QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) All Versions of this Article: mol.106.028662v1 71/2/483    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Kloda, J. H. Articles by Czajkowski, C. Search for Related Content PubMed PubMed Citation Articles by Kloda, J. H. Articles by Czajkowski, C.

Agonist-, Antagonist-, and Benzodiazepine-Induced Structural Changes in the ₁ Met¹¹³-Leu¹³² Region of the GABA_A Receptor

Department of Physiology and Molecular and Cellular Pharmacology Program, University of Wisconsin-Madison, Madison, Wisconsin

The structural basis by which agonists, antagonists, and allosteric modulators exert their distinct actions on ligand-gated ion channels is poorly understood. We used the substituted cysteine accessibility method to probe the structure of the GABA_A receptor in the presence of ligands that elicit different pharmacological effects. Residues in the ₁ Met¹¹³-Leu¹³² region of the GABA binding site were individually mutated to cysteine and expressed with wild-type ₂ and ₂ subunits in Xenopus laevis oocytes. Using electrophysiology, we determined the rates of reaction of N-biotinaminoethyl methaneth-iosulfonate (MTSEA-biotin) with the introduced cysteines in the resting (unliganded) state and compared them with rates determined in the presence of GABA (agonist), 4-[6-imino-3-(4-methoxyphenyl)pyridazin-1-yl]butanoic acid hydrobromide (SR-95531; antagonist), pentobarbital (allosteric modulator), and flurazepam (allosteric modulator). ₁N115C, ₁L117C, ₁T129C, and ₁R131C are predicted to line the GABA binding pocket because MTSEA-biotin modification of these residues decreased the amount of current elicited by GABA, and the rates/extents of modification were decreased both by GABA and SR-95531. Reaction rates of some substituted cysteines were different depending on the ligand, indicating that barbiturate- and GABA-induced channel gating, antagonist binding, and benzodiazepine modulation induce specific structural rearrangements. Chemical reactivity of ₁E122C was decreased by either GABA or pentobarbital but was unaltered by SR-95531 binding, whereas ₁L127C reactivity was decreased by agonist and antagonist binding but not affected by pentobarbital. Furthermore, ₁E122C, ₁L127C, and ₁R131C changed accessibility in response to flurazepam, providing structural evidence that residues in and near the GABA binding site move in response to benzodiazepine modulation.

Address correspondence to: Dr. Cynthia Czajkowski, Department of Physiology, University of Wisconsin at Madison, 601 Science Drive, Madison, WI 53711. E-mail: czajkowski{at}physiology.wisc.edu

This article has been cited by other articles:

				L. M. Sharkey and C. Czajkowski Individually Monitoring Ligand-Induced Changes in the Structure of the GABAA Receptor at Benzodiazepine Binding Site and Non-Binding-Site Interfaces Mol. Pharmacol., July 1, 2008; 74(1): 203 - 212. [Abstract] [Full Text] [PDF]

				J. Mercado and C. Czajkowski {gamma}-Aminobutyric Acid (GABA) and Pentobarbital Induce Different Conformational Rearrangements in the GABAA Receptor {alpha}1 and {beta}2 Pre-M1 Regions J. Biol. Chem., May 30, 2008; 283(22): 15250 - 15257. [Abstract] [Full Text] [PDF]

				S. M. Hanson and C. Czajkowski Structural Mechanisms Underlying Benzodiazepine Modulation of the GABAA Receptor J. Neurosci., March 26, 2008; 28(13): 3490 - 3499. [Abstract] [Full Text] [PDF]

				C. L. Padgett and S. C. R. Lummis The F-loop of the GABAA Receptor {gamma}2 Subunit Contributes to Benzodiazepine Modulation J. Biol. Chem., February 1, 2008; 283(5): 2702 - 2708. [Abstract] [Full Text] [PDF]