Chap. 11A. Biological Membranes &
Transport
• The Composition and Architecture of Membranes
• Membrane Dynamics
• Solute Transport Across Membranes
Fig. 11-3. Fluid mosaic model for plasma
membrane structure.
Intro. to Membranes & Transport
Membranes set the external boundaries of cells and control the
molecular traffic across those boundaries. They also divide the
internal space of eukaryotic cells into different compartments (Fig.
11-1). Membranes also play important roles in energy production
and cell-cell communication. Membranes are flexible (e.g., allow
amoeboid movement of cells), self-sealing (e.g., reform on
membrane fission and fusion events), and selectively permeable to
polar solutes (e.g., retain/exclude wanted and unwanted solutes).
Most biological membranes contain many proteins involved in solute
transport, reception of extracellular signals, mediation of cell-cell
contacts, synthesis of membrane lipids and proteins, and energy
transductions. We first will cover the
composition and chemical architecture
of biological membranes. Second we
will cover the dynamic features of
membranes. Third, we will present an
in-depth discussion of solute transport
through protein transporters and ion
channels.
Lipid & Protein Components of Membranes I
The major molecular components of membranes include polar lipids,
membrane proteins, and carbohydrates attached to glycoproteins
and glycolipids. Each type of membrane has its characteristic lipid
and protein abundance (Table 11-1). Gross compositional
differences are reflective of the biological roles of membranes. For
example, myelin sheath membranes, which play an electrical
insulator role about neurons in animals, are enriched in lipids. Many
other membranes actually contain more protein than lipid, reflecting
their many roles in solute transport, catalysis, signaling, etc.
Lipid & Protein Components of Membranes II
Each cell in each species in all three
domains of life has a characteristic
composition of lipids in its membrane(s).
The lipid compositions of the plasma
membrane and organelle membranes of a
rat hepatocyte are given in Fig. 11-2.
In hepatocytes, the plasma membrane is
enriched in cholesterol due to its close
contact with lipoproteins in the blood.
On the other hand, cholesterol makes
up only a small percentage of
mitochondrial membrane lipids. In most
cases, the functional significance of
variations in membrane lipid composition
remain unknown. Lastly, the protein
composition of membranes varies even
more widely than the lipid composition
and is determined by the functional
specialization of the membrane.
Fluid Mosaic Model for Membrane Structure
Physical studies of the permeability and motions of individual
proteins and lipids within membranes led to the seminal concept of
the “fluid mosaic” model for membrane structure. In this model,
proteins form the ceramic tiles that float in the lipid mortar. The
whole structure is held together by noncovalent interactions and
the hydrophobic effect. Due to the lack of covalent interactions
between components, both lipids and proteins are able to undergo
lateral diffusion in the bilayer. The acyl chains of membrane lipids
are maintained in their melted states to allow lateral diffusion of
components. On the other hand, movement of lipids, and
particularly proteins, from one
bilayer leaflet to the other is
restricted. The carbohydrate
moieties of both glycolipids and
glycoproteins face outside the
cell, and the two sides of the
bilayer are said to be
asymmetrical. We will further
discuss the various features of
the fluid mosaic properties of
membranes, and will note some
exceptions and refinements.
Amphipathic Lipid Aggregates in Water
The major amphipathic structural lipids of membranes-glycerophospholipids, sphingolipids, and sterols--form microscopic
aggregates when placed in water. The most common structure
formed is the bilayer, which is unstable unless its edges are curved
around and sealed as in vesicles (liposomes). Bilayers form for the
above lipids whose head groups and acyl chains have roughly the
same cross-sectional areas. Lipids such as free fatty acids and
detergents such as SDS form spherical micelles since the crosssectional areas of their head groups are larger than their acyl
chains. The formation of these structures is driven by the
hydrophobic effect. The lipid bilayer is about 30Å thick. The
hydrocarbon core is about as nonpolar as decane. Pure lipid vesicles
made in the laboratory are essentially impermeable to polar solutes.
Asymmetric Distribution of Phospholipids in
Cell Membranes
Plasma membrane lipids are
asymmetrically distributed between
the two leaflets of the bilayer.
However, the asymmetry is not
absolute as it is for membrane
proteins. The distribution of
phospholipids between the inner and
outer monolayers (leaflets) of the
plasma membrane of erythrocytes is
shown in Fig. 11-5. The table shows
that the choline-containing lipids,
phosphatidylcholine and sphingomyelin,
are typically found in the outer
leaflet. In contrast, the amino group
containing phospholipids,
phosphatidylethanolamine and
phosphatidylserine, along with other
phospholipids are primarily found in
the inner leaflet. The transbilayer
distribution of phospholipids is
maintained by specific proteins (see
below).
