Structure of both the ligand- and lipid-dependent channel-inactive states of the nicotinic acetylcholine receptor probed by FTIR spectroscopy and hydrogen exchange.
FTIR spectra have been recorded both as a function of time and after prolonged exposure to 2H2O buffer in order to study the structural changes that lead to both the ligand- and lipid-dependent channel-inactive states of the nicotinic acetylcholine receptor (nAChR). The hydrogen/deuterium exchange spectra provide insight into both the overall rates and extent of peptide 1H/2H exchange and the individual rates and extent to which peptide hydrogens in alpha-helix and beta-sheet conformations exchange for deuterium. The spectra are also sensitive to the conformation of the polypeptide backbone and thus the secondary structure of the nAChR. The various spectral features monitored in the presence and absence of carbamylcholine and tetracaine are essentially identical, indicating that there are no large net changes in secondary structure in the channel-inactive desensitized state. The various spectral features monitored for the nAChR reconstituted into lipid membranes either with or without cholesterol are very similar, indicating that cholesterol is not a major structural regulator of the nAChR. However, in the absence of both cholesterol and anionic lipids, there is a slightly enhanced rate of exchange of alpha-helical peptide hydrogens for deuterium that occurs as a result of either an increase in nAChR dynamics or an increase in the accessibility of transmembrane peptide hydrogens to 2H2O. The latter may simply be due to an increase in the "fluidity" and thus permeability of the lipid bilayers to aqueous solvent.(ABSTRACT TRUNCATED AT 250 WORDS)
Méthot
N
N
Department of Biochemistry, University of Ottawa, Ontario, Canada.
Demers
C N
CN
Baenziger
J E
JE
D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D002784:Cholesterol / Q000494:pharmacology; D003903:Deuterium; D006859:Hydrogen; D006860:Hydrogen Bonding; D007473:Ion Channels / Q000737:chemistry* / Q000187:drug effects; D008051:Lipid Bilayers / Q000737:chemistry; D008563:Membrane Lipids / Q000494:pharmacology*; D010712:Phosphatidic Acids / Q000494:pharmacology; D010713:Phosphatidylcholines / Q000494:pharmacology; D017433:Protein Structure, Secondary; D011978:Receptors, Nicotinic / Q000737:chemistry*; D013053:Spectrophotometry; D017550:Spectroscopy, Fourier Transform Infrared; D013748:Tetracaine / Q000494:pharmacology; D014101:Torpedo
D007473:Ion Channels; D008051:Lipid Bilayers; D008563:Membrane Lipids; D010712:Phosphatidic Acids; D010713:Phosphatidylcholines; D011978:Receptors, Nicotinic; D013748:Tetracaine; C037657:dioleoylphosphatidic acid; D006859:Hydrogen; D002217:Carbachol; D002784:Cholesterol; D003903:Deuterium; C017251:1,2-oleoylphosphatidylcholine