Download Chem*3560 Lecture 27: Membrane transport

Chem*3560
Lecture 27:
Membrane transport
Diffusion across permeable membranes
If solutions are separated by a permeable membrane, solutes will
cross the membrane by random diffusion. Molecules leave the side
with higher concentration at a faster rate, causing the concentration to
decrease on the source side and increase on the destination side, until
equilibrium is reached when both sides are at equal concentration.
If the solute is charged, a concentration difference across the
membrane may induce a voltage difference or membrane potential Vm (same as ∆ψ used in the
context of oxidative phosphorylation). If negative ions are in excess on the left, the membrane potential
will be negative on the left. The membrane potential also provides a driving force, because negative ions
will tend to move towards the positive side (Lehninger p.408).
Ionic substances move across a permeable membrane in response to combination of the membrane
potential and the concentration difference, which is referred to as the electrochemical gradient.
The net direction of ion movement will decrease both concentration difference and membrane potential,
until both are zero and net flux ceases.
Lipid membranes are impermeable to most polar molecules
Membranes surround cells to enclose their contents - leaving open the question of how necessary
nutrients enter cells or pass through internal membranes. The bilayer portion of the membrane is
impermeable to most polar molecules, which represent most of the intermediates of metabolism.
Membrane proteins play a direct role in allowing transport of substances across membranes.
In addition to simple transport, which is governed by the relative concentrations on each side of the
membrane, some membrane proteins are also involved in active transport, in which a particular solute
can be driven across a membrane from a region of low concentration to a region of higher
concentration. This requires an external energy source such as ATP hydrolysis.
Facilitated diffusion
Many transport proteins simply provide a passage through the membrane for specific solutes, governed
solely by the electrochemical gradient, without any external energy being applied. This is facilitated
diffusion. The transport protein permits passage of solute, but does not drive solute against an
electrochemical gradient.
The problem for polar solutes and lipid bilayers is that solutes exist in aqueous solution in a hydrated
state. In order to enter the hydrocarbon core of the bilayer, they must lose their hydration layer, and
the energy required to dehydrate a solute may be hundreds of kJ/mol. Desolvation contributes to the
activation free energy ∆GI for the transport process, hence transport of polar solutes across a lipid
bilayer is imperceptibly slow (Lehninger p.409).
A transport protein provides a channel through the membrane , and provides binding sites for selected
solutes such that the binding energy of solute in the channel is roughly equal to the solvation
energy in the surroundings. The result is that solute can leave the aqueous medium, and become
dehydrated, but immediately passes to a binding site of equivalent energy. The energy change is thus
minimal so that activation energy is low. When the solute reaches the other side, it must be
released from the binding site, but immediately becomes hydrated again.
If the channel binds a solute too tightly, it will enter the channel easily, but now there's an activation
energy barrier for the solute to reenter the aqueous solution on the other side.
Many passive substrate transporters share a 12-helix structure
These proteins belong to the Major Facilitator Superfamily. A superfamily is a grouping of proteins
from many different organisms that share a common structural arrangement. The Facilitator superfamily
perform facilitated diffusion of substrates across membranes. There were several structures published
during 2002-2003 illustrating how these proteins may function. (The current edition of Lehninger was
written 1998-1999 so the description on pp. 411-413 is speculative but not far off the mark).
These proteins are arranged in two subgroups of six helices each that surround the central substrate
binding site like a pair of cupped hands. The outward faces of the helices are lined with nonpolar amino
acids where they contact the hydrocarbon core of the bilayer, but have inward facing polar amino acids
facing the substrate binding site.
The schematic layout of the twelve
helices I-XII is shown above viewed
looking down with the bilayer in the
plane of the paper. This produces a
favourable binding site for a polar
substrate in the middle of the protein (S in the schematic).
On the right above is a cross section through the actual protein structure of LacY, the lactose sugar
transporter in the E. coli cell membrane. Yellow amino acids are nonpolar, and face the hydrocarbon
core of the bilayer. Green amino acids are polar, and face the substrate (grey C and red O atoms).
The section has been cut in a plane at mid bilayer level. Atoms
are shown in their true volume, so there are few gaps.
The helical wheel
The asymmetric spatial arrangement of polar and non-polar
amino acids in the α-helices that make up this structure can be
detected in the amino acid sequence by a technique called
helical wheel analysis.
The method is based on the geometry of the α-helix, which has
3.6 amino acids per turn of helix, or in other words the
orientation of each amino acid is 100o away from the
previous.
Amino acids in a sequence are written out around the
perimeter of a circle, one amino acid every 100o . If the amino
acids have the correct distribution in the sequence, one side of
the helix will be predominantly polar and the side that contacts
bilayer will be non polar.
The two halves of LacY alternate between two conformations
IColour coding in the figure runs from blue at N-terminus to red at the C-terminus. If colours are traced
it can be seen that the six helices making the N-terminal half are all on the left of the substrate, and the
six making the C-terminal half are all on the right.
In the left hand structure, the substrate side is exposed to the exterior ///s \\\ and can bind substrate.
The two halves rock about a pivot point roughly at the midline, so that the channel closes on the exterior
half and opens on the cytoplasmic side, \\\s ///, and this allows the substrate to be released into the cell.
The channel is therefore always closed on one side or the other, so non-substrates can't leak through
(see Lehninger p. 412, Fig. 12-26 for a good guess at how the erythrocyte glucose transporter might
work).
Substrate transport follows
Michaelis-Menten kinetics
Transport depends on 1) a substrate
occupying a binding site, and
2) the conformational changes which
accesses the opposite side. This
pattern is little different from the
standard Michaelis-Menten behaviour
of an enzyme.
The transport of a single substrate is called uniport
Uniport: a single substrate is transported in the direction
governed by the electrochemical gradient. After passage of
substrate, the transporter randomly flips between conformations. If
there is more substrate on the left, the site is more likely to become
occupied when open to the left, so net transport is towards the right
where the concentration is lower.
Many transporters operate by a cotransport
mechanism
Symport: two substrates are
co-transported in the same
direction. This could be due to
the requirements of the substrate
site, so the conformational change
only occurs when both
components are present.
Alternatively, there could be two
channels and passage of the
second substrate is necessary for the transporter to return to its initial state. In many cases, the second
substrate is H+, and the transporter takes advantage of the H+ gradient of oxidative phosphorylation.
Net direction of transport is governed by whichever of the two substrates has the steeper
electrochemical gradient.
Antiport: substrates are exchanged; for each molecule A that
enters, a molecule of B must leave. Substrate A flips the
conformation in one direction, substrate B brings it back to the
original state. Net direction of transport is governed by whichever of
the two substrates has the steeper gradient.
An example of antiport is the ATP:ADP exchange translocator of
the mitochondrial membrane, which exports ATP while bringing in
ADP:
ATPin + ADPout ‡ ATPout + ADPin
The advantage of antiport is that it maintains balance of
substrates between compartments. The mitochondrion should
never accumulate too much or run out of adenosine compounds.