Effect of staphylococcal delta-lysin on the thermotropic phase behavior and vesicle morphology of dimyristoylphosphatidylcholine lipid bilayer model membranes. Differential scanning calorimetric, P-31 nuclear magnetic resonance and Fourier transform infrared spectroscopic, and X-ray diffraction studies

We investigated the effects of various concentrations of staphylococcal del ta-lysin on the thermotropic phase behavior of large multilamellar dimyrist oylphosphatidylcholine (DMPC) vesicles by differential scanning calorimetry (DSC), P-31 nuclear magnetic resonance (NMR) and Fourier transform infrare d (FTIR) spectroscopy, and X-ray diffraction. The DSC studies revealed that at all concentrations, the addition of delta-lysin progressively decreases the enthalpy of the pretransition of DMPC bilayers without significantly a ffecting its temperature or cooperativity. Similarly, the addition of small er quantities of peptide has little effect on the temperature of the main p hase transition of DMPC bilayers but does reduce the cooperativity and enth alpy of this transition somewhat. However, at higher peptide concentrations , a second phase transition with a slightly increased temperature and a mar kedly reduced cooperativity and enthalpy is also induced, and this latter p hase transition resolves itself into two components at the highest peptide concentrations that are tested. Moreover, our P-31 NMR spectroscopic studie s reveal that at relatively low delta-lysin concentrations, essentially all of the phospholipid molecules produce spectra characteristic of the lamell ar phase, whereas at the higher peptide concentrations, an increasing propo rtion exhibit an isotropic signal. Also, at the highest delta-lysin concent rations that are. studied, the isotropic component of the P-31 NMR spectrum also resolves itself into two components. At the highest peptide concentra tion that was tested, we are also able to effect a macroscopic separation o f our sample into two fractions by centrifugation, a pellet containing rela tively smaller amounts of delta-lysin and a supernatant containing larger a mounts of peptide relative to the amount of lipid present. We are also able to show that the more cooperative phase transition detected calorimetrical ly, and the lamellar phase P-31 NMR signal, arise from the pelleted materia l, while the less cooperative phase transition and the isotropic P-31 NMR s ignal arise from the supernatant. In addition, we demonstrate by X-ray diff raction that the pelleted material corresponds to delta-lysin-containing la rge multilamellar vesicles and the supernatant to a mixture of delta-lysin- containing small unilamellar vesicles and discoidal particles. We also show by FTIR spectroscopy that delta-lysin exists predominantly in the alpha-he lical conformation in aqueous solution or when interacting with DMPC, and t hat a large fraction of the peptide bonds undergo H-D exchange in D2O, Howe ver, upon interaction with DMPC, the fraction of exchangeable amide protons decreases. We also demonstrate by this technique that both of the phase tr ansitions detected by DSC correspond to phospholipid hydrocarbon chain-melt ing phase transitions. Finally, we show by several techniques that. the abs olute concentrations of delta-lysin and the thermal history, as well as the lipid:peptide ratio, can affect the thermotropic phase behavior and morpho logy of peptide-lipid aggregates.