Deuterium Nuclear Magnetic Resonance Studies of the Interaction between Dimyristoylphosphatidylcholine and Gramicidin A

Deuterium nuclear magnetic resonance spectra of dimyristoylphosphatidylcholines (DMPCs) specifically labeled with deuterium at one of positions 2‘, 3‘, 4‘, 6‘, 8‘, 10‘, 12‘, or 14‘ of the 2-chain have been recorded at 34.1 MHz in the presence of varying concentrations of the linear pentadecapeptide antibiotic gramicidin A‘. Deuterium quadrupole splittings (Δvq) have been used to partially characterize the motion and hydrocarbon chain order of phospholipid in contact with the polypeptide surface. At lipid concentrations below 4 lipids/gramicidin molecule the quadrupole splitting of a terminal methyl-labeled DMPC collapses to a single line. The quadrupole splittings of the methylene labels are decreased, and the line shapes are dominated by large intrinsic line widths. The time constants characterizing the decays of the echo intensity (T2e) are correspondingly reduced in the lipid-polypeptide complexes. At lipid-polypeptide molar ratios of greater than 15:1, the quadrupole splittings of the labels increase linearly with gramicidin concentration to a value ~30% greater than that of pure lipid. Between ratios of 15:1 and 4:1, the splittings decrease slowly. At a ratio of about 4:1 a rather abrupt transition occurs to the high gramicidin phase, and on the time scale of the deuterium NMR experiment (~5.0 μs) lipid adjacent to polypeptide appears disordered (smaller Δvq) compared to pure lipid. At all lipid― protein ratios investigated above the gel-to-liquid crystalline phase transition temperature of the pure lipid (Tc), the experimental spectrum is shown to be accurately simulated by using one quadrupole splitting together with a Lorentzian line broadening corresponding to the quadrupolar echo decay rate (πT2e)-1. In some gramicidin-lecithin complexes, T2e values as short as 49 μs are observed. This implies in general for studies of protein-lipid organization in both model and biological membranes that T2e values should be determined routinely in order to eliminate spectral distortions due to relaxation.