Download 5. Membrane Transporters

Molecules to Medicine- Cell Physiology
5. Membrane Transporters 20 Nov 9AM
Bill Betz
http://www.cuphys.net/cellphys
[email protected]
Reading: Alberts et al, 4th ed: pp 615-631.
Lehninger, 4th ed: pp 389-411.
Key words: primary active transport, secondary active transport, cotransporter, exchanger,
electrogenic
5. Membrane Transporters
Earlier we talked about the famous sodium-potassium pump, the ubiquitous transporter
that renders cells ‘functionally impermeable’ to sodium ions, and keeps the cells from swelling
uncontrollably. Today we will consider other types of carrier mechanisms.
The first thing to note is that none of them, including the Na/K pump, actually ‘carries’
solute cargo across the membrane. There is an antibiotic (valinomycin) that actually diffuses back
and forth across the membrane (shuttling potassium ions), but no biological molecule is known to
act this way, so transporter is a better word than carrier. How do protein transporters work? For
most, we don’t really know, but the general model of the Na/K pump (a tube with a gate on each
end and dynamic (changeable) binding affinity) is probably a good model for the others as well
(see Handout #4, page 5).
Conceptually, matters are pretty simple. Some transporters act like ion channels, shuttling a
single solute species in either direction. This is called facilitated diffusion (a historic term applied
to molecules that shouldn’t be able to diffuse across lipid membranes
because of their large size or charge, but do get across). The best known
example of this is the glucose transporter (cartoon at right). The glucose
transporter will transport glucose in either direction, and burns no energy
in the process. Thus, it is not a pump. You might then wonder how cells
accumulate glucose. The answer is that as soon as a glucose molecule gets
into the cell, it is phosphorylated to Glucose-6-Phosphate, which doesn't
fit on the transporter, and so is "trapped" inside.
Glucose uptake by cells is regulated by insulin, a hormone secreted by specialized cells in
the pancreas when plasma glucose levels rise. How does insulin turn on the transporter? It turns out
that in the absence of glucose, the transporter is not even present in the plasma membrane; it is
sequestered inside the cell, in the membrane of intracellular vesicles. Insulin triggers a biochemical
cascade that causes the vesicle membranes to fuse with the surface membrane (exocytosis),
exposing the glucose transporter to the ECF. The transporter then gets busy and ‘carries’ glucose
inside. When insulin subsides, the transporter molecules are reinternalized (endocytosis).
Pumps. Now we turn to transport proteins that can cause a substance to be accumulated or
expelled against its electrochemical gradient. Dozens have been identified, pumping nearly every
substance of importance in the body in one cell or another. The only evident exceptions are water
and urea (the end product of nitrogen metabolism in mammals). Isn’t it odd that no water pumps
exist anywhere in nature, given its importance? Water always moves down its concentration
(osmotic) gradient. By definition, pumps move solute against an energy gradient, and so require an
energy source. We subdivide pumps into two categories, depending on the source of that energy.
Primary active transporters, like the Na/K pump, derive their energy directly from the
splitting of ATP. There are no other ubiquitous primary active transporters in the plasma
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membrane of cells. Some specialized cells have them, and you will study them later. For example,
the cells in the stomach that secrete acid, and certain cells in the kidney geared for excreting
protons from the body possess proton pumps in their plasma membranes that rely directly on ATP
for their energy source.
Inside cells (as opposed to the surface membrane) there are other primary active
transporters. One pumps protons into intracellular membrane-bound organelles (endosomes,
vesicles, lysosomes). Another pumps calcium ions into membrane-bound compartments. All
proton and calcium pumps are oriented to pump these ions out of the cytoplasm.
Inside mitochondria is a very special proton pump (the F1-ATPase, or ATP synthase) that,
when running backwards, lets protons leak across a membrane and synthesizes, rather than
hydrolyzes ATP.
Secondary active transport is the mechanism by which most substances are pumped. In this
case, the energy to do the direct work of pumping comes not from metabolism (ATP), but from a
secondary source. Usually this energy source is the 'downhill leak' of Na+ into the cell. For
example, cells can accumulate amino acids against their energy gradients. This active uptake is
dependent on external Na+; if external Na+ is removed, amino acid uptake is abolished.
Conversely, removing the amino acid reduces the entry of Na+. The carrier ingeniously captures
the energy released by the inward leak of Na+ and instead of letting it escape as heat, uses it to
pump the amino acid into the cell.
There are two basic types of secondary active transporters, those that move different solute
species in the same direction (cotransport), and those that move solute in opposite directions
(antiport, or exchange). The cartoons below illustrate these two types (arrows pointing up mean the
substance is pumped; down means leak).
Secondary active transporters do not necessarily always run in the same direction. They
will always tap the bigger leak to drive the smaller pump. Consequently, they can reverse direction
sometimes. One of the most important examples of this is the sodium-calcium exchanger, which
reverses direction in heart muscle cells every time the heart beats (more on this later).
All secondary transport mechanisms depend ultimately on the Na+/K+ pump (and therefore
on ATP). For example, if the Na+/K+ pump is blocked, cells fill up with Na+, and thus the Na+
electrochemical gradient is reduced. Because this is the energy source for secondary active
transport, all of these transport mechanisms suffer.
