Investigating Cholesterol Orientation in Lipid Bilayer by Raman Spectroscopy
Demers, Steven Meyer
Hafner, Jason H
Doctor of Philosophy
Biological membranes are composed of numerous types of molecules ranging from different sterols to phospholipids to membrane partitioning proteins. While the overall compositions of these various elements are understood in part, how the phospholipid bilayer in cells accommodates the different processes for the copious types of proteins is not fully understood. Cholesterol however has been proven to play a key role in animal membrane functions. By using gold nanorods as a substrate, lipid membranes encapsulated the nanorods for the lipid structure to be analyzed by SERS. Optical excitation of gold nanoparticles at their size and shape-dependent plasmon resonant frequency induces strong oscillations of the nanoparticle’s free electron gas, leading to surfaced enhanced Raman scattering (SERS) – an enhancement of Raman scattering signals with a molecular scale distance dependence from the gold surface. The gold nanorods used for this project were tuned to an excitation laser wavelength of 785 nm. From previous lipid membrane studies, dioleoylphosphatidylcholine (DOPC) was demonstrated to have a well ordered, gold supported bilayer. Once a lipid bilayer was exchanged to the gold nanorod surface, a lipid solution interspersed with cholesterol was exchanged to the surface. The presence of cholesterol was confirmed in SERS by additional Raman peaks being observed that were characteristic of only cholesterol. Structural analysis by the surfaced enhanced Raman scattering (SABERS) method combines SERS and unenhanced Raman spectra with theoretical calculations of the optical field and molecular polarizability. Raman measurements of the samples are orientationally averaged while SERS spectra contain information on molecular position and orientation relative to the surface. Together these reveal the molecular orientation and position of cholesterol in phospholipid bilayers. This method offers an approach to analyzing lipid membrane molecular structure under ambient conditions, with microscopic quantities, and without molecular labels.