Download The Electron Transport System of Mitochondria

Overview of Mitochondria Structure and
Function
The cytoplasm of nearly all eukaryotic cells contain mitochondria, although there is at
least one exception, the protist Chaos (Pelomyxa) carolinensis. They are especially
abundant in cells and parts of cells that are associated with active processes. For example,
in flagellated protozoa or in mammalian sperm, mitochondria are concentrated around the
base of the flagellum or flagella. In cardiac muscle, mitochondria surround the contractile
elements. Hummingbird flight muscle is one of the richest sources of mitochondria
known. Thus, from their distribution alone one would suspect that they are involved in
energy production. Multicellular organisms probably could not exist without
mitochondria. The inability to remove electrons from the system and the buildup of
metabolic end products restrict the utility of anaerobic metabolism. Through oxidative
phosphoryation mitochondria make efficient use of nutrient molecules. They are the
reason that we need oxygen at all.
Mitochondria vary considerably in shape and size, but all have the same basic
architecture. There is a smooth outer membrane, surrounding a very convoluted inner
membrane. The convolutions form recognizable structures called cristae. The two
membranes have very different properties. Together they create two compartments,
namely the intermembrane space (the comparment between the membranes), and the
matrix (the very interior of the mitochondria).
The double-membraned mitochondrion can be loosely described as a large wrinkled bag
packed inside of a smaller, unwrinkled bag. The two membranes create distinct
compartments within the organelle, and are themselves very different in structure and in
function.
The outer membrane is a relatively simple phospholipid bilayer, containing protein
structures called porins which render it permeable to molecules of about 10 kilodaltons or
less (the size of the smallest proteins). Ions, nutrient molecules, ATP, ADP, etc. can pass
through the outer membrane with ease.
The inner membrane is freely permeable only to oxygen, carbon dioxide, and water. Its
structure is highly complex, including all of the complexes of the electron transport
system, the ATP synthetase complex, and transport proteins. The wrinkles, or folds, are
organized into lamillae (layers), called the cristae (singlular: crista). The cristae greatly
increase the total surface area of the inner membrane. The larger surface area makes
room for many more of the above-named structures than if the inner membrane were
shaped like the outer membrane.
The membranes create two compartments. The intermembrane space, as implied, is the
region between the inner and outer membranes. It has an important role in the primary
function of mitochondria, which is oxidative phosphorylation.
The matrix contains the enzymes that are responsible for the citric acid cycle reactions.
The matrix also contains dissolved oxygen, water, carbon dioxide, the recyclable
intermediates that serve as energy shuttles, and much more. Diffusion is a very slow
process. Because of the folds of the cristae, no part of the matrix is far from the inner
membrane. Therefore matrix components can diffuse to inner membrane complexes and
transport proteins within a relatively short time.
Electron micrographs have revealed the three dimensional structure of mitochondria.
However, since micrographs are themselves two dimensional, their interpretation can be
misleading. Texts frequently show a picture of a 'typical' mitochondrion as a bacteriasized ellipsoid (perhaps 0.5 by 1 micrometer). However, they vary widely in shape and
size. Electron micrographs seldom show such variation, because they are twodimensional images.
Isolated mitochondria, such as from homogenized muscle tissue, show a rounded
appearance in electron micrographs, implying that mitochondria are spherical organelles.
Mitochondria in situ can be free in the cytoplasm or packed in among more rigid
structures, such as among the myofibrils of cardiac muscle tissue. In cells such as muscle,
it is clear that mitochondria are not spherical, and often are not even ellipsoid. In some
tissues, the mitochondria are almost filamentous, a characteristic that two dimensional
micrographs may fail to reveal.
Their most immediate function is to produce adenosine triphosphate (ATP) by
systematically extracting energy from nutrient molecules (substrates). ATP is the
universal energy-yielding commodity in cells, used by enzymes to perform a wide range
of cellular functions. We refer to the process as aerobic metabolism because it requires
constant removal of excess electrons through the reduction of oxygen. The need for
oxygen as an electron acceptor is the sole reason that we breathe air. In fact, the most
immediate purpose of our respiratory and circulatory systems is to deliver oxygen to the
tissues for use by mitochondri (and to eliminate carbon dioxide). By the way, the term
respiration is synonymous with the phrase "cellular respiration." It refers to the
reduction of oxygen to water. Simply inhaling and exhaling is simply breathing, not
respiration.
