Solid-state 2H NMR spectroscopy

For mechanically aligned samples, peptides were incorporated into lipid bilayers, as previously described (12), with a peptide/lipid molar ratio of 1:60 using dilauroylphosphatidylcholine (DLPC), dimyristoylphosphatidylcholine (DMPC), or dioleoylphosphatidylcholine (DOPC) from Avanti Polar Lipids (Alabaster, AL) and a final hydration of 45% w/w using deuterium-depleted water from Cambridge Isotope Laboratories. Bilayer alignment in the liquid-crystalline samples was confirmed by ³¹P NMR using a Bruker (Billerica, MA) Avance 300 spectrometer with broadband ¹H decoupling. Sample orientations at both β = 0° (bilayer normal parallel to the magnetic field) and β = 90° were tested. A quadrupole echo pulse sequence was utilized with full phase cycling (30) at 50°C at both sample orientations. The quadrupole echo delay was 4.5 μs, and the recycle delay was 90 ms. Between 0.7 and 1.5 million scans were acquired for each ²H NMR experiment, and the spectra were processed with 150 Hz line broadening.

For experiments in which magnetically aligned bicelle samples containing ²H-labeled peptides were examined, the samples were prepared as described below.