Benzodiazepines induce a conformational change in the region of the γ-aminobutyric acid type a receptor α1-subunit M3 membrane-spanning segment

Daniel B. Williams, Myles Akabas

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

Benzodiazepine binding to γ-aminobutyric acid type A (GABA(A)) receptors allosterically modulates GABA binding and increases the currents induced by submaximal GABA concentrations. Benzodiazepines induce conformational changes in the GABA-binding site in the extracellular domain, but it is uncertain whether these conformational changes extend into the membrane-spanning domain where the channel gate is located. Alone, benzodiazepines do not open the channel. We used the substituted-cysteine-accessibility method to investigate diazepam-induced conformational changes in the region of the α1-subunit M3 membrane-spanning segment. In the absence of diazepam or GABA, pCMBS- did not react at a measurable rate with cysteine-substitution mutants between α1 Phe296 and α1Glu303. In the presence of 100 nM diazepam, pCMBS- reacted with α1F296C, α1F298C, and α1L301C but not with the other cysteine mutants between α1Phe296 and α1Glu303. These three mutants are a subset of the five residues that we previously showed reacted with pCMBS- applied in the presence of GABA. The pCMBS- reaction rates with these three cysteine mutants were similar in the presence of diazepam and GABA. Thus, diazepam, which binds to the extracellular domain, induces a conformational change in the membrane-spanning domain that is similar to a portion of the change induced by GABA. Because diazepam does not open the channel, these results provide structural evidence that the diazepam-bound state represents an intermediate conformation distinct from the open and resting/closed states of the receptor. The diazepam-induced conformational change in the M3 segment vicinity may be related to the mechanism of allosteric potentiation.

Original languageEnglish (US)
Pages (from-to)1129-1136
Number of pages8
JournalMolecular Pharmacology
Volume58
Issue number5
StatePublished - 2000

Fingerprint

Aminobutyrates
Diazepam
Benzodiazepines
gamma-Aminobutyric Acid
Membranes
Cysteine
GABA-A Receptors
Binding Sites

ASJC Scopus subject areas

  • Pharmacology

Cite this

@article{bbb9e9ac69ba4244818d3b65fbe97122,
title = "Benzodiazepines induce a conformational change in the region of the γ-aminobutyric acid type a receptor α1-subunit M3 membrane-spanning segment",
abstract = "Benzodiazepine binding to γ-aminobutyric acid type A (GABA(A)) receptors allosterically modulates GABA binding and increases the currents induced by submaximal GABA concentrations. Benzodiazepines induce conformational changes in the GABA-binding site in the extracellular domain, but it is uncertain whether these conformational changes extend into the membrane-spanning domain where the channel gate is located. Alone, benzodiazepines do not open the channel. We used the substituted-cysteine-accessibility method to investigate diazepam-induced conformational changes in the region of the α1-subunit M3 membrane-spanning segment. In the absence of diazepam or GABA, pCMBS- did not react at a measurable rate with cysteine-substitution mutants between α1 Phe296 and α1Glu303. In the presence of 100 nM diazepam, pCMBS- reacted with α1F296C, α1F298C, and α1L301C but not with the other cysteine mutants between α1Phe296 and α1Glu303. These three mutants are a subset of the five residues that we previously showed reacted with pCMBS- applied in the presence of GABA. The pCMBS- reaction rates with these three cysteine mutants were similar in the presence of diazepam and GABA. Thus, diazepam, which binds to the extracellular domain, induces a conformational change in the membrane-spanning domain that is similar to a portion of the change induced by GABA. Because diazepam does not open the channel, these results provide structural evidence that the diazepam-bound state represents an intermediate conformation distinct from the open and resting/closed states of the receptor. The diazepam-induced conformational change in the M3 segment vicinity may be related to the mechanism of allosteric potentiation.",
author = "Williams, {Daniel B.} and Myles Akabas",
year = "2000",
language = "English (US)",
volume = "58",
pages = "1129--1136",
journal = "Molecular Pharmacology",
issn = "0026-895X",
publisher = "American Society for Pharmacology and Experimental Therapeutics",
number = "5",

}

TY - JOUR

T1 - Benzodiazepines induce a conformational change in the region of the γ-aminobutyric acid type a receptor α1-subunit M3 membrane-spanning segment

AU - Williams, Daniel B.

AU - Akabas, Myles

PY - 2000

Y1 - 2000

N2 - Benzodiazepine binding to γ-aminobutyric acid type A (GABA(A)) receptors allosterically modulates GABA binding and increases the currents induced by submaximal GABA concentrations. Benzodiazepines induce conformational changes in the GABA-binding site in the extracellular domain, but it is uncertain whether these conformational changes extend into the membrane-spanning domain where the channel gate is located. Alone, benzodiazepines do not open the channel. We used the substituted-cysteine-accessibility method to investigate diazepam-induced conformational changes in the region of the α1-subunit M3 membrane-spanning segment. In the absence of diazepam or GABA, pCMBS- did not react at a measurable rate with cysteine-substitution mutants between α1 Phe296 and α1Glu303. In the presence of 100 nM diazepam, pCMBS- reacted with α1F296C, α1F298C, and α1L301C but not with the other cysteine mutants between α1Phe296 and α1Glu303. These three mutants are a subset of the five residues that we previously showed reacted with pCMBS- applied in the presence of GABA. The pCMBS- reaction rates with these three cysteine mutants were similar in the presence of diazepam and GABA. Thus, diazepam, which binds to the extracellular domain, induces a conformational change in the membrane-spanning domain that is similar to a portion of the change induced by GABA. Because diazepam does not open the channel, these results provide structural evidence that the diazepam-bound state represents an intermediate conformation distinct from the open and resting/closed states of the receptor. The diazepam-induced conformational change in the M3 segment vicinity may be related to the mechanism of allosteric potentiation.

AB - Benzodiazepine binding to γ-aminobutyric acid type A (GABA(A)) receptors allosterically modulates GABA binding and increases the currents induced by submaximal GABA concentrations. Benzodiazepines induce conformational changes in the GABA-binding site in the extracellular domain, but it is uncertain whether these conformational changes extend into the membrane-spanning domain where the channel gate is located. Alone, benzodiazepines do not open the channel. We used the substituted-cysteine-accessibility method to investigate diazepam-induced conformational changes in the region of the α1-subunit M3 membrane-spanning segment. In the absence of diazepam or GABA, pCMBS- did not react at a measurable rate with cysteine-substitution mutants between α1 Phe296 and α1Glu303. In the presence of 100 nM diazepam, pCMBS- reacted with α1F296C, α1F298C, and α1L301C but not with the other cysteine mutants between α1Phe296 and α1Glu303. These three mutants are a subset of the five residues that we previously showed reacted with pCMBS- applied in the presence of GABA. The pCMBS- reaction rates with these three cysteine mutants were similar in the presence of diazepam and GABA. Thus, diazepam, which binds to the extracellular domain, induces a conformational change in the membrane-spanning domain that is similar to a portion of the change induced by GABA. Because diazepam does not open the channel, these results provide structural evidence that the diazepam-bound state represents an intermediate conformation distinct from the open and resting/closed states of the receptor. The diazepam-induced conformational change in the M3 segment vicinity may be related to the mechanism of allosteric potentiation.

UR - http://www.scopus.com/inward/record.url?scp=0033789013&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033789013&partnerID=8YFLogxK

M3 - Article

VL - 58

SP - 1129

EP - 1136

JO - Molecular Pharmacology

JF - Molecular Pharmacology

SN - 0026-895X

IS - 5

ER -