Biology Reference
In-Depth Information
membranes the term 'fluidity' is complex, technique-dependent, and quantifies a variety of
parameters including:
1.
Rotational correlation times
2.
Order parameters
3.
Steady state anisotropy
4.
Partitioning of probes between the membrane and water
Although a number representing 'fluidity' can be obtained, its precise definition may be
a matter of contention. Further complicating the problem is the enormous number of
membrane probes that are commercially available. The major provider of membrane probes
has for many years been Molecular Probes in Eugene, Oregon (now owned by Invitrogen,
Carlsbad, CA). Chapter 13 of the Molecular Probes catalog, entitled 'Probes for Lipids and
Membranes', provides an excellent review of the topic.
It is clear from several different types of measurements that 'fluidity' gradients exist from
the aqueous interface through to the bilayer interior. This has often been demonstrated using
fluorescence polarization as well as ESR. Both of these techniques require adding an aniso-
tropic bulky, and hence perturbing, probe at different locations down the acyl chain. Both
FP and ESR show a gradual, continuous increase in 'fluidity' from the carbons near the
a
or carboxylate end to the omega or terminal carbon
[54,55]
. Unfortunately, the bulkiness
of the fluoro or spin probes disturbs the chains to such an extent that important detail of
the 'fluidity' gradient can be lost.
NMR has been used to monitor the 'fluidity' gradient without perturbing the system.
Without getting bogged down in experimental details, two types of NMR techniques (order
parameter (
Figure 9.32
) and T
1
relaxation measurements) (
Figure 9.33
) have demonstrated
that 'fluidity' increases at a steady but slow rate for about the first 8 to 10 carbons (called
the plateau region), after which 'fluidity' progressively increases through the rest of the
chain. This pattern is characteristic of all bilayers. However, details concerning the size
and location of the plateau and the 'fluidity' of the most fluid terminal region vary depending
on the lipid composition of the bilayer. In fact the shape of the 'fluidity' gradient can provide
a crude fingerprint of the bilayer. The NMR-derived order parameter (S
CD
) can be used to
estimate how isotropic the motion is at a particular carbon. A low S
CD
indicates a large range
of motion and hence high 'fluidity'. The range of values can vary depending on what is being
measured, but often range from ~0 (no motion) to ~1.0 (isotropic motion).
Figure 9.32
pres-
ents S
CD
values for the sn-1 palmitoyl chain of DPPC and POPC (16:0,18:1 PC). The typical
trans-membrane 'fluidity' gradient pattern with a plateau region can be easily seen but it
is also clear that the two profiles, while having a similar general shape, do exhibit differences.
For example, the plateau region of POPC is more pronounced than for DPPC. The profile
shown for DPPC was obtained at a relatively high temperature of 67
C. At a reduced temper-
ature the plateau region would be longer and more pronounced.
Fast molecular motion can be followed by T
1
relaxation using NMR. A large T
1
indicates
more rapid motion.
Figure 9.33
shows T
1
s for various carbons on DPPC. Again a substantial
plateau region is evident. The viscosity varies about 70 fold from the highly fluid omega
methyl terminus to carbons on the less 'fluid' polar head group.
The biophysical techniques used to study membrane 'fluidity' are best suited to
compare changes in membrane physical properties brought about by environmental and