Enzymes and energetics
In order to understand the role of mitochondria in cells, you will need some very basic
concepts concerning how cells store and transfer energy. All molecules contain energy,
stored in the molecular structure itself. A portion of that energy, called free energy, can
be used to do work. A chemical reaction that adds free energy to a molecule is said to
reduce the molecule. Removing free energy from a molecule is called oxidation. When a
reaction results in transfer of free energy from one molecule to another we call it an
oxidation/reduction, or redox reaction. In a redox reaction one or more molecules is
reduced (gains energy) while one or more molecules is oxidized (loses energy).
In cells, enzymes are responsible for reactions that transfer free energy from one
molecule to another. A typical enzyme is extremely selective for its substrates (the
reactants). Enzymes bind substrates in such a way that they are brought in to positions
favoring a particular reaction, greatly lowering the activation energy for the reaction. By
controlling enzyme activity a cell can control what reactions take place, because the types
of reactions catalyzed by enzymes are extremely unlikely to occur spontaneously.
The second law of thermodynamics states that spontaneous reactions occur in directions
that increase the overall disorder of the universe. A consequence is that with each energy
transfer some energy is lost to the chaotic motion of molecules that we measure as
temperature. While enzymes are designed to conserve free energy, some energy is always
'wasted' with each process (although endotherms use the 'wasted' energy to maintain body
temperature). The stepwise oxidation of substrates by enzymes can be thought of as a
'controlled burn,' in which much of the available free energy is retained in a useful form.
Sources of nutrients
Nutrients are organic molecules that are ultimately derived from food sources (or, in
green organisms, from photosynthesis). They start off as fats, carbohydrates, and
proteins. Enzymes involved in intermediate metabolism oxidize nutrient molecules to a
form that can be converted to energy by mitochondria. Thus fats, carbohydrates, and
proteins are broken down to individual fatty acids, simple sugars, and amino acids.
Various metabolic pathways 'channel' the molecules into a limited number of forms,
which we call Krebs substrates (after Hans Krebs, who is credited with the discovery of
the basic metabolic cycle within mitochondria). Conversion to Krebs substrates is
accomplished in the cytoplasm, in the mitochondria matrix, or in both places, depending
on the reaction. A limited number of enzymes need be maintained in the mitochondria,
and energy from a variety of sources is funneled into what ultimately becomes a single
pathway.
Krebs' cycle
Enzymes within the mitochondria matrix are designed to oxidize the substrates within, in
a cyclic manner. That is, every product of a reaction is a potential substrate for another
reaction. The most reduced of the substrates (greatest available free energy) is citric acid
(citrate). The most oxidized of the substrates (least available free energy) is oxaloacetic
acid (oxaloacetate). Citric acid can be regenerated by coupling oxaloacetate to a two
carbon unit, acetyl-coenzyme A. Acetyl-coenzyme A, generated from metabolism of fats
and sugars, brings free energy back into the system. Energy also enters Krebs cycle as
amino acids are converted to various Krebs intermediates.
At certain points in the cycle, carbon dioxide is lopped off, so the number of carbon
atoms from citrate to oxaloacetate is reduced from 6 to 4. Hydrogens are removed when
water molecules are taken off of the substrates. So the system is in balance. But what
happens to the free energy that is being released by progressive oxidation of substrates?
Substrate Oxidation: Krebs Reactions
The citric acid cycle is also called the Krebs cycle, after Hans Krebs, who first proposed
its cyclic nature. The Krebs' cycle reactions take place in the matrix of the mitochondria.
Some of the final steps of intermediate metabolism take place there as well. For example,
in the matrix as well as the cytoplasm, glutamate (the amino acid glutamic acid) loses its
amino group and is oxidized to alpha-ketoglutarate.
Most texts and lecture courses start with glycolysis, then proceed to describe Krebs' cycle
as the "next step" in metabolism of sugars. Unfortunately, such a presentation may leave
students with the impression that metabolites follow a linear progression through
glycolysis to acetyl-coenzyme A to citric acid, through the Krebs intermediates to
oxaloacetate, which is then coupled to the two carbons of another acteyl-coenzyme A to
regenerate citric acid. Even in biochemistry texts, chapters on Krebs' cycle may start with
a picture showing only pyruvate as the starting point.
This figure is what sticks in the minds of most introductory level students, and it isn't
complete at all. First, it isn't just glycolysis that leads to the generation of acetyl
coenzyme A. Carbohydrates, fats, and proteins are all nutrients, and in fact fatty
acid metabolism results in the generation of acetyl coenzyme A as well.