Distribution of Lipids in a Typical Eukaryotic
Cell
Each membrane in a cell has its
own characteristic composition. As
membrane vesicles transport
components through the cell from
the ER to the Golgi, then on to
the plasma membrane, changes
occur in vesicular lipid composition
and the distribution across the
membranes. For instance,
phosphatidylcholine is the major
phospholipid in the lumenal leaflet
of the Golgi, and vesicles moving
to the trans-Golgi network.
However, in transport vesicles
moving from the trans-Golgi network to the plasma membrane,
phosphatidylcholine is largely replaced by sphingolipids and
cholesterol, which enter the outer leaflet of the plasma
membrane on fusion of the transport vesicles with the plasma
membrane. In some cells, changes in membrane lipid distribution
have functional consequences. For example, phosphatidylserine
must move to the outer leaflet of the platelet plasma membrane
for a platelet to participate in blood coagulation.
Classification of Membrane Proteins
Membrane proteins are classified into
three broad categories that relate to
conditions needed to remove them from
bilayers. Integral membrane proteins are
deeply embedded in the bilayer and have
multiple hydrophobic amino acid residues
in contact with bilayer lipids. They can
only be removed from the membrane by
reagents, such as detergents, that
disrupt hydrophobic interactions between
the protein and membrane lipids and coat
the hydrophobic domain with a micellar
like phase. Peripheral membrane proteins
are associated with the membrane
through electrostatic and hydrogen
bonding interactions to other membrane
proteins or the polar head groups of
phospholipids. They can readily be
removed from the membrane by changing
the pH or ionic strength of the solution.
Finally, amphitrophic protein attachment
to membranes is regulated by a biological
modification such as attachment of a lipid
anchor or phosphorylation. Like peripheral
membrane proteins, they are not
embedded in the bilayer.
Membrane Protein Topology (I)
The term membrane protein topology refers
to the localization of protein sequences and
domains with respect to the plane of the
lipid bilayer. All copies of a given protein
adopt the same topology in the membrane,
and are oriented asymmetrically with
respect to the bilayer plane. The topology
of the erythrocyte plasma membrane
protein, glycophorin, is shown in Fig. 11-8.
The N-terminus of glycophorin resides
outside the membrane, while the C-terminus
is located in the cytoplasm. Glycophorin is
an integral membrane protein, and a 19residue  helix (amino acids 75-93)
consisting predominantly of nonpolar amino
acids anchors it in the membrane. (Note
that about 20 amino acids are needed to
span a 30Å wide membrane in the  helical
conformation). The attachment points for
N- and O-linked oligosaccharide chains are
located in the polar region located outside
the membrane. Glycosylated domains are
invariably found outside the membrane in
membrane glycoproteins.
Membrane Protein Topology (II)
Six topological categories of integral membrane proteins are now
known. Types I and II have a single transmembrane helix, where
the C-terminal end of the protein is located outside or inside,
respectively. Note that glycophorin is a Type II integral membrane
protein. Type III proteins have multiple transmembrane segments,
while Type IV proteins are assemblies of multiple different
polypeptide chains usually oriented to form a channel in the bilayer.
Type V membrane proteins are held to the bilayer primarily by
covalently linked lipids. Lastly, Type VI membrane proteins have
both transmembrane segments and lipid anchors. The structures of
very few integral membrane proteins have been solved by X-ray
crystallography. Usually structures are determined by treatment
with proteases or chemical reagents that are membrane impermeable
and modify amino acids on only one side of the bilayer.
V
IV
Structure of Bacteriorhodopsin
The integral membrane protein
called bacteriorhodopsin is one of
few membrane proteins whose
structure has been determined at
atomic detail by X-ray
crystallography. Bacteriorhodopsin
is a light-driven proton pump that
is located in the purple membranes
of the photosynthetic bacterium,
Halobacterium salinarum. The
protein contains a light-absorbing
pigment, retinal, which is attached
to one of the protein’s seven
transmembrane segments
and is buried deep in the interior of the membrane.
Conformational changes in retinal caused by light absorption drive
conformational changes in apobacteriorhodopsin which cause
protons to be pumped across the membrane. This generates a
proton gradient across the membrane which is exploited for
energy production.The seven transmembrane segments are tilted
slightly with respect to the plane of the membrane. Each segment
contains about 20 amino acids in  helical conformation. The seven
transmembrane segments are clustered together and are
surrounded by membrane lipids. Some membrane lipids actually
occupy spaces between the segments.
