incorporation of tripartite nanoluciferase (tNLuc) into the SIMPL system.
  • Homogenous liquid phase operation for luminescence-based detection.
  • Validation using a reference PPI set.

Main Results:

  • The SIMPL-tNLuc (SIMPL2) assay demonstrated improved sensitivity and specificity over the original SIMPL.
  • The SIMPL2 platform offers modularity for flexible measurement of PPI modulators.
  • Successful application in detecting inhibitors, molecular glues, and PROTACs.

Conclusions:

  • SIMPL2 is a sensitive, cost-effective, and labor-efficient tool for PPI research.
  • The platform is suitable for high-throughput screening (HTS) in PPI mapping.
  • SIMPL2 facilitates drug discovery efforts targeting PPIs.