GABAA receptor M2-M3 loop secondary structure and changes in accessibility during channel gating

Amal K. Bera, Maya Chatav, Myles Akabas

Research output: Contribution to journalArticle

81 Citations (Scopus)

Abstract

The γ-aminobutyric acid type A (GABAA) receptor M2-M3 loop structure and its role in gating were investigated using the substituted cysteine accessibility method. Residues from α1Arg-273 to α1aIle-289 were mutated to cysteine, one at a time. MTSET+ or MTSES- reacted with all mutants from α1R273C to α1Y281C, except α1P277C, in the absence and presence of GABA. The MTSET+ closed-state reaction rate was >1000 liters/ mol-s at α1N274C, α1S275C, α1K278C, and α1Y281C and was <300 liters/mol-s at <1R273C, α1L276C, α1V279C, α1A280C, and α1A284C. These two groups of residues lie on opposite sides of an α-helix. The fast reacting group lies on a continuation of the M2 segment channel-lining helix face. This suggests that the M2 segment α-helix extends about two helical turns beyond α1N274 (20′), aligned with the extracellular ring of charge. At α1S275C, α1V279C, α1A280C, and α1A284C the reaction rate was faster in the presence of GABA. The reagents had no functional effect on the mutants from α1A282C to α1I289C, except α1A284C. Access may be sterically hindered possibly by close interaction with the extracellular domain. We suggest that the M2 segment α-helix extends beyond the predicted extracellular end of the M2 segment and that gating induces a conformational change in and/or around the N-terminal half of the M2-M3 loop. Implications for coupling ligand-evoked conformational changes in the extracellular domain to channel gating in the membrane-spanning domain are discussed.

Original languageEnglish (US)
Pages (from-to)43002-43010
Number of pages9
JournalJournal of Biological Chemistry
Volume277
Issue number45
DOIs
StatePublished - Nov 8 2002

Fingerprint

GABA-A Receptors
gamma-Aminobutyric Acid
Reaction rates
Cysteine
Aminobutyrates
Linings
Ligands
Membranes

ASJC Scopus subject areas

  • Biochemistry

Cite this

GABAA receptor M2-M3 loop secondary structure and changes in accessibility during channel gating. / Bera, Amal K.; Chatav, Maya; Akabas, Myles.

In: Journal of Biological Chemistry, Vol. 277, No. 45, 08.11.2002, p. 43002-43010.

Research output: Contribution to journalArticle

@article{db5ecea75dde4274821824103dfa53bd,
title = "GABAA receptor M2-M3 loop secondary structure and changes in accessibility during channel gating",
abstract = "The γ-aminobutyric acid type A (GABAA) receptor M2-M3 loop structure and its role in gating were investigated using the substituted cysteine accessibility method. Residues from α1Arg-273 to α1aIle-289 were mutated to cysteine, one at a time. MTSET+ or MTSES- reacted with all mutants from α1R273C to α1Y281C, except α1P277C, in the absence and presence of GABA. The MTSET+ closed-state reaction rate was >1000 liters/ mol-s at α1N274C, α1S275C, α1K278C, and α1Y281C and was <300 liters/mol-s at <1R273C, α1L276C, α1V279C, α1A280C, and α1A284C. These two groups of residues lie on opposite sides of an α-helix. The fast reacting group lies on a continuation of the M2 segment channel-lining helix face. This suggests that the M2 segment α-helix extends about two helical turns beyond α1N274 (20′), aligned with the extracellular ring of charge. At α1S275C, α1V279C, α1A280C, and α1A284C the reaction rate was faster in the presence of GABA. The reagents had no functional effect on the mutants from α1A282C to α1I289C, except α1A284C. Access may be sterically hindered possibly by close interaction with the extracellular domain. We suggest that the M2 segment α-helix extends beyond the predicted extracellular end of the M2 segment and that gating induces a conformational change in and/or around the N-terminal half of the M2-M3 loop. Implications for coupling ligand-evoked conformational changes in the extracellular domain to channel gating in the membrane-spanning domain are discussed.",
author = "Bera, {Amal K.} and Maya Chatav and Myles Akabas",
year = "2002",
month = "11",
day = "8",
doi = "10.1074/jbc.M206321200",
language = "English (US)",
volume = "277",
pages = "43002--43010",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "45",

}

TY - JOUR

T1 - GABAA receptor M2-M3 loop secondary structure and changes in accessibility during channel gating

AU - Bera, Amal K.

