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UBC Theses and Dissertations

Integral protein and cholesterol in model membranes : a ²H NMR study Nezil, Frank Arthur


Deuterium (²H) nuclear magnetic resonance (NMR) is used to study bilayer hydrophobic thickness and thickness expansivity in La phase lipid bilayer membranes containing combinations of POPC , cholesterol, and synthetic amphiphillic polypeptides of different hydrophobic lengths. Subsequent analysis relates ²H NMR thickness expansivity measurements to micromechanical measurements of the area expansivity, άA. Model membrane area expansivity to date has seldom been considered i n typical analyses of NMR data. We study a case, using ²H and 31P NMR , in which considerations of άA are central: spontaneous vesiculation i n model membranes composed of POPC and POPS , with and without cholesterol. Subsequent freeze-fracture electron microscopy confirms that membrane eruptions occur in response to positive temperature changes and upon addition of, for instance, myelin basic protein (MBP) . Eruptions are shown to occur more readily in the presence of cholesterol. Estimations of άA based upon NMR data in these systems is found to be in agreement with micromechanical measurements of άA. Orientation-dependent spin-spin (T2) relaxation in some of the above systems potentially confounds the interpretation of apparent thickness changes as being due to inclusion of integral polypeptides. A procedure is outlined which allows empirical correction of distorsion in inhomogeneously broadened NMR spectra due to orientation-dependent relaxation processes. This is then applied to data of previous studies, thereby ruling out T2 distorsion as an artifactuaL "cause" of previously described changes in bilayer hydrophobic thickness. The method is then used to suggest some directions for future study.

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