Residues in the seventh membrane-spanning segment of the dopamine D2 receptor accessible in the binding-site crevice

Biochemistry. 1996 Sep 3;35(35):11278-85. doi: 10.1021/bi960928x.


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. Using the substituted-cysteine accessibility method, we previously mapped the residues that form the surface of the binding-site crevice in the third and fifth membrane-spanning segments (M3 and M5). We have now mutated to cysteine, one at a time, 26 consecutive residues in and flanking the seventh membrane-spanning segment (M7) and expressed the mutant receptors in HEK 293 cells. Nine of these mutants reacted with charged, hydrophilic, lipophobic, sulfhydryl-specific reagents, added extracellularly, and were protected from reaction by a reversible dopamine antagonist, sulpiride. Thus, we infer that the side chains of these residues are in the water-accessible surface of the binding-site crevice. The pattern of accessibility of the cysteine-substitution mutants is consistent with M7 being a kinked alpha-helix.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Cell Line
  • Cysteine / genetics
  • Dopamine Antagonists / pharmacology
  • Ethyl Methanesulfonate / analogs & derivatives
  • Ethyl Methanesulfonate / metabolism
  • Ethyl Methanesulfonate / pharmacology
  • GTP-Binding Proteins / metabolism
  • Humans
  • Kinetics
  • Membrane Proteins / chemistry
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Receptors, Dopamine D2 / chemistry*
  • Receptors, Dopamine D2 / metabolism
  • Signal Transduction / physiology
  • Spiperone / analogs & derivatives
  • Spiperone / metabolism
  • Sulfhydryl Reagents / metabolism
  • Sulfhydryl Reagents / pharmacology
  • Sulpiride / pharmacology


  • Dopamine Antagonists
  • Membrane Proteins
  • Receptors, Dopamine D2
  • Sulfhydryl Reagents
  • methanethiosulfonate ethylammonium
  • Spiperone
  • Sulpiride
  • 3-N-methylspiperone
  • Ethyl Methanesulfonate
  • GTP-Binding Proteins
  • Cysteine