Lipid Annuli Surrounding Membrane Proteins
Included in the X-ray crystal structures of integral membrane
proteins are tightly bound membrane lipids surrounding the proteins
as a bilayer shell or annulus. It is presumed that these lipids also
bind tightly to the proteins in membranes. In Fig. 11-11a, the lipid
annulus associated with the transmembrane water channel, sheep
aquaporin, is shown. In the figure, aquaporin protein is colored dark
blue. Tightly bound lipids have their head groups colored in light
blue and their fatty acyl chains in yellow. The polar head groups of
the lipids contact polar amino acids in aquaporin, whereas the fatty
acyl chains contact hydrophobic residues. It is not uncommon for a
membrane protein to require the presence of certain lipids in the
annulus surrounding it.
Membrane Protein Topology Prediction (I)
The locations of transmembrane
segments in a membrane protein can be
predicted from computer-based analysis
of its amino acid sequence. In essence,
what is done is that an algorithm is
used to scan the amino acid sequence
looking for stretches of residues on the
order of 20 amino acids long that are
sufficiently nonpolar to be embedded in
a lipid bilayer. To achieve this end,
amino acids are assigned a hydropathy
index for transfer from water to a
hydrocarbon phase such as listed in
Table 3-1. Then the membrane protein
sequence is scanned and the average
hydropathy index is calculated for a
window of 7 to 20 residues, depending
on the program. The average
hydropathy is plotted on the y-axis
against the middle residue number in
the window. Plots like those shown in
Fig. 11-12 result. In this figure it can
be seen that the sequence of
glycophorin yields one predicted
transmembrane segment, whereas the
sequence of bacteriorhodopsin yields 6
or 7 segments.
Membrane Protein Topology Prediction (II)
Membrane protein topology can further be predicted in two other
ways. First, as the crystal structures of the five membrane
proteins in Fig. 11-13 reveals, Trp and Tyr residues commonly are
located at the interface between the membrane surface and
surrounding water. These residues serve as membrane interface
anchors because they are simultaneously able to interact with the
lipid and aqueous phases on either side of the bilayer surface.
Second the orientations of segments in the bilayer can generally be
predicted by the “positive-inside rule”. Experimental analysis has
revealed that Lys, Arg, and His residues are more likely to be
adjacent to the ends of  helical transmembrane segments emerging
into the cytoplasm of cells than outside the membrane.
ß-Barrel Membrane Proteins
The  helical conformation is an ideal secondary structure for
building transmembrane segments. All backbone hydrogen bonds
occur in the interior of the helix, and the R groups radiate out from
the helix axis; if composed of nonpolar residues, the helix is ideally
designed for interaction with membrane fatty acyl chains. On the
other hand individual ß strands are poorly structured to serve as
transmembrane segments. Nonetheless, there are many integral
membrane proteins that are rich in ß conformation. These proteins
are known as ß-barrel proteins and they make up a large fraction of
the outer membrane proteins of Gram-negative bacteria and
mitochondria (e.g., the porins, Fig. 11-14). To allow the ß
conformation to be located in the membrane interior, antiparallel ß
sheets form cylindrical (barrel-like) structures. Backbone hydrogen
bonding is satisfied through interactions within the cylindrical ß
sheets. And in most cases every other residue in the strands is a
nonpolar one whose R group points out toward membrane fatty acyl
chains. Because ß conformation is more extended than 
conformation, only 7-to-9 residues are needed for a ß strand to
span the bilayer.
Lipid-linked Membrane Proteins
Some membrane proteins contain
one or more covalently attached
lipids that anchor them to
membranes (Fig. 11-15). The
tethering to the membrane is
not that strong, so most lipidlinked proteins have more than
one attached lipid and/or also
bind to the head groups of
membrane lipids via ionic and
hydrogen bond interactions.
Lipid-linked membrane proteins
can be attached to the membrane via palmitoyl groups covalently
bound to an internal Cys or Ser residue, an N-myristoyl group on
an N-terminal Gly residue, a farnesyl or geranylgeranyl group on a
C-terminal Cys residue, or a GPI (glycosylated phosphatidylinositol)
linkage. The first three types of lipid-linked proteins reside in the
cytoplasm. GPI-linked membrane proteins project outside the cell.
N-myristoylated proteins commonly also contain a hydrophobic
transmembrane segment. Cysteine-palmitoyl-linked proteins are
weakly attached to the membrane and membrane binding often is
reversible. In intestinal epithelial cells, GPI-linked proteins are
exclusively targeted to the apical (lumenal) instead of basal
(bloodstream) membranes of the cells.
