Gamma-aminobutyric acid increases the water accessibility of M3 membrane-spanning segment residues in gamma-aminobutyric acid type A receptors

Biophys J. 1999 Nov;77(5):2563-74. doi: 10.1016/s0006-3495(99)77091-8.


Gamma-aminobutyric acid type A (GABA(A)) receptors are members of the ligand-gated ion channel gene superfamily. Using the substituted cysteine accessibility method, we investigated whether residues in the alpha(1)M3 membrane-spanning segment are water-accessible. Cysteine was substituted, one at a time, for each M3 residue from alpha(1)Ala(291) to alpha(1)Val(307). The ability of these mutants to react with the water-soluble, sulfhydryl-specific reagent pCMBS(-) was assayed electrophysiologically. Cysteines substituted for alpha(1)Ala(291) and alpha(1)Tyr(294) reacted with pCMBS(-) applied both in the presence and in the absence of GABA. Cysteines substituted for alpha(1)Phe(298), alpha(1)Ala(300), alpha(1)Leu(301), and alpha(1)Glu(303) only reacted with pCMBS(-) applied in the presence of GABA. We infer that the pCMBS(-) reactive residues are on the water-accessible surface of the protein and that GABA induces a conformational change that increases the water accessibility of the four M3 residues, possibly by inducing the formation of water-filled crevices that extend into the interior of the protein. Others have shown that mutations of alpha(1)Ala(291), a water-accessible residue, alter volatile anesthetic and ethanol potentiation of GABA-induced currents. Water-filled crevices penetrating into the interior of the membrane-spanning domain may allow anesthetics and alcohol to reach their binding sites and thus may have implications for the mechanisms of action of these agents.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • 4-Chloromercuribenzenesulfonate / metabolism
  • Amino Acid Substitution
  • Animals
  • Cell Membrane / metabolism*
  • Cysteine
  • Hydrophobic and Hydrophilic Interactions
  • Ion Channel Gating / drug effects
  • Mesylates / metabolism
  • Mutation
  • Protein Binding / drug effects
  • Protein Conformation / drug effects
  • Rats
  • Receptors, GABA-A / chemistry*
  • Receptors, GABA-A / genetics
  • Receptors, GABA-A / metabolism*
  • Sulfhydryl Compounds / metabolism
  • Surface Properties
  • Water / metabolism*
  • gamma-Aminobutyric Acid / pharmacology*


  • Mesylates
  • Receptors, GABA-A
  • Sulfhydryl Compounds
  • Water
  • methanethiosulfonate
  • gamma-Aminobutyric Acid
  • 4-Chloromercuribenzenesulfonate
  • Cysteine