High-throughput screen for small molecule inhibitors of Mint1-PDZ domains

Xuesong Chen, Jamie C. Longgood, Carolyn Michnoff, Shuguang Wei, Doug E. Frantz, Llya Bezprozvanny

Research output: Contribution to journalArticlepeer-review

22 Scopus citations


Several hundred PDZ (postsynaptic density-95, Drosophila disks-large, ZO-1) domain-containing proteins have been identified in the human genome. PDZ domains play a critical role in organization and function of cellular signaling pathways. Thus, small molecule inhibitors of PDZ domain association with their targets have wide potential applications as research and therapeutic agents. PDZ domains typically bind to a carboxyl-terminal tail of the target protein. Here we describe a high-throughput screening (HTS) assay for small molecule inhibitors of association between Mint1-PDZ domains and N-type Ca2+ channel carboxyl-terminal peptide (NC peptide). The performance of a homogeneous time-resolved fluorescence resonance energy transfer (HTRF) and an amplified luminescent proximity homogeneous assay (ALPHA) were systematically compared in parallel pilot HTS experiments with glutathione S-transferase-Mint1-PDZ1/2 protein and biotinylated NC peptide. Both of the two assays showed similar sensitivities in our target protein assay. Using HTRF-based assay we screened a library of 100,000 small molecule compounds and identified a number of potential "hits." The activity of isolated "hits" was confirmed by ALPHA assay. However, further evaluation revealed that isolated "hits" most likely act as "promiscuous binders," not as specific Mint-PDZ inhibitors, and that additional screening will be required to identify the true Mint-PDZ inhibitors. The assays described provided an example of HTS for a small molecule inhibitor of Mint-PDZ domain that can be easily adapted to other PDZ domain-mediated interactions.

Original languageEnglish (US)
Pages (from-to)769-783
Number of pages15
JournalAssay and Drug Development Technologies
Issue number6
StatePublished - Dec 1 2007

ASJC Scopus subject areas

  • Molecular Medicine
  • Drug Discovery


Dive into the research topics of 'High-throughput screen for small molecule inhibitors of Mint1-PDZ domains'. Together they form a unique fingerprint.

Cite this