Internal dynamics of the nicotinic acetylcholine receptor in reconstituted membranes.
The structure and 1H/2H exchange kinetics of affinity-purified nAChR reconstituted into egg phosphatidylcholine membranes with increasing levels of either dioleoylphosphatidic acid (DOPA) or cholesterol (Chol) have been examined using infrared spectroscopy. All spectra of the reconstituted nAChR membranes recorded after 72 h in 2H2O exhibit comparable amide I band shapes, suggesting a similar secondary structure for the nAChR in each lipid environment. Increasing levels of either DOPA or Chol, however, lead to an increasing intensity of the amide II band, indicating a decreasing proportion of nAChR peptide hydrogens that have exchanged for deuterium. Spectra recorded as a function of time after exposure of the nAChR to 2H2O show that the presence of either lipid slows down the 1H/2H exchange of those peptide hydrogens that normally exchange on the minutes to hours time scale. The slowing of peptide 1H/2H exchange correlates with both an increasing ability of the nAChR to undergo agonist-induced conformational change [Baenziger, J. E., Morris, M.-L., Darsaut, T. E., and Ryan, S. E. (1999) in preparation] and possibly a decreasing membrane fluidity. Our data suggest that lipid composition dependent changes in nAChR peptide 1H/2H exchange kinetics reflect altered internal dynamics of the nAChR. Lipids may influence protein function by changing the internal dynamics of integral membrane proteins.
Baenziger
J E
JE
Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada.
Darsaut
T E
TE
Morris
M L
ML
D000818:Animals; D002784:Cholesterol / Q000737:chemistry; D017666:Deuterium Oxide / Q000737:chemistry; D004295:Dihydroxyphenylalanine / Q000737:chemistry; D007700:Kinetics; D008560:Membrane Fluidity; D008563:Membrane Lipids / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D010713:Phosphatidylcholines / Q000737:chemistry; D011487:Protein Conformation; D011978:Receptors, Nicotinic / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D014101:Torpedo
D008563:Membrane Lipids; D010455:Peptides; D010713:Phosphatidylcholines; D011978:Receptors, Nicotinic; D004295:Dihydroxyphenylalanine; D002784:Cholesterol; D017666:Deuterium Oxide