AU - Chatav, Maya

AU - Akabas, Myles

PY - 2002/11/8

Y1 - 2002/11/8

N2 - The γ-aminobutyric acid type A (GABAA) receptor M2-M3 loop structure and its role in gating were investigated using the substituted cysteine accessibility method. Residues from α1Arg-273 to α1aIle-289 were mutated to cysteine, one at a time. MTSET+ or MTSES- reacted with all mutants from α1R273C to α1Y281C, except α1P277C, in the absence and presence of GABA. The MTSET+ closed-state reaction rate was >1000 liters/ mol-s at α1N274C, α1S275C, α1K278C, and α1Y281C and was <300 liters/mol-s at <1R273C, α1L276C, α1V279C, α1A280C, and α1A284C. These two groups of residues lie on opposite sides of an α-helix. The fast reacting group lies on a continuation of the M2 segment channel-lining helix face. This suggests that the M2 segment α-helix extends about two helical turns beyond α1N274 (20′), aligned with the extracellular ring of charge. At α1S275C, α1V279C, α1A280C, and α1A284C the reaction rate was faster in the presence of GABA. The reagents had no functional effect on the mutants from α1A282C to α1I289C, except α1A284C. Access may be sterically hindered possibly by close interaction with the extracellular domain. We suggest that the M2 segment α-helix extends beyond the predicted extracellular end of the M2 segment and that gating induces a conformational change in and/or around the N-terminal half of the M2-M3 loop. Implications for coupling ligand-evoked conformational changes in the extracellular domain to channel gating in the membrane-spanning domain are discussed.

AB - The γ-aminobutyric acid type A (GABAA) receptor M2-M3 loop structure and its role in gating were investigated using the substituted cysteine accessibility method. Residues from α1Arg-273 to α1aIle-289 were mutated to cysteine, one at a time. MTSET+ or MTSES- reacted with all mutants from α1R273C to α1Y281C, except α1P277C, in the absence and presence of GABA. The MTSET+ closed-state reaction rate was >1000 liters/ mol-s at α1N274C, α1S275C, α1K278C, and α1Y281C and was <300 liters/mol-s at <1R273C, α1L276C, α1V279C, α1A280C, and α1A284C. These two groups of residues lie on opposite sides of an α-helix. The fast reacting group lies on a continuation of the M2 segment channel-lining helix face. This suggests that the M2 segment α-helix extends about two helical turns beyond α1N274 (20′), aligned with the extracellular ring of charge. At α1S275C, α1V279C, α1A280C, and α1A284C the reaction rate was faster in the presence of GABA. The reagents had no functional effect on the mutants from α1A282C to α1I289C, except α1A284C. Access may be sterically hindered possibly by close interaction with the extracellular domain. We suggest that the M2 segment α-helix extends beyond the predicted extracellular end of the M2 segment and that gating induces a conformational change in and/or around the N-terminal half of the M2-M3 loop. Implications for coupling ligand-evoked conformational changes in the extracellular domain to channel gating in the membrane-spanning domain are discussed.

UR - http://www.scopus.com/inward/record.url?scp=0037044793&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0037044793&partnerID=8YFLogxK

U2 - 10.1074/jbc.M206321200

DO - 10.1074/jbc.M206321200

M3 - Article

VL - 277

SP - 43002

EP - 43010

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 45

ER -