Spontaneous thermal motion of the GABAA receptor M2 channel-lining segments

Amal K. Bera, Myles H. Akabas

Research output: Contribution to journalArticle

16 Scopus citations

Abstract

The γ-aminobutyric acid type A (GABAA) receptor channel opening involves translational and rotational motions of the five channel-lining, M2 transmembrane segments. The M2 segment's extracellular half is loosely packed and undergoes significant thermal motion. To characterize the extent of the M2 segment's motion, we used disulfide trapping experiments between pairs of engineered cysteines. In α1β 1γ25 receptors the single γ subunit is flanked by an α and β subunit. The γ2M2-14′ position is located in the α-γ subunit interface. γ 213′ faces the channel lumen. We expressed either the γ214′ or the γ213′ cysteine substitution mutants with α1 cysteine substitution mutants between 12′ and 16′ and wild-type β1. Disulfide bonds formed spontaneously between γ214′C and both α115′C and α116′C and also between γ213′C and α113′C. Oxidation by copper phenanthroline induced disulfide bond formation between γ214′C and α1113′C. Disulfide bond formation rates with γ214′C were similar in the presence and absence of GABA, although the rate with α113′C was slower than with the other two positions. In a homology model based on the acetylcholine receptor structure, αM2 would need to rotate in opposite directions by ∼80° to bring α113′ and α115′ into close proximity with γ 214′. Alternatively, translational motion of αM2 would reduce the extent of rotational motion necessary to bring these two a subunit residues into close proximity with the γ214′ position. These experiments demonstrate that in the closed state the M2 segments undergo continuous spontaneous motion in the region near the extracellular end of the channel gate. Opening the gate may involve similar but concerted motions of the M2 segments.

Original languageEnglish (US)
Pages (from-to)35506-35512
Number of pages7
JournalJournal of Biological Chemistry
Volume280
Issue number42
DOIs
StatePublished - Oct 21 2005

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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