Investigation of membrane-disruption by peptides using solid-state NMR techniques.

In this thesis, mechanically aligned lipid bilayers were used to investigate the importance of membrane composition in determining the behavior of amphipathic, alpha-helical peptides in lipid bilayers. Mechanically aligned lipid bilayers are often used as model membranes to obtain structural details of membrane-associated molecules, such as peptides, proteins, drugs, and cholesterol. However, well-aligned bilayer samples can be challenging to prepare, particularly those containing phosphatidylethanolamine lipids and hydrophobic components. A new method for producing mechanically aligned lipid bilayer samples using naphthalene, a sublimable solid, was developed and the utility of the naphthalene procedure was demonstrated on bilayers of many different compositions, including the previously mentioned difficult components. These results showed that the naphthalene procedure is a generally applicable method of preparing mechanically aligned samples for NMR spectroscopy. Using this newly developed procedure, several peptides were studied including the carboxy-amide of pardaxin (P1a) and MSI-78. 31P NMR experiments exploring the effect of P1a on lipid bilayers showed that P1a significantly disrupts lipid bilayers composed solely of zwitterionic lipids, causing the formation of an isotropic phase in POPE bilayers and a narrowing of the 31P CSA span in POPC bilayers; the presence of anionic lipids (POPG) or cholesterol reduced the peptide's ability to disrupt these bilayers. 31P NMR of samples combining MSI-78 and POPE or POPC showed that this peptide did not induce the formation of non-lamellar phases up to 15 mol% in POPE, but caused significant changes in the POPC bilayers, particularly at high peptide concentrations. At lower concentrations (1--5%), the peptide altered the morphology of the bilayer in a way consistent with the formation of a toroidal pore. Higher concentrations of peptide (10--15%) led to the formation of a mixture of normal hexagonal phase and lamellar phase lipids. The results from our studies of peptides demonstrate the importance of membrane composition in shaping the behavior of antimicrobial peptides in lipid bilayers.PhDBiochemistryBiological SciencesBiophysicsPhysical chemistryPure SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/123187/2/3068878.pd