Phase Transitions in Membrane Lipids
Membrane lipids have characteristic melting
points that are determined largely by the
composition of their fatty acids. Saturated
fatty acids pack better next to one another
in bilayers than do unsaturated and
polyunsaturated fatty acids. Therefore,
bilayer lipids rich in saturated fatty acids are
less fluid than lipids rich in unsaturated fatty
acids at a given temperature. For any
membrane lipid composition, lowering the
temperature significantly will create what is
termed the liquid-ordered state, wherein
fatty acyl chains are extended and thermal
motions are greatly restricted (Fig. 11-16).
At significantly higher temperatures, carboncarbon single bonds in fatty acyl chains begin
to rotate, chains become less extended, and
thermal motions become considerable
(liquid-disordered state). Lipids become free to rotate and diffuse
laterally in the bilayer. All cells regulate the degree of fluidity of
their membrane lipids (see Fig. 11-17 below). Animals regulate
fluidity by adjusting the levels of unsaturated fatty acids and
cholesterol. Low concentrations of cholesterol in membranes increase
fluidity, whereas high concentrations decrease fluidity.
Affect of Temperature on Bacterial
Membrane Fatty Acid Composition
The fluidity of biological membranes must be maintained at a near
constant level for proper function of membrane-bound enzymes,
transporters, and receptors. Warm blooded animals typically need
not adjust membrane composition to regulate fluidity due to changes
in environmental temperature. However, bacteria, for example,
often experience wide shifts in temperature that would affect
membrane fluidity. Therefore, bacteria alter the compositions of
the fatty acids in their membrane lipids to compensate for
temperature fluctuations. Namely, at lower growth temperatures
bacteria incorporate a higher amount of unsaturated fatty acids
into membrane lipids, and vice versa (Table 11-2). This helps
maintain fluidity at the correct level regardless of temperature.
Diffusion and Translocation of Membrane
Lipids (I)
In membranes existing in the liquid-disordered state, lipid motion is
great. Uncatalyzed lateral diffusion occurs very rapidly allowing
lipids to move about in each leaflet of the bilayer (Fig. 11-17b).
Uncatalyzed transbilayer (“flip-flop”) diffusion (Fig. 11-17a) is also
possible, although it occurs very slowly owing to the thermodynamic
cost of moving the polar head group of a membrane lipid through
the nonpolar fatty acyl chain interior of the bilayer.
Diffusion and Translocation of Membrane
Lipids (II)
As discussed previously, different species of membrane
phospholipids maintain characteristic and asymmetric distributions
between the leaflets of cell membranes. Research in recent years
has revealed the existence of membrane proteins that catalyze
transbilayer translocation of phospholipids (Fig. 11-17c). Proteins
called flippases catalyze translocation of the amino-phospholipids,
phosphatidylethanolamine and phosphatidylserine, from the outer
to the inner leaflet of the cytoplasmic membrane. Keeping PS out
of the outer leaflet reduces the possibility of the cell undergoing
apoptosis. Proteins called floppases move phospholipids in the
opposite direction. Lastly, proteins called scramblases move lipids
in both directions down their
concentration gradients randomizing
the distribution across leaflets.
Scramblase activity is regulated, and
increases during apopotosis and other
physiological situations. Flippases and
floppases build up a gradient of
phospholipids across the leaflets of
the bilayer and acquire the energy
needed for transbilayer translocations
from the hydrolysis of ATP.
Measurement of Lateral Diffusion Rates
The rates of lateral diffusion for membrane
lipids (and proteins) have been measured using
techniques such as fluorescence recovery
after photobleaching (FRAP) (Fig. 11-18).
With FRAP, membrane components are labeled
with a membrane impermeant fluorescent
probe (red). The surface becomes uniformly
labeled with the probe. While viewed with a
fluorescence microscope, a small area of the
membrane surface is bleached (becomes nonfluorescent) by irradiation with an intense
laser beam. The time required for nonbleached lipids to diffuse back into the
bleached area by lateral diffusion is then
measured. From this time period it has been
calculated that some lipids diffuse laterally at
rates of 1 m/s. At this velocity, a lipid could
could circumnavigate an E. coli cell in about
one second.
Hop Diffusion of Membrane Lipids
Other methods have been used to track the movements of
individual fluorescently-labeled lipids in membranes. These
techniques show that lateral diffusion occurs rapidly within
small discrete regions of the membrane (Fig. 11-19). However,
movement from one region to another is limited, as if the lipids
are restricted by boundaries around them. Occasionally a lipid
moves from one region to another by what is called “hop
diffusion”. Immobilized proteins in the membrane may be
responsible for restricted movement of lipids.
Restriction of Membrane Protein Lateral
Diffusion
The lateral diffusion of many
membrane proteins is unrestricted.
