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Multiple binding sites for the general anesthetic isoflurane identified in the nicotinic acetylcholine receptor transmembrane domain.

ABSTRACT: An extensive search for isoflurane binding sites in the nicotinic acetylcholine receptor (nAChR) and the proton gated ion channel from Gloebacter violaceus (GLIC) has been carried out based on molecular dynamics (MD) simulations in fully hydrated lipid membrane environments. Isoflurane introduced into the aqueous phase readily partitions into the lipid membrane and the membrane-bound protein. Specifically, isoflurane binds persistently to three classes of sites in the nAChR transmembrane domain: (i) An isoflurane dimer occludes the pore, contacting residues identified by previous mutagenesis studies; analogous behavior is observed in GLIC. (ii) Several nAChR subunit interfaces are also occupied, in a site suggested by photoaffinity labeling and thought to positively modulate the receptor; these sites are not occupied in GLIC. (iii) Isoflurane binds to the subunit centers of both nAChR alpha chains and one of the GLIC chains, in a site that has had little experimental targeting. Interpreted in the context of existing structural and physiological data, the present MD results support a multisite model for the mechanism of receptor-channel modulation by anesthetics.

SUBMITTER: Brannigan G

PROVIDER: S-EPMC2922517 | biostudies-literature | 2010 Aug

REPOSITORIES: biostudies-literature

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Multiple binding sites for the general anesthetic isoflurane identified in the nicotinic acetylcholine receptor transmembrane domain.

Brannigan Grace G LeBard David N DN Hénin Jérôme J Eckenhoff Roderic G RG Klein Michael L ML

Proceedings of the National Academy of Sciences of the United States of America 20100726 32

An extensive search for isoflurane binding sites in the nicotinic acetylcholine receptor (nAChR) and the proton gated ion channel from Gloebacter violaceus (GLIC) has been carried out based on molecular dynamics (MD) simulations in fully hydrated lipid membrane environments. Isoflurane introduced into the aqueous phase readily partitions into the lipid membrane and the membrane-bound protein. Specifically, isoflurane binds persistently to three classes of sites in the nAChR transmembrane domain: ...[more]

PMID: 20660787

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Project description:Photoreactive derivatives of the general anesthetic etomidate have been developed to identify their binding sites in ?-aminobutyric acid, type A and nicotinic acetylcholine receptors. One such drug, [(3)H]TDBzl-etomidate (4-[3-(trifluoromethyl)-3H-diazirin-3-yl]benzyl-[(3)H]1-(1-phenylethyl)-1H-imidazole-5-carboxylate), acts as a positive allosteric potentiator of Torpedo nACh receptor (nAChR) and binds to a novel site in the transmembrane domain at the ?-? subunit interface. To extend our understanding of the locations of allosteric modulator binding sites in the nAChR, we now characterize the interactions of a second aryl diazirine etomidate derivative, TFD-etomidate (ethyl-1-(1-(4-(3-trifluoromethyl)-3H-diazirin-3-yl)phenylethyl)-1H-imidazole-5-carboxylate). TFD-etomidate inhibited acetylcholine-induced currents with an IC(50) = 4 ?M, whereas it inhibited the binding of [(3)H]phencyclidine to the Torpedo nAChR ion channel in the resting and desensitized states with IC(50) values of 2.5 and 0.7 mm, respectively. Similar to [(3)H]TDBzl-etomidate, [(3)H]TFD-etomidate bound to a site at the ?-? subunit interface, photolabeling ?M2-10 (?Ser-252) and ?Met-295 and ?Met-299 within ?M3, and to a site in the ion channel, photolabeling amino acids within each subunit M2 helix that line the lumen of the ion channel. In addition, [(3)H]TFD-etomidate photolabeled in an agonist-dependent manner amino acids within the ? subunit M2-M3 loop (?Ile-288) and the ? subunit transmembrane helix bundle (?Phe-232 and ?Cys-236 within ?M1). The fact that TFD-etomidate does not compete with ion channel blockers at concentrations that inhibit acetylcholine responses indicates that binding to sites at the ?-? subunit interface and/or within ? subunit helix bundle mediates the TFD-etomidate inhibitory effect. These results also suggest that the ?-? subunit interface is a binding site for Torpedo nAChR negative allosteric modulators (TFD-etomidate) and for positive modulators (TDBzl-etomidate).

Dataset Information

Multiple binding sites for the general anesthetic isoflurane identified in the nicotinic acetylcholine receptor transmembrane domain.

Publications

Multiple binding sites for the general anesthetic isoflurane identified in the nicotinic acetylcholine receptor transmembrane domain.

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