Prevalence, specificity and determinants of lipid-interacting PDZ domains from an in-cell screen and in vitro binding experiments

PLoS One. 2013;8(2):e54581. doi: 10.1371/journal.pone.0054581. Epub 2013 Feb 4.


Background: PDZ domains are highly abundant protein-protein interaction modules involved in the wiring of protein networks. Emerging evidence indicates that some PDZ domains also interact with phosphoinositides (PtdInsPs), important regulators of cell polarization and signaling. Yet our knowledge on the prevalence, specificity, affinity, and molecular determinants of PDZ-PtdInsPs interactions and on their impact on PDZ-protein interactions is very limited.

Methodology/principal findings: We screened the human proteome for PtdInsPs interacting PDZ domains by a combination of in vivo cell-localization studies and in vitro dot blot and Surface Plasmon Resonance (SPR) experiments using synthetic lipids and recombinant proteins. We found that PtdInsPs interactions contribute to the cellular distribution of some PDZ domains, intriguingly also in nuclear organelles, and that a significant subgroup of PDZ domains interacts with PtdInsPs with affinities in the low-to-mid micromolar range. In vitro specificity for the head group is low, but with a trend of higher affinities for more phosphorylated PtdInsPs species. Other membrane lipids can assist PtdInsPs-interactions. PtdInsPs-interacting PDZ domains have generally high pI values and contain characteristic clusters of basic residues, hallmarks that may be used to predict additional PtdInsPs interacting PDZ domains. In tripartite binding experiments we established that peptide binding can either compete or cooperate with PtdInsPs binding depending on the combination of ligands.

Conclusions/significance: Our screen substantially expands the set of PtdInsPs interacting PDZ domains, and shows that a full understanding of the biology of PDZ proteins will require a comprehensive insight into the intricate relationships between PDZ domains and their peptide and lipid ligands.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins
  • Binding Sites
  • Cell Line, Tumor
  • Genes, Reporter
  • High-Throughput Screening Assays
  • Humans
  • Immunoblotting
  • Kinetics
  • Ligands
  • Luminescent Proteins
  • Membrane Proteins / chemistry
  • Membrane Proteins / metabolism*
  • Models, Molecular
  • Molecular Sequence Data
  • PDZ Domains*
  • Peptides / chemistry
  • Peptides / metabolism*
  • Phosphatidylinositols / chemistry
  • Phosphatidylinositols / metabolism*
  • Protein Binding
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Surface Plasmon Resonance
  • Syntenins / chemistry
  • Syntenins / metabolism*


  • Bacterial Proteins
  • Ligands
  • Luminescent Proteins
  • Membrane Proteins
  • Peptides
  • Phosphatidylinositols
  • Recombinant Proteins
  • Syntenins
  • yellow fluorescent protein, Bacteria

Grants and funding

This work was supported by the Fund for Scientific Research - Flanders (FWO), the Cell Imaging Core and the Concerted Actions Program of K. U. Leuven, the Hercules Foundation (AKUL005 HER/08/061) and the Belgian Foundation against cancer. AMW was supported by a Ph.D. fellowship from FWO, YI was an EMBO long-term fellow and PZ was an EMBO young investigator. JPB is supported by La Ligue contre le Cancer (Label 2010). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.