Characterization of the A2AR-D2R interface: focus on the role of the C-terminal tail and the transmembrane helices

Biochem Biophys Res Commun. 2010 Nov 26;402(4):801-7. doi: 10.1016/j.bbrc.2010.10.122. Epub 2010 Oct 30.

Abstract

A single serine point mutation (S374A) in the adenosine A(2A) receptor (A(2A)R) C-terminal tail reduces A(2A)R-D(2)R heteromerization and prevents its allosteric modulation of the dopamine D(2) receptor (D(2)R). By means of site directed mutagenesis of the A(2A)R and synthetic transmembrane (TM) α-helix peptides of the D(2)R we further explored the role of electrostatic interactions and TM helix interactions of the A(2A)R-D(2)R heteromer interface. We found evidence that the TM domains IV and V of the D(2)R play a major role in the A(2A)R-D(2)R heteromer interface since the incubation with peptides corresponding to these domains significantly reduced the ability of A(2A)R and D(2)R to heteromerize. In addition, the incubation with TM-IV or TM-V blocked the allosteric modulation normally found in A(2A)R-D(2)R heteromers. The mutation of two negatively charged aspartates in the A(2A)R C-terminal tail (D401A/D402A) in combination with the S374A mutation drastically reduced the physical A(2A)R-D(2)R interaction and lost the ability of antagonistic allosteric modulation over the A(2A)R-D(2)R interface, suggesting further evidence for the existence of an electrostatic interaction between the C-terminal tail of A(2A)R and the intracellular loop 3 (IL3) of D(2)R. On the other hand, molecular dynamic model and bioinformatic analysis propose that specific AAR, AQE, and VLS protriplets as an important motive in the A(2A)R-D(2L)R heteromer interface together with D(2L)R TM segments IV/V interacting with A(2A)R TM-IV/V or TM-I/VII.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation
  • Amino Acid Sequence
  • Aspartic Acid / chemistry
  • Aspartic Acid / genetics
  • Cell Line
  • Humans
  • Molecular Sequence Data
  • Mutation
  • Protein Multimerization
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Receptor, Adenosine A2A / chemistry*
  • Receptor, Adenosine A2A / genetics
  • Receptors, Dopamine D2 / chemistry*
  • Receptors, Dopamine D2 / genetics
  • Serine / chemistry
  • Serine / genetics

Substances

  • Receptor, Adenosine A2A
  • Receptors, Dopamine D2
  • dopamine D2L receptor
  • Aspartic Acid
  • Serine