The structure of an integral membrane peptide: a deuterium NMR study of gramicidin.
Solid state deuterium NMR was employed on oriented multilamellar dispersions consisting of 1,2-dilauryl-sn-glycero-3-phosphatidylcholine and deuterium (2H) exchange-labeled gramicidin D, at a lipid to protein molar ratio (L/P) of 15:1, in order to study the dynamic structure of the channel conformation of gramicidin in a liquid crystalline phase. The corresponding spectra were used to discriminate between several structural models for the channel structure of gramicidin (based on the left- and right-handed beta 6.3 LD helix) and other models based on a structure obtained from high resolution NMR. The oriented spectrum is complicated by the fact that many of the doublets, corresponding to the 20 exchangeable sites, partially overlap. Furthermore, the asymmetry parameter, eta, of the electric field gradient tensor of the amide deuterons is large (approximately 0.2) and many of the amide groups are involved in hydrogen bonding, which is known to affect the quadrupole coupling constant. In order to account for these complications in simulating the spectra in the fast motional regime, an ab initio program called Gaussian 90 was employed, which permitted us to calculate, by quantum mechanical means, the complete electric field gradient tensor for each residue in gramicidin (using two structural models). Our results indicated that the left-handed helical models were inconsistent with our observed spectra, whereas a model based on the high-resolution structure derived by Arseniev and coworkers, but relaxed by a simple energy minimization procedure, was consistent with our observed spectra. The molecular order parameter was then estimated from the motional narrowing assuming the relaxed (right-handed) Arseniev structure. Our resultant order parameter of SZZ = 0.91 translates into an rms angle of 14 degrees, formed by the helix axis and the local bilayer normal. The strong resemblance between our spectra (and also those reported for gramicidin in 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) multilayers) and the spectra of the same peptide incorporated in a lyotropic nematic phase, suggests that the lyotropic nematic phase simulates the local environment of the lipid bilayer.
Prosser
R S
RS
Department of Physics, University of Guelph, Ontario, Canada.
Daleman
S I
SI
Davis
J H
JH
D000595:Amino Acid Sequence; D055592:Biophysical Phenomena; D001703:Biophysics; D002942:Circular Dichroism; D003903:Deuterium; D006096:Gramicidin / Q000737:chemistry*; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D008051:Lipid Bilayers / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010713:Phosphatidylcholines / Q000737:chemistry; D011487:Protein Conformation
D008051:Lipid Bilayers; D008565:Membrane Proteins; D010713:Phosphatidylcholines; D006096:Gramicidin; C015153:1,2-dilauroylphosphatidylcholine; D003903:Deuterium