Second, the 'starting point' for Krebs' cycle need not be acetyl-coenzyme A at all. In fact,
it really isn't appropriate to refer to a cycle as having a starting point (e.g., where does a
circle start?). Amino acids and odd-chain fatty acids can be metabolized into Krebs
intermediates, and enter at several points.
Finally, intermediates can be 'siphoned off' for use in biosynthetic pathways. They must
be replaced in order to maintain energy balance, however the fate of a Krebs intermediate
is not necessarily to cycle through the enzymes until it is completely oxidized to carbon
dioxide and water.
Since cycle intermediates can be incorporated into both anabolic and catabolic pathways,
the cycle is really amphibolic, not just catabolic.
Glutamate to alpha-ketoglutarate
The conservation of energy by Krebs reactions can be illustrated by looking at the fate of
glutamic acid, or glutamate. There are, in fact, several Krebs reactions that conserve the
energy of oxidation of substrates. Glutamate enters the intermembrane space through the
porins. A transport mechanism in the inner membrane called the glutamate-aspartate
exchange carrier takes the glutamate molecule into the matrix.
The enzyme complex known as glutamate dehydrogenase binds the glutamate molecule,
a molecule of oxidized nicotine adenine dinucleotide (NAD), and a water molecule. Off
comes the amino group, and glutamate is partially oxidized to alpha-ketoglutarate, which
you should recognize as a Krebs intermediate. In vivo, the alpha-ketoglutarate would be
further oxidized to succinyl-coenzyme A by alpha-ketoglutarate dehydrogenase, then the
succinyl-coenzyme A would be oxidized to succinate, etc. However, remember the
amphibolic nature of Krebs cycle - substrates may be utilized for biosynthesis rather than
undergoing further oxidations. In addition, limitations on Krebs cycle in isolated
mitochondria must be considered.
The oxidation of organic compounds releases free energy. This energy would be wasted
as heat, except the enzyme also catalyzes the reduction of the NAD. The enzyme is
designed so that the reaction cannot take place unless all of the reactants are present. Free
energy from the partial oxidation of glutamate is used to reduce the NAD. The result is a
molecule of NADH, which retains in its structure some of the free energy lost by the
glutamate.
Glutamate dehydrogenase catalyzes a reaction in which the energy from an exothermic
reaction is used to power an endothermic reaction. Therefore it performs what we call a
coupled reaction. The second law of thermodynamics, paraphrased, states that some of
the useful energy in any system is converted to useless energy during any process. That
is, any spontaneous process causes the entropy of the universe to increase. In coupled
reactions, only some of the free energy is conserved in the form of a reduced product.
The remainder is released as heat.
NADH is an energy carrier. After release from glutamate dehydrogenase it can be used to
power other reactions. It can also bind a specific enzyme on the mitochondria inner
membrane, transferring some of its free energy to the electron transport system. In
mitochondria, this is the principle role of NADH.
Succinate to fumarate
An important substrate in our studies of mitochondria function is succinic acid
(succinate). When succinate is brought into or generated in the mitochondria matrix in
sufficient quantity, succinate molecules bind the enzyme complex called succinate
dehydrogenase. The succinate dehydrogenase complex is also known as complex II of the
electron transport system, thus the oxidation of succinate to fumarate is the only Krebs
reaction that takes place on the inner membrane itself, as opposed to the other reactions
that are catalyzed by soluble enzymes. The energy carrier flavin adenine dinucleotide
(FAD) is also a part of the succinate dehydrogenase complex.
Because the enzyme and FAD are both part of the same complex, the only step needed to
initiate succinate oxidation is the binding of succinate to the enzyme. Even in severely
compomised mitochondria succinate supported respiration can usually be accomplished,
as long as fragments of the inner membrane remain.
Energy carriers
At specific points in Krebs' cycle, the enzyme responsible for the reaction conserves
some of the released free energy by reducing a molecule of either NAD+ or FAD. Thus
an oxidation is coupled to a reduction. Some of the free energy released by the oxidation
is conserved in the structure of the reduced molecule, either NADH or FADH2. With
coupled reactions, the enzymes are designed so that they must carry out both reactions, or
not at all.
