A highly conserved δ-opioid receptor region determines RGS4 interaction

FEBS J. 2020 Feb;287(4):736-748. doi: 10.1111/febs.15033. Epub 2019 Sep 9.


The δ-opioid receptor (δ-OR) couples to Gi/Go proteins to modulate a variety of responses in the nervous system. Τhe regulator of G protein signalling 4 (RGS4) was previously shown to directly interact within the C-terminal region of δ-OR using its N-terminal domain to negatively modulate opioid receptor signalling. Herein, using molecular dynamics simulations and in vitro pull-down experiments we delimit this interaction to 12 helix 8 residues of δ-ΟR and to the first 17 N-terminal residues (NT) of RGS4. Monitoring the complex arrangement and stabilization between RGS4 and δ-OR by molecular dynamics simulations combined with mutagenesis studies, we defined that two critical interactions are formed: one between Phe329 of helix8 of δ-ΟR and Pro9 of the NT of RGS4 and the other a salt bridge between Glu323 of δ-ΟR and Lys17 of RGS4. Our observations allow drafting for the first time a structural model of a ternary complex including the δ-opioid receptor, a G protein and a RGS protein. Furthermore, the high degree of conservation among opioid receptors of the RGS4-binding region, points to a conserved interaction mode between opioid receptors and this important regulatory protein.

Keywords: G proteins; RGS4; helix 8; molecular dynamics simulation; opioid receptors.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Cattle
  • Cloning, Molecular
  • Conserved Sequence
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • GTP-Binding Protein alpha Subunits, Gi-Go / chemistry*
  • GTP-Binding Protein alpha Subunits, Gi-Go / genetics
  • GTP-Binding Protein alpha Subunits, Gi-Go / metabolism
  • Gene Expression
  • Genetic Vectors / chemistry
  • Genetic Vectors / metabolism
  • Humans
  • Molecular Dynamics Simulation
  • Mutation
  • Protein Binding
  • Protein Conformation, alpha-Helical
  • Protein Conformation, beta-Strand
  • Protein Interaction Domains and Motifs
  • RGS Proteins / chemistry*
  • RGS Proteins / genetics
  • RGS Proteins / metabolism
  • Receptors, Opioid, delta / chemistry*
  • Receptors, Opioid, delta / genetics
  • Receptors, Opioid, delta / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Alignment
  • Sequence Homology, Amino Acid


  • RGS Proteins
  • Receptors, Opioid, delta
  • Recombinant Proteins
  • RGS4 protein
  • GNAI3 protein, human
  • GTP-Binding Protein alpha Subunits, Gi-Go