Mapping the Interface of a GPCR Dimer: A Structural Model of the A 2A Adenosine and D 2 Dopamine Receptor Heteromer.

The A2A adenosine (A2A R) and D2 dopamine (D2 R) receptors form oligomers in the cell membrane and allosteric interactions across the A2A R-D2 R heteromer represent a target for development of drugs against central nervous system disorders. However, understanding of the molecular determinants of A2A R-D2 R heteromerization and the allosteric antagonistic interactions between the receptor protomers is still limited. In this work, a structural model of the A2A R-D2 R heterodimer was generated using a combined experimental and computational approach. Regions involved in the heteromer interface were modeled based on the effects of peptides derived from the transmembrane (TM) helices on A2A R-D2 R receptor-receptor interactions in bioluminescence resonance energy transfer (BRET) and proximity ligation assays. Peptides corresponding to TM-IV and TM-V of the A2A R blocked heterodimer interactions and disrupted the allosteric effect of A2A R activation on D2 R agonist binding. Protein-protein docking was used to construct a model of the A2A R-D2 R heterodimer with a TM-IV/V interface, which was refined using molecular dynamics simulations. Mutations in the predicted interface reduced A2A R-D2 R interactions in BRET experiments and altered the allosteric modulation. The heterodimer model provided insights into the structural basis of allosteric modulation and the technique developed to characterize the A2A R-D2 R interface can be extended to study the many other G protein-coupled receptors that engage in heteroreceptor complexes.