PHASE-EQUILIBRIA AND MOLECULAR PACKING IN THE N,N-DIMETHYLDODECYLAMINE OXIDE GRAMICIDIN-D WATER-SYSTEM STUDIED BY H-2 NUCLEAR-MAGNETIC-RESONANCE SPECTROSCOPY

A partial phase diagram of the system N,N-dimethyldodecylamine oxide ( DDAO)/water/gramicidin D was determined by H-2-NMR. Both (H2O)-H-2 and perdeuterated DDAO (DDAO-d(31)) were studied by solid state NMR techn iques, Addition of gramicidin D to the micellar (L(1)), normal hexagon al (H-1) and cubic (1) phases of DDAO induces phase separations, givin g two-phase regions, which all contain a lamellar (L,) phase. The L, p hase containing gramicidin is characterized by larger order parameters for DDAO-d(31) compared with the corresponding order parameters in th e L(alpha) and H-1 phases of DDAO-d(31)/H2O. The L(alpha) phase may st ay in equilibrium with any other phase in the phase diagram. The DDAO exchange between the coexisting phases is slow on the NMR timescale, w hich is why the recorded NMR spectrum consists of superimposed spectra from the different phases occurring in the sample. Gramicidin D can b e solubilized in appreciable quantities only in the lamellar phase of DDAO-d(31). Increasing amounts of gramicidin in the liquid crystalline phases result in a continuous increase in the molecular ordering up t o about 5 mol% gramicidin, where a plateau is reached. This is consist ent with a recent theoretical model describing the influence on the or dering of lipids by a membrane protein with larger hydrophobic thickne ss than the lipid bilayer. The solvent used for dissolving gramicidin at the incorporation of the peptide in the lipid aggregates has no eff ect on the H-2-NMR lineshapes of DDAO-d(31). It is concluded that gram icidin is solubilized in the L alpha phase and that it always adopts t he channel conformation independent of a particular solvent. The chann el conformation is also supported by CD studies. In some of the sample s, macroscopic orientation of the lipid aggregates is observed. It is concluded that DDAO-d(31) in the binary system favors an orientation w ith the long axis of the hydrocarbon chain perpendicular to the magnet ic field, whereas when gramicidin D is present the hydrocarbon chain o rients parallel to the magnetic field. This is explained by the fact t hat gramicidin aligns with its helical axis parallel to the magnetic f ield, thereby forcing also the DDAO-d(31) molecules to obtain such an orientation.