Investigations of the orientation of a membrane peptide by sum frequency spectroscopy

Sum frequency spectra of gramicidin A at the air/water interface in partially deuterated dimyristoylphosphatildycholine monolayers were collected as a function of surface pressure and mole fraction of the peptide. Reorganization of the lipid and peptide molecules were followed between 5 and 35 mN/m. Selective deuteration of the lipid enabled independent observation of the ordering of the lipid tail and headgroup regions. The results from pure lipid monolayer studies revealed that both portions of the amphiphile rearranged roughly in tandem as a function of pressure. When the membrane peptide was added to the monolayer, changes in its orientation were noted at mole fractions below 0.5, but not above this value. Specifically, two sets of data for the peptide could be obtained. One was for the aliphatic residues and the other for the Trp residues as each gave a distinct sum frequency signal in the CH stretch range. The aliphatic residues, which are dispersed throughout gramicidin, revealed that the whole peptide could facilely track changes in lipid orientation at 0.1 mole fraction but did so less readily at 0.4 mole fraction. On the other hand, the Trp residues, which all reside near the C-terminus and interact strongly with the lipid headgroups, showed a greater propensity to track lipid reorientation at the higher mole fraction. These results indicated that changes in the C-terminus probably precede realignment in other parts of the membrane peptide at higher peptide concentration. Furthermore, peptide-peptide interactions probably inhibit gramicidin A reorientation.