SPECTROSCOPY, IMAGING, OTHER TECHNIQUES |
Optimizing Oriented Planar Supported Lipid Samples for Solid-State Protein NMR
Jan K. Rainey 1 and Brian D. Sykes 1*
1 University of Alberta
* To whom correspondence should be addressed. E-mail: brian.sykes{at}ualberta.ca.
Submitted on April 13, 2005
Revised on June 18, 2005
Accepted on 20 July 2005
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Abstract |
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Sample orientation relative to the static magnetic field of an NMR spectrometer allows study of membrane proteins in the lipid bilayer setting. The straightforward preparation and handling of extremely thin mica substrates with consistent surface properties has prompted us to examine oriented phospholipid bilayer and hexagonal phases on mica. The spectral characteristics of oriented lipid samples formed on mica are as good as or better than those on glass. Nine solvents with varying dielectric constant were used to cast lipid films or for vesicle spreading, film characteristics were compared, and static solid-state 31P-NMR was used to characterize the degree of orientation of the hydrated lipid species. Lipids with four headgroup chemistries were tested: 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG), 1,2-dioleoyl-sn-glycero-3-phosphate (DOPA), and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE). Solvent affected orientation of POPG, DOPA, and DOPE, but not POPC. Film characteristics varied with solvent, with ramifications for producing homogeneous oriented lipid samples. POPC was used to optimize the amount of lipid per substrate and compare hydration methods. POPG did not orient reproducibly, whereas POPG-POPC mixtures did. DOPA showed 1-2 oriented states depending upon hydration level and deposition method. DOPE formed oriented hexagonal phase which underwent a reversible temperature induced phase transition to the oriented bilayer phase.
Key Words:
hexagonal phase, membrane protein, muscovite mica, oriented sample, phospholipid bilayers and multilayers, solid-state NMR
