Discovery and mapping of an intracellular antagonist binding site at the chemokine receptor CCR2

Research output: Contribution to journalJournal articleResearchpeer-review

  • Annelien J M Zweemer
  • Julia Bunnik
  • Margo Veenhuizen
  • Fabiana Miraglia
  • Eelke B Lenselink
  • Maris Vilums
  • Henk de Vries
  • Arthur Gibert
  • Stefanie Thiele
  • Rosenkilde, Mette
  • Adriaan P IJzerman
  • Laura H Heitman

The chemokine receptor CCR2 is a G protein-coupled receptor that is involved in many diseases characterized by chronic inflammation, and therefore a large variety of CCR2 small molecule antagonists has been developed. On the basis of their chemical structures these antagonists can roughly be divided into two groups with most likely two topographically distinct binding sites. The aim of the current study was to identify the binding site of one such group of ligands, exemplified by three allosteric antagonists, CCR2-RA-[R], JNJ-27141491, and SD-24. We first used a chimeric CCR2/CCR5 receptor approach to obtain insight into the binding site of the allosteric antagonists and additionally introduced eight single point mutations in CCR2 to further characterize the putative binding pocket. All constructs were studied in radioligand binding and/or functional IP turnover assays, providing evidence for an intracellular binding site for CCR2-RA-[R], JNJ-27141491, and SD-24. For CCR2-RA-[R] the most important residues for binding were found to be the highly conserved tyrosine Y(7.53) and phenylalanine F(8.50) of the NPxxYx(5,6)F motif, as well as V(6.36) at the bottom of TM-VI and K(8.49) in helix-VIII. These findings demonstrate for the first time the presence of an allosteric intracellular binding site for CCR2 antagonists. This contributes to an increased understanding of the interactions of diverse ligands at CCR2 and may allow for a more rational design of future allosteric antagonists.

Original languageEnglish
JournalMolecular Pharmacology
Issue number4
Pages (from-to)358-68
Number of pages11
Publication statusPublished - 2014

    Research areas

  • Allosteric Site, Amino Acid Motifs, Amino Acid Sequence, Animals, CHO Cells, COS Cells, Cell Line, Tumor, Cercopithecus aethiops, Cricetinae, Cricetulus, Humans, Imidazoles, Ligands, Molecular Sequence Data, Point Mutation, Protein Binding, Pyrrolidines, Receptors, CCR2, Sulfonamides

ID: 137817839