Some secondary active transporters are electrogenic, in that one cycle produces a net
charge transfer across the membrane. For example, Na/amino acid transporters are electrogenic,
because one cycle transfers a net positive charge (Na+) into the cell. Other secondary active
transporters are not electrogenic; an example is the Na/K/2Cl cotransporter, which each cycle
moves one sodium ion, one potassium ion, and two chloride ions into the cell. The main feature of
electrogenic secondary active transporters is that their activity is governed by the membrane
potential (as described in the Appendix). Electrically silent transporters could not care less about
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membrane potential.
Here are some important examples of secondary active transporters driven by the inward
sodium leak:
Calcium transport. There is a huge electrochemical gradient for calcium ions across cell
membranes. In fact, no other ion is further from equilibrium than calcium. The extracellular
(ionized) calcium concentration (about 1 mM) is nearly 10,000 times greater than the intracellular
concentration (about 0.0002 mM, or 200 nM); thus its concentration gradient is inward. And the
electrical gradient is also inward, of course, because the ICF is electrically negative and calcium is
positively charged. From the Nernst equation, ECa is calculated to be about +111 mV (ECa =
(60/2)*log(1/0.0002) = +111 mV). Thus, given the opportunity (i.e., an open calcium channel),
Ca++ ions will always leak into cells, so there must be a pump to extrude them. The Na/Ca
exchanger’s main job is to pump calcium ions out of the cell. The inward leak of sodium ions
provides the energy source.
The Na/Ca exchange pump takes on a special significance in the heart, where it actually
switches direction during each heartbeat. (You will study muscle contraction shortly, and will learn
that calcium ions are necessary for contraction.) At rest (while the ventricles are refilling with
blood during diastole), the exchanger runs forward, pumping Ca++ out (keeping the muscle
relaxed) as Na+ leaks in. When the ventricles contract (systole), the exchanger switches direction,
letting Ca++ leak into the cell, where it strengthens the force of contraction. The direction of Ca++
movement is controlled by the value of the membrane potential; when Vm is more negative than
about -60 mV, the sodium leak rules, and drives the outward pumping of calcium; when Vm is
more positive than -60 (during the action potential), the pump reverses direction, and calcium leaks
in (pumping sodium out).
Digitalis. For centuries it has been known that an extract of the beautiful purple foxglove
can help a weak heart beat stronger. Digitalis (and related drugs) exert their action by acting
directly on, not the Na/Ca exchanger, but the Na/K pump! In fact, digitalis blocks the Na/K pump.
How does this lead to an increase in the strength of contraction of heart muscle? Blocking the
Na/K pump of course allows intracellular sodium ion concentration to increase. In turn, this
reduces the energy available to all sodium-driven secondary active transporters, including the
Na/Ca exchanger. Thus, digitalis indirectly inhibits the Na/Ca exchanger, allowing intracellular
calcium ion concentration to rise, which increases cardiac contractility.
Hydrogen ions (protons) are also pumped out of most cells by a Na+/H+ exchange carrier,
which operates under the same principles as the Na/Ca exchanger. Protons are harder to study than
UFO's, because they capriciously vanish and reappear as they bind to and unbind from various
buffers. Typically, only about 1 in a million protons is free, as H+; the rest are hiding, bound to
buffers. The free concentration of protons in the ICF is about 100 nM (pH=7.0); in the ECF it’s
even lower, about 40 nM (pH=7.4). From the Nernst equation, EH = -24 mV. That means in cells
with membrane potentials more negative than -24 mV (most cells), H+ must be pumped out of the
cell. The mechanism is a secondary active transport system, in which the inward leak of Na+ drives
the outward pumping of H+.
Chloride ions are pumped into some cells by a secondary active transport process
(Na/K/2Cl cotransporter). As a result, ECl moves in a positive direction, away from the resting
membrane potential.
The (non-existent) ‘H+/K+ exchanger’. There are several clinical situations that suggest the
presence of a system that will exchange K+ for H+, and vice versa. For example, infusing K+ causes
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acidemia (the K+ is taken up by cells ‘in exchange’ for H+), and infusing acid causes hyperkalemia
(elevated (hyper-) potassium (-kal- for Latin kalium) in the blood (-emia)). While it is conceptually
simple (and useful) to think in terms of an H/K exchanger, the reality is that such a transporter
probably does not exist. Rather, the process evidently involves different transporters, perhaps
working in pairs in parallel, the upshot being hydrogen/potassium exchange. For example,
hyperkalemia will cause extra K+ uptake via the Na/K pump. Hyperkalemia also will depolarize
cells (by shifting EK in a positive direction), and the change in membrane potential can affect the
rate of activity of electrogenic transporters. One such transporter, which transports 3 bicarbonate
ions and one sodium ion from the ICF to the ECF, is inhibited by depolarization. Thus, the
reduction in activity will reduce bicarbonate extrusion. Because bicarbonate is a base, its slower
extrusion will cause acidemia. So, which is easier to remember, the idea of a (non-existent) H/K
exchanger, or trying to keep track of several different cotransporters and exchangers that might be
playing a role?
Below are 10 cartoons of membrane transporters. One of them has been mislabeled. Which one?
answer: the last one (it is a sodium-bicarbonate
cotransporter, not an exchanger)
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