Energy carriers in turn lose energy to molecules that are embedded in the inner
membrane. They are then reoxidized and ready to be used again (note that FAD is part of
complex II in the inner membrane). The purpose of the carriers is to shuttle energy from
the Krebs reactions to specific structures in the inner membrane. The molecules
embedded in the inner membrane are energy carriers of a different type. They transfer
energy by passing along a pair of electrons. Taken together, the electron-transporting
molecules in the inner membrane are called the electron transport system.
Electron transport
Components of the electron transport system include complexes I, II, III, and IV, plus
two individual molecules, coenzyme Q and cytochrome c.
NADH is reoxidized at complex I. A pair of electrons from NADH is then passed through
a series of electron transport carriers within complex I to coenzyme Q (NOT to complex
II). The electrons flow just like water rolling downhill, with energy released upon each
electron exchange. Within complex I, as each electron pair passes through a pair of
hydrogen ions (protons) is forced from the matrix to the intermembrane space.
Electrons cannot be passed through unless the protons are translocated at the same time.
The energy used to force the protons across the inner membrane is released by the
passage of the electron pair from NADH to coenzyme Q. The electrons are then passed
through complex III, where more protons are forced across, and complex IV, where still
more protons are forced across. Complex IV, called cytochrome oxidase, uses the energy
it receives along with an electron pair to reduce oxygen to water. Note that since
molecular oxygen is diatomic it takes two cytochrome oxidase complexes, two electron
pairs, and four hydrogen ions to complete the reaction.
Oxygen must be replenished, otherwise electron transport cannot proceed. Each carrier,
once reduced, would have to stay that way because there would be no place for the
electrons to go. The need to deliver oxygen to the electron transport system is why we
have respiratory and circulatory systems. Oxygen is necessary to "drain" electrons from
the system, otherwise all of the carriers would remain reduced and electron transport
would have to stop.
As electrons are tranferred from carrier to carrier, the total amount of free energy drops.
Each carrier is more electronegative than the carrier "upstream." Much of the energy is
lost as heat, but a significant amount is stored in the form of a proton gradient.
Chemiosmotic gradient and respiratory control
The electron transport system cannot force an infinite number of protons into the
intermembrane space. The inner membrane is impermeable to protons, and they
accumulate in the intermembrane space, creating what is called a chemiosmotic gradient.
A mitochondrion in vivo maintains its energy gradient at a constant level. That situation
does not change because of a mechanism that we call respiratory control.
Electron transport cannot proceed if protons cannot be pumped across the inner
membrane. Protons cannot be pumped unless the available energy to move them out of
the matrix exceeds the required amount plus what energy is lost to heat. If the inner and
outer membranes were sealed and there was no way of altering the gradient, electron
transport would have to stop. In reality, a number of processes act constantly to dissipate
the gradient, so electron transport never completely stops. Instead, the rate of electron
transport is regulated by the rate at which the energy is removed from the gradient. The
ability of the chemiosmotic gradient to limit electron transport is called respiratory
control.
Many students have trouble understanding the cause and effect relationship between
chemiosmotic gradient and electron transport. Nevertheless it is the most critical concept
toward a complete understanding of how mitochondria function.
Oxidative phosphorylation
Of course, the primary purpose of mitochondria is to phosphorylate ADP. The energy
needed to do that is stored as a gradient of protons. Presumably, if a proton were allowed
to come back into the matrix there would be a release of energy. How can that energy be
captured and exploited?
Embedded in the inner membrane among the structures of the electron transport system
are structures called the ATP synthase complex. The complex consists of a proton
channel and catalytic sites for the synthesis of ATP from ADP and phosphate. When
ADP and phosphate are available, they bind the catalytic sites on the ATP synthase.
When this happens, the channel opens, and protons can come whooshing back in. The
energy released is used to couple the phosphate to ADP, to make ATP.
The mechanism can be likened to a water wheel, where the flow of protons resembles a
flow of water downhill, and the turning of the wheel is the turning of ADP toward
phosphate to cause the bond to form.
Stimulation of electron transport by ADP
When ADP binds to ATP synthase, protons are driven into the matrix and as they do so
energy is drained from the gradient. As fast as energy is removed, the electron transport
system can force protons into the intermembrane space. The faster or slower energy is
removed from the gradient, the faster or slower electron transport can proceed. Again, the
rate of electron transport is subject to respiratory control.
