HIGH PRESSURE X-RAY SURFACE DIFFRACTION ON HIGHLY ALIGNED LIPID SYSTEMS UNDER EXCESS WATER CONDITIONS.
M.Kriechbaum(1), M. Steinhart(2), S. Bernstorff(3) and P. Laggner(1).
A hydrostatic high-pressure
X-ray cell  was modified to perform studies on highly aligned
lamellar lipid systems. With this experimental set-up at the
SAXS-beamline at ELETTRA, Trieste, Italy,
simultaneously wide- and small-angle X-ray scattering (SAXS) studies to elucidate the complex morphology of
these systems were possible. In contrast to isotropic bilayer systems (usually liposomal preparations, also
called "powder samples") highly aligned bilayer membrane systems give the possibility to derive
additional structural information by diffraction.
Aligned samples allow the clear differentiation of in-plane and out-of-plane correlation as well as
the unique identification of the Miller indices in higher (>1) dimensional phases. However, aligning
them under physiological conditions (i.e. excess water, ionic strength and physiological pH) has turned out
to be a difficult task, e.g. the "vapor pressure paradox" [2, 3].
Using this modified high pressure cell, studies on these systems under physiological conditions,
pressures between 0 - 3000 bar and temperatures between 0 and 80 °C can be performed.
Furthermore the cell offers the possibility to do p-jump experiments on these systems with
jump amplitudes of up to 3 kbar/10 ms.
In Fig.1 the 2-dimensional diffraction pattern of the aligned phospholipid
(DMPC) obtained at 20 °C and ambient pressure is shown.
It clearly exhibits the in-plane reflection from the Pb´ phase after cooling down from the La phase,
which is lamellar and has no in-plane reflections (see e.g. [4-6]).
The real space model of the Pb´ phase is given in Fig.2. For the first time
the set-up allowed to perform the La - Pb´ phase transition on aligned lipids using pressure as
the thermodynamical variable increasing the pressure from 0 to 1080 bar at a temperature of
30 °C. To compare the results the d-spacings obtained at different temperatures and pressures
are compiled in Table 1.
|Phase of DMPC
|Pb´ cooling from 30°C
|Pb´ pressurizing from 1 bar
Table 1. Liquid crystalline and stable ripple phase of
Diffraction pattern of a highly aligned lipid (DMPC) in the Pb´ phase at constant grazing angle
at 20 °C. Letter A shows the in-plane reflections due to the ripple structure (distance lambda),
letter B gives the off-plane reflections due to the lamellar repeat distance d (see Fig.2).
C denotes the specular reflectivity peak.
Schematic sketch showing asymmetric rippled bilayer, separated by a layer of water. The unit cell
(shown in red) is given by the dimensions d and lambda and the monoclinic angle gamma.
 Steinhart, M., Kriechbaum, M., Pressl, K., Amenitsch, H., Laggner, P. & Bernstorff, S. (1999),
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 Katsaras, J., Watson, M.J. (2000), Rev. Sci. Instrum. 71, 1737.
 Tardieu, A., Luzzati, V., Reman, F.C. (1973), J.Mol.Biol. 75, 711.
 Wack, D.C., Webb, W.W., (1989), Phys.Rev. A 40, 2712.
 Katsaras, J., et.al. (2000), Phys.Rev. E 61, 5668.
|Author: Heinz Amenitsch
firstname.lastname@example.org (Sept.18th 2000) |