Biology Reference
In-Depth Information
technique that can accurately monitor thermotropic phase behavior in lipid bilayers
[37
42]
.
From DSC one can obtain T
m
s, transition enthalpies, and information on phase. In general
this technique involves heating a reference solution devoid of the membrane of interest
and a sample containing a dilute aqueous subject membrane suspension. The technique
measures the difference in the amount of heat required to increase the temperature of the
sample and reference at a chosen rate. During the scan, the temperature of the sample and
reference is very accurately kept the same.
The phospholipid most studied by DSC has been DPPC (16:0,16:0 PC), the workhorse of
DSC membrane studies. This lipid is inexpensive and readily available in highly pure
form from a number of commercial sources. The lipid is resistant to oxidation (it has two
saturated acyl chains) and has a T
m
that is not near the troublesome ice transition (0
C),
nor is it so high as to make the lipid susceptible to massive heat-induced hydrolysis. A typical
DSC scan for DPPC is shown in
Figure 9.22
. The main transition occurs at 41.3
C (Table 5.1).
A much smaller transition, called the pre-transition, is observed at ~35.6
C. Since the pre-
transition is not found in all phospholipid classes, it probably is related to the polar head
group. For example DPPE (16:0,16:0 PE) has no pre-transition and has a T
m
of 63
C. Profound
differences in the DSC scans between DPPC and DPPE have been attributed to differences in
head group size (
Figure 9.23
). PC has a much larger and more hydrated head group than does
PE. As a result, at temperatures below T
m
(in the gel state), the diameter of the PC head group
(S) is wider than the combined diameters of the two all trans acyl chains (2
e
S
). Therefore
S
.The chains must then tilt relative to the head group until their combined diameters
are the same size, S
>
2
S
.Without tilting there would be a gap beneath the head that would
have to be filled by water, a thermodynamic disaster. The larger the head group, the larger is
the tilt angle. For example, in the gel state DMPC (14:0,14:0 PC) has a tilt angle of 12
while
the much larger cerebroside (with a sugar head group) has a tilt angle of 41
. For PCs, upon
melting the gauche kinks substantially increase the diameter of the acyl chains whereupon
their sum becomes equal to, or greater than the head diameter (S
¼
2
S
), eliminating the
tilt angle. For DPPE which has a small, poorly hydrated head group, in the gel state
S
¼
/
<
2
S
and the tilt angle is zero.
As depicted in
Figure 9.23
, gel state DPPC exhibits chain tilt, while DPPE does not. This
may explain why PCs have a lower T
m
than PEs. The chain tilt puts DPPC in a configuration
more amenable to melting. The typical DSC scan shown in
Figure 9.22
for DPPC shows both
the main chain melting transition and the minor pre-transition. Both transitions are believed
to be part of the same melting process. In the gel state (L
¼
2
S
'
), the chains of DPPC are tilted. The
tilt is lost upon transitioning into the melted liquid crystalline (L
b
) state. Something unusual
happens to DPPC in the temperature range spanning the pre-transition and main transition.
Fluorescence and ESR methodologies (discussed below and in Chapter 10, respectively) indi-
cate a co-existence of gel and fluid domains. More unusual, however, is the shape of the
membrane over this temperature range. At the pre-transition, the flat membrane in the gel
phase transforms into a periodically undulated bilayer, named the ripple or P
a
'
phase
(
Figure 9.23
). The ripple phase has been shown by electron density profiles obtained from
small angle X-ray scattering to be an asymmetric undulation pattern resembling a saw-tooth
with a wavelength of 120
b
160
˚
, depending on acyl chain length. It is not at all clear if ripple
structure has any biological importance as very small lipid 'contaminants,' including other
membrane lipids, eliminate the pre-transition and hence the ripple phase. A biological
e
