Biology Reference
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over which several important biophysical techniques, commonly employed in membrane
studies, can be used. Of these techniques, the fastest is Raman
¼
IR
>
ESR
¼
Fluorescence
Polarization
NMR. There are, however, modifications of each of these techniques that
can substantially expand the normal range to much faster times. The technique must match
the question being addressed. For example, take the case of one well-studied, fundamental
membrane problem that is discussed in more detail in Chapter 10. Early ESR studies indi-
cated that lipids in immediate contact with membrane proteins (termed annular lipids)
have different properties than the bulk bilayer lipids. However, according to NMR, all lipids
are the same and therefore annular lipids do not exist. The reason for this discrepancy is that
ESR detects events that are ten times faster than can be observed by the slower NMR.
>
B. MEMBRANE THICKNESS
Although not fully appreciated at the time, the thickness of a biological membrane was
accurately determined by Hugo Fricke in 1925 [1] . Using trans-membrane electrical measure-
ments, Fricke correctly determined that the hydrophobic center of a membrane was ~33 ˚
thick. This was in close agreement with the early pioneers working on lipid monolayers,
Rayleigh, Pokels, and Langmuir, who determined the thickness of a triolein lipid monolayer
was ~13
16 ˚ (Chapter 2). Doubling this value to account for membrane lipids existing in
bilayers, reported in 1925 by Gorter and Grendel, yields a correct hydrophobic membrane
interior of ~26
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32 ˚ . Fricke's failure to appreciate the importance of his own measurements
stemmed from his lack of understanding of the bilayer. It was not until 1957, when Robertson
imaged the membrane using electron microscopy (see Chapter 8), that the full membrane
span of about 75 ˚ , including lipid bilayer and protein, was established.
Two other important methods that have been used to determine membrane thickness
(planar bimolecular lipid membrane analysis and X-ray diffraction) are considered below.
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Planar Bimolecular Lipid Membranes
For the decades between 1925 and 1960, the lipid bilayer was believed to be an essential
feature of membranes. However, it was not clear if the lipid bilayer was stable enough to
actually exist. Several investigators, including Irwin Langmuir in ~1937, tried but failed to
make a lipid bilayer using the reverse process involved in making soap bubbles. Whereas
a lipid bilayer is water
lipid bilayer
water, a soap bubble is air
inverse lipid bilayer
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air ( Figure 9.3 ). Fascination with soap bubbles is centuries old and its early scientific inves-
tigation is summarized in a classic 1911 topic by C.V. Boys [2] . In 1961 Mueller and
co-workers reported the first preparation of a large, stable lipid bilayer they termed the
planar bimolecular lipid membrane or BLM [3] . They found that BLMs could be made
from a variety of polar lipids dissolved in organic solvents. Originally these investigators
used a chloroform/methanol/brain lipid extract since they were interested in the nerve
process of excitability. The solution was spread over a submerged 1
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2 mm hole in a Teflon
cup ( Figure 9.4 ) by use of a small artist's brush or later by a syringe. The hole was monitored
by 90 degree reflected light through a small telescope. After application of the lipid solution,
the hole appeared gray, but soon developed many bands of color, the result of partial
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