Solid-state nuclear magnetic resonance derived model for dynamics in the polypeptide backbone of the gramicidin A channel.

The dynamics of the backbone of the gramicidin A transmembrane cation channel in dimyristoylphosphatidylcholine bilayers have been investigated using solid state 15N nuclear magnetic resonance (n.m.r.) spectroscopy. With the temperature-dependent fluidity of the bilayer, the rates of motions in the helical gramicidin channel can be modulated. It is shown that in the gel phase, all substantial motions of the channel are slow on the timescale of the n.m.r. experiment (3.5 kHz). The use of oriented samples in which the axis of global channel rotation is aligned parallel to the magnetic field enables separation of global and local dynamics. Spectra obtained from oriented bilayer samples containing single-site 15N-labeled gramicidin at 8 degrees C are analyzed to yield a spatial model for local backbone motion. This model includes the axis of motion, the mean orientation, and the maximum amplitude of displacement for individual peptide planes. Specific sites in the first turn of the amino terminus were investigated, with emphasis on the Ala3 and Leu4 linkages, for which the orientation of the 15N chemical shift tensor with respect to the molecular frame has been determined. The effect of two well-characterized bilayer defect structures, parabolic focal conics and oily streaks, is included in the spectral simulations. It is found that only relatively small amplitude motions are possible at the two sites, with amplitudes of not more than +/- 8 degrees and +/- 15 degrees for the Ala3 and Leu4 sites, respectively. Detailed characterization of the bilayer surface geometry in the oriented samples is presently the major limiting factor in the use of this technique for probing the spatial extent of local motions in integral membrane proteins.