Spectroscopic Characterization Of Spin-Labeled Magnetically Oriented Phospholipid Bilayers By EPR Spectroscopy

Document Type

Article

Publication Date

7-26-2000

Published In

Journal Of The American Chemical Society

Abstract

This paper reports the EPR spectroscopic characterization of a recently developed magnetically oriented spin-labeled model membrane system. The oriented membrane system is composed of a mixture of a bilayer forming phospholipid and a short chain phospholipid that breaks up the extended bilayers into bilayered micelles or bicelles that are highly hydrated (approximately 75% aqueous). Paramagnetic lanthanide ions (Tm3+) were added to align the bicelles such that the bilayer normal is collinear with the direction of the static magnetic field. Optimal bicelle alignment was obtained when the temperature was increased slowly (approximately 15 mini from 298 K (gel phase) to 318 K (L-alpha phase) at 0.64 T. The nitroxide spin probe 3 beta-doxyl-5 alpha-cholestane (cholestane) was used to demonstrate the effects of macroscopic bilayer alignment through the measurement of orientational dependent hyperfine splittings that were close to A(yy). The EPR signals of cholestane inserted into oriented and randomly dispersed DMPC-rich bilayers have been investigated over the temperature range 298-348 K. Also, the time dependence of the loss of orientation upon cessation of the magnetic field has been characterized. Power saturation EPR experiments indicate that for the sample compositions described here, the lanthanide ions do not induce spectral line broadening of the cholestane EPR signal in DMPC-rich Lipid bilayers. Recently, there has been a great deal of excitement over the use of magnetically oriented systems for both solution and solid-state NMR spectroscopy. This study demonstrates the feasibility of conducting bicelle experiments in the relatively low magnetic field of a conventional EPR spectrometer. The system offers the opportunity to carry out EPR studies using a well-oriented highly hydrated model membrane system whose preparation is amenable to the reconstitution of labile membrane components such as integral membrane proteins.

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