However, for other membrane
proteins movement is limited, often
by their interactions with the
cytoskeleton located adjacent to
the membrane. For example, in
erythrocytes the lateral diffusion
of glycophorin and the chloridebicarbonate exchange protein is
limited due to the attachment of
their cytoplasmic domains to the
filamentous protein known as
spectrin (Fig. 11-20). Such
immobilized proteins are thought to
create the localized domains to
which lipids are confined. In other
cases, such as the acetylcholine
receptor of neurons, membrane
proteins cluster together in patches
where the interacting proteins
remain immobilized.
Membrane Microdomains (Rafts) (I)
In addition to the asymmetric distribution of lipids across a
bilayer, lipid distribution is not uniform even within a single leaflet.
Instead microdomains or “rafts” form wherein the lipid composition
differs from the surrounding sea of lipids. One common type of
raft is that formed in the outer leaflet of the cytoplasmic
membrane which is enriched in sphingolipids and cholesterol. Unlike
glycerophospholipids, which typically contain one saturated and one
unsaturated fatty acyl chain, sphingolipids often contain two longchain saturated fatty acids. This drives their self-association, and
cholesterol packs well within the sphingolipid chains. Its
concentration therefore is enriched in these rafts. Overall, the
raft is thicker and more ordered than neighboring microdomains
more abundant in glycerophospholipids.
Membrane Microdomains (Rafts) (II)
Sphingolipid/cholesterol rafts are enriched in two classes of
membrane proteins--GPI-anchored proteins and cysteine-palmitoyl
(myristoyl)-linked proteins. The fatty acyl groups tethering these
two types of proteins interact well with the acyl groups present in
the raft. In contrast, the prenyl groups of prenylated proteins,
such as the Ras GTPase involved in signaling by growth factors,
does not interact well with the acyl chains of the raft, and is
excluded from entry. Thus different classes of membrane proteins
become functionally segregated into different regions of a
membrane. Based on detergent extraction studies, in which
sphingolipid/cholesterol rafts are resistant to extraction, it is
estimated that about half of the cytoplasmic leaflet of the plasma
membrane of a eukaryotic cell is composed of rafts. Raft
diameters average about 50 nm, and rafts contain a few thousand
sphingolipid molecules.
Caveolin-induced Membrane Curvature
Caveolin is an integral membrane protein
with two globular domains connected by a
hairpin-shaped hydrophobic domain (Fig.
11-21 & 22). It further is anchored to the
membrane by three palmitoyl groups in the
C-terminal globular domain. Caveolin dimers
are attracted to cholesterol-rich regions of
the membrane (such as in sphingolipid/
cholesterol rafts). Due to the intrinsic
shape of the caveolin dimer, the associated
bilayer curves inward forming caveolae
(“little caves”) in the surface of the cell.
Caveolae are implicated in cellular functions
such as membrane trafficking and
transduction of external signals into cellular
responses.
Membrane Fusion
Changes in membrane curvature
are central to the ability of
membranes to undergo fusion. As
shown in Fig. 11-23, processes
that require membrane fusion
events are common within cells. In
membrane fusion, membrane
continuity is preserved as
illustrated below for the process
of neurotransmitter release at a
synapse.
Models for Protein-induced Membrane
Curvature
Bilayer curvature can be induced by the
binding of an intrinsically curved protein,
such as modeled in Fig. 11-24a & b.
Alternatively, scaffolding proteins that when
assembled form curved supramolecular
complexes can curve an attached membrane
(Fig. 11-24c). One well known family of
proteins that functions in this manner, BAR
domain proteins, forms crescent shaped
scaffolds that curve the attached bilayer.
BAR domains consist of coiled coils that
form long, thin curved dimers with a
positively charged concave surface that
interacts with negatively charged head
groups of membrane phospholipids.
Membrane Fusion: Neurotransmitter Release
The fusion of two membranes requires
that they first recognize one another,
their surfaces come into close contact with
the exclusion of water, the outer leaflets
of the two bilayers become disrupted and
fuse together (hemifusion), and finally
their bilayers fuse to form a single
continuous bilayer. In neurotrans-mitter
release (Fig. 11-25), fusion proteins called
v-SNAREs in secretory vesicles, and tSNAREs in the plasma membrane interact
with one another drawing the two
membranes together. The protein known as
SNAP25 also plays a role in this process.
Once in contact, these three proteins coil
around one another and exert lateral
tension on the bilayers that leads to
hemifusion. Complete fusion subsequently
occurs releasing the contents of the
secretory vesicle outside the cell.
Clostridium botulinum toxin is a protease
that cleaves SNAREs and SNAP25,
resulting in inhibition of neurotransmitter
release and death.