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Differential polarized phase fluorometric studies of phospholipid bilayers under high hydrostatic pressure
Authors:Joseph R Lakowicz  Richard B Thompson
Institution:University of Maryland, School of Medicine, Department of Biological Chemistry, Baltimore, MD 21201 U.S.A.
Abstract:Differential polarized phase fluorometry was used to quantify the rotational rate (R) and limiting anisotropy (r) of the membrane probe diphenylhexatriene (DPH) in solvents and lipid vesicles exposed to hydrostatic pressures ranging from 1 bar to 2 kbar. These measurements reveal the effect of pressure on the phase-transition temperatures of the phosphatidylcholine vesicles, and the effects of pressure on order parameter of the acyl side-chain region of the membranes, the latter as indicated by r. In addition to the well-known elevation of the transition temperature (Tc) with pressure, our results demonstrate that increased pressure restores the order of the bilayers to that representative of temperatures below the transition temperature. We also found that solvents which allowed free isotropic rotation of DPH at 1 bar no longer allowed free rotation when sufficiently compressed; moreover, the apparent DPH rotational rate increased with r. Pressure studies using both DPH and the charged DPH analogue, trimethylammonium DPH (TMA-DPH) indicated that the Tc of dipalmitoylphosphatidylcholine vesicles increased 23 K/kbar and an apparent volume change of 0.036 ml/mol lipid at the phase transition. Assuming, as has been proposed, that TMA-DPH is localized near the glycerol backbone region of the bilayers, these results indicate a similar temperature- and pressure-dependent phase transition in this region and the acyl side-chain region of the membrane.
Keywords:Phase fluorimetry  High pressure  Membrane probe  Phase transition  (Phospholipid bilayer)  DPH  1  6-diphenyl-1  3  5-hexatriene  TMADPH  1-(4-(trimethylamino)phenyl)-6-phenylhexyl-1  3  5-triene  DOPC  DMPC  DPPC  dioleoyl-  dimyristoyl-  fluorescence lifetime by the phase shift demodulation methods  respectively
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