Investigations of polypeptide rotational diffusion in aligned membranes by 2H and 15N solid-state NMR spectroscopy

Transmembrane and in-plane oriented peptides have been prepared by solid-phase peptide synthesis, labeled with 3,3,3-H-2(3)-alanine and N-15-leucine at two selected sites, and reconstituted into oriented phophaticlylcholine membranes. Thereafter, proton-decoupled N-15 and H-2 solid-state NMR spectroscopy at sample orientations of the membrane normal parallel to the magnetic field direction have been used to characterize the tilt and rotational pitch angle of these peptides in some detail. In a second step the samples have been tilted by 90degrees. In this setup the spectral line shapes are sensitive indicators of the rate of rotational diffusion. Whereas monomeric transmembrane peptides exhibit spectral averaging and well-defined resonances, larger complexes are characterized by broad spectral line shapes. In particular the deuterium line shape is sensitive to association of a few transmembrane helices. In contrast, the formation of much larger complexes affects the N-15 chemical shift spectrum. The spectra indicate that in liquid crystalline membranes an amphipathic peptide of 14 amino acids exhibits fast rotational diffusion on both the H-2 and N-15 time scales (> 10(-5) s). Extending the sequences to 26 amino acids results in pronounced changes of the H-2 solid-state NMR spectrum, whereas the signal intensities of N-15 solid-state NMR spectra degrade. Below the phase transition temperature of the phospholipid bilayers, motional averaging on the time scale of the H-2 solid-state NMR spectrum ceases for transmembrane and in-plane oriented peptides. Furthermore at temperatures close to the phase transition the total signal intensities of the deuterium solid-state NMR spectra strongly decrease.