AlphaScreen selectivity assay for β-catenin/B-cell lymphoma 9 inhibitors

Anal Biochem. 2015 Jan 15:469:43-53. doi: 10.1016/j.ab.2014.09.018. Epub 2014 Oct 13.

Abstract

The aberrant formation of the β-catenin/B-cell lymphoma 9 (BCL9) protein-protein complex is the driving force for many diseases, including cancer. Crystallographic analyses demonstrate that the surface area in β-catenin for interacting with BCL9 is overlapped with that for the β-catenin/E-cadherin interaction. In this study, a robust AlphaScreen selectivity assay was developed to quantify inhibitor potency for the β-catenin/BCL9 interaction and selectivity for β-catenin/BCL9 over β-catenin/E-cadherin interactions. A pilot screen was performed to demonstrat the feasibility of this assay. This selectivity assay is highly sensitive and suitable for adaptation to high-throughput screening. The establishment of this assay lays the foundation for the discovery of selective inhibitors specific for β-catenin/BCL9 interactions.

Keywords: AlphaScreen; B-cell lymphoma 9; E-cadherin; Protein–protein interactions; Selectivity; β-Catenin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Abietanes / analysis
  • Cell Line, Tumor
  • Enzyme-Linked Immunosorbent Assay
  • Fluorescence Polarization*
  • Humans
  • Immunoprecipitation
  • Kinetics
  • Microscopy, Confocal
  • Neoplasm Proteins / antagonists & inhibitors
  • Neoplasm Proteins / genetics
  • Neoplasm Proteins / metabolism*
  • Plant Extracts / analysis
  • Protein Binding
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Surface Plasmon Resonance*
  • Transcription Factors
  • beta Catenin / antagonists & inhibitors
  • beta Catenin / genetics
  • beta Catenin / metabolism*

Substances

  • Abietanes
  • BCL9 protein, human
  • Neoplasm Proteins
  • Plant Extracts
  • Recombinant Proteins
  • Transcription Factors
  • beta Catenin
  • salvin