Summary of key concepts
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Reactions that remove free energy are oxidation reactions; reactions that add free
energy are reduction reactions
Enzymes facilitate oxidation and reduction reactions at the same time, by
transferring free energy from one molecule to another; some energy is always lost
in such transfers
Locations: Krebs reactions take place in the matrix; electron transport takes place
on the inner membrane; a gradient is maintained across the inner membrane; ATP
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synthesis takes place in the matrix as protons cross the inner membrane from the
intermembrane space
Nutrients from a variety of sources are modified so that mitochondrial metabolism
involves a limited number of Krebs substrates
Metabolic products can enter Krebs cycle in a number of places
Krebs reactions transfer energy from the progressive oxidation of substrates to
two types of energy carriers, NAD and FAD
Energy carriers donate electrons and free energy to the electron transport system
(ETS)
Each successive carrier in the ETS, when reduced, carries less available free
energy than the one upstream
Electron transport cannot take place without pumping protons out of the matrix
Proton pumping produces a chemiosmotic gradient that in turn restricts the rate of
electron transport
Various processes drain energy from the gradient, allowing electron transport to
proceed
The most important process that exploits the gradient is ATP synthesis
Mitochondria in vivo constantly exercise respiratory control
Electron transport always proceeds as fast as energy is removed from the gradient
The Electron Transport System of
Mitochondria
Embedded in the inner membrane are proteins and complexes of molecules that are
involved in the process called electron transport. The electron transport system (ETS), as
it is called, accepts energy from carriers in the matrix and stores it to a form that can be
used to phosphorylate ADP. Two energy carriers are known to donate energy to the ETS,
namely nicotine adenine dinucleotide (NAD) and flavin adenine dinucleotide (FAD).
Reduced NAD carries energy to complex I (NADH-Coenzyme Q Reductase) of the
electron transport chain. FAD is a bound part of the succinate dehydrogenase complex
(complex II).
It is reduced when the substrate succinate binds the complex.
What happens when NADH binds to complex I? It binds to a prosthetic group called
flavin mononucleotide (FMN), and is immediately re-oxidized to NAD. NAD
is"recycled," acting as an energy shuttle. What happens to the hydrogen atom that comes
off the NADH? FMN receives the hydrogen from the NADH and two electrons. It also
picks up a proton from the matrix. In this reduced form, it passes the electrons to ironsulfur clusters that are part of the complex, and forces two protons into the
intermembrane space.
The obligatory forcing of protons into the intermembrane space is a key concept.
Electrons cannot pass through complex I without accomplishing proton translocation. If
you prevent the proton translocation, you prevent electron transport. If you prevent
electron transport, you prevent proton translocation. The events must happen together or
not at all.
Electron transport carriers are specific, in that each carrier accepts electrons (and
associated free energy) from a specific type of preceeding carrier. Electrons pass from
complex I to a carrier (Coenzyme Q) embedded by itself in the membrane. From
Coenzyme Q electrons are passed to a complex III which is associated with another
proton translocation event. Note that the path of electrons is from Complex I to
Coenzyme Q to Complex III. Complex II, the succinate dehydrogenase complex, is a
separate starting point, and is not a part of the NADH pathway.
From Complex III the pathway is to cytochrome c then to a Complex IV (cytochrome
oxidase complex). More protons are translocated by Complex IV, and it is at this site that
oxygen binds, along with protons, and using the electron pair and remaining free energy,
oxygen is reduced to water. Since molecular oxygen is diatomic, it actually takes two
electron pairs and two cytochrome oxidase complexes to complete the reaction sequence
for the reduction of oxygen. This last step in electron transport serves the critical function
of removing electrons from the system so that electron transport can operate
continuously.
The reduction of oxygen is not an end in itself. Oxygen serves as an electron acceptor,
clearing the way for carriers in the sequence to be reoxidized so that electron transport
can continue. In your mitochondria, in the absence of oxygen, or in the presence of a
poison such as cyanide, there is no outlet for electrons. All carriers remain reduced and
Krebs products become out of balance because some Krebs reactions require NAD or
FAD and some do not. However, you don't really care about that because you are already
dead. The purpose of electron transport is to conserve energy in the form of a
chemiosmotic gradient. The gradient, in turn, can be exploited for the phosphorylation of
ADP as well as for other purposes. With the cessation of aerobic metabolism cell damage
is immediate and irreversible.
From succinate, the sequence is Complex II to Coenzyme Q to Complex III to
cytochrome c to Complex IV. Thus there is a common electron transport pathway beyond
the entry point, either Complex I or Complex II. Protons are not translocated at Complex
II. There isn't sufficient free energy available from the succinate dehydrogenase reaction
to reduce NAD or to pump protons at more than two sites.
