Mapping the binding-site crevice of the dopamine D2 receptor by the substituted-cysteine accessibility method

Neuron. 1995 Apr;14(4):825-31. doi: 10.1016/0896-6273(95)90226-0.


The binding site of the dopamine D2 receptor, like that of other homologous G protein-coupled receptors, is contained within a water-accessible crevice formed among its seven membrane-spanning segments. We have developed a method to map systematically all the residues forming the surface of this binding-site crevice, and we have applied this method to the third membrane-spanning segment (M3). We mutated, one at a time, 23 residues in and flanking M3 to cysteine and expressed the mutant receptors heterologously. Ten of these mutants reacted with charged, hydrophilic, lipophobic, sulfhydryl-specific reagents, added extracellularly, and were protected from reaction by a reversible dopamine antagonist. Thus, the side chains of these residues are exposed in the binding-site crevice, which like M3 extends from the extracellular to the intracellular side of the membrane. The pattern of exposure is consistent with a short loop followed by six turns of an alpha helix.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Binding Sites
  • Cell Line
  • Cysteine* / chemistry
  • Cysteine* / genetics
  • Embryo, Mammalian
  • Ethyl Methanesulfonate / analogs & derivatives
  • Ethyl Methanesulfonate / chemistry
  • Humans
  • Indicators and Reagents
  • Kidney
  • Mesylates / chemistry
  • Mutagenesis, Site-Directed
  • Peptide Mapping / methods*
  • Protein Structure, Secondary
  • Quaternary Ammonium Compounds / chemistry
  • Receptors, Dopamine D2 / chemistry*
  • Receptors, Dopamine D2 / metabolism
  • Transfection


  • Indicators and Reagents
  • Mesylates
  • Quaternary Ammonium Compounds
  • Receptors, Dopamine D2
  • methanethiosulfonate ethylammonium
  • trimethylethylammonium
  • methanethiosulfonate
  • Ethyl Methanesulfonate
  • Cysteine