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|Title:||A Deuterium Nuclear Magnetic Resonance Study of Protein Dynamics and Protein-Lipid Interactions in Model and Biological Membranes (Nmr)|
|Author(s):||Kintanar, Agustin Balagot|
|Department / Program:||Chemistry|
|Degree Granting Institution:||University of Illinois at Urbana-Champaign|
|Abstract:||The dynamics of the Halobacterium halobium purple membrane protein, bacteriorhodopsin; and the motional order of the fluorescent lipid probe, diphenylhexatriene, in various model membrane systems, have been investigated by deuterium nuclear magnetic resonance (NMR) spectroscopy. ('2)H NMR spectra and spin-lattice relaxation times of selectively-deuterated amino acids in the crystalline solid state and in biosynthetically enriched bacteriorhodopsin, have been obtained. Analysis of the data has led to the determination of the type and rate of amino acid motion in these systems. The motions of amino acid residues are shown to be very sensitive to packing considerations. Large differences in both the type and rate of motion are observed for a particular amino acid in various crystal lattice forms and in the membrane protein. Moreover, motional heterogeneity is found to occur in some crystal lattice forms and especially in bacteriorhodopsin. The major types of motion that are observed include methyl group rotation (by three-fold hops), phenyl ring flips (180(DEGREES) hops), ring libration, chain libration, side-chain hops, and isotropic reorientation. The last two motions are seen only in the membrane protein. All of these motions can occur at widely different rates depending on the packing environment, but in bacteriorhodopsin, these rates are generally greater than 10('6) sec('-1).
('2)H NMR spectra of selectively-deuterated diphenylhexatriene in dimyristoylphosphatidylcholine multilamellar bilayers in the liquid crystalline phase, as a function of temperature and membrane composition, have also been obtained. The data have been analyzed in terms of order parameters of the probe molecules, and for the case of pure lipid bilayers and cholesterol-containing systems, are shown to agree with order parameters determined from fluorescence depolarization. In membranes containing the polypeptide gramicidin, however, the ('2)H NMR data reveal complex changes in the order parameter that are not observed by fluorescence techniques. These differences are attributed to the different timescales of the methods. The faster fluorescence technique cannot resolve motions that occur in the range of 10('-6)-10('-7) sec, and which are critical to the full understanding of protein-lipid interactions.
Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1984.
|Date Available in IDEALS:||2014-12-15|