Is the ETS a sequence?
Before the development of the fluid mosaic model of membranes, the ETS was pictured
as a chain, in which each complex was fixed in position relative to the next. Now it is
accepted that while the complexes form 'islands' in the fluid membrane, they move
independently of one another, and exchange electrons when they are in mutual proximity.
Textbooks necessarily show the ETS as a physical sequence of complexes and carriers.
This has the unintentional effect of implying that they are all locked in place. The fluid
nature of membranes allows electron exchange to take place in a test tube containing
membrane fragments.
The location of ETS complexes on the inner membrane has two major consequences. By
floating in two-dimensional space, the likelihood of carriers making an exchange is much
higher than if they were in solution in the three dimensional space of the matrix. They are
exposed to the matrix side of the membrane, of course, for access to succinate and
NADH, but have limited mobility. Second, the location of the ETS on the inner
membrane enables them to establish a chemiosmotic gradient.
Electron pathways and inhibition
Electron transport inhibitors act by binding one or more electron carriers, preventing
electron transport directly. Changes in the rate of dissipation of the chemiosmotic
gradient have no effect on the rate of electron transport with such inhibition. In fact, if
electron transport is blocked the chemiosmotic gradient cannot be maintained. No matter
what substrate is used to fuel electron transport, only two entry points into the electron
transport system are known to be used by mitochondria. A consequence of having
separate pathways for entry of electrons is that an ETS inhibitor can affect one part of a
pathway without interfering with another part. Respiration can still occur depending on
choice of substrate.
An inhibitor may competely block electron transport by irreversibly binding to a binding
site. For example, cyanide binds cytochrome oxidase so as to prevent the binding of
oxygen. Electron transport is reduced to zero. Breathe all you want - you can't use any of
the oxygen you take in. Rotenone, on the other hand, binds competitively, so that a trickle
of electron flow is permitted. However, the rate of electron transport is too slow for
maintenance of a gradient.
Chemiosmotic Gradient: Generation and
Maintenance
Energetics of proton translocation at Compex I
The energy needed to push protons out of the matrix and into the intermembrane space
comes from the oxidation of either reduced NAD (NADH) or reduced FAD (FADH2). In
the case of NADH, passage of an electron pair to Coenzme Q provides a "pulling" force.
That is, NADH is much less electronegative than Coenzyme Q, while the iron-sulfur
protein carriers in between are intermediate in electronegativity. The amount of available
free energy is 69.5 kJ/mole of NADH (kiloJoules per mole). The efficiency of electron
transport can be represented by the standard reduction potential difference, namely the
voltage generated by a redox reaction under standard biochemical conditions.
The standard reduction potential of NADH is -0.315V, while that of coenzyme Q is
0.045V (difference of 0.345 V). Therefore there is a strong 'pull' by Coenzyme Q on
electrons through the components of Complex I.
Just for the sake of understanding the principles, let Complex I (NADH dehydrogenase
complex), embedded in an intact inner membrane, be the only component of an
experimental electron transport system. We'll simply take the electrons from Coenzyme
Q when they reach it, so the system can keep going. The removal of protons from the
matrix and deposition of protons in the intermembrane space creates a concentration
difference of protons across the inner membrane. This is called the chemiosmotic
gradient. As the gradient builds up, more and more energy is required to push protons
across. When the amount of energy required to push protons reaches 69.5 kJ/mole,
electron transport has to stop. In fact, the second law of thermodynamics requires that
electron transport stop before the gradient builds up to that point.
If there was no way of draining energy from the system, electron transport could not
continue despite the presence of adequate substrate. However, a mitochondrion is always
in a steady state of respiration, in which the energy lost by processes that dissipate the
gradient is constantly replaced by electron transport.
Respiratory Control
The limitation placed on electron transport by the chemisosmotic gradient is termed
respiratory control. Mitochondria are said to exercise respiratory control as long as they
can restrict electron transport by means of the gradient. If the gradient is destroyed by
damaging the membranes, respiratory control is abolished and electron transport can run
freely.
Cause and effect relationship
Electron transport is driven by the free energy that is available from the energy carriers,
in turn obtained from substrates such as glutamate or Krebs intermediates. It is restricted
by the chemiosmotic gradient. The only way electron transport can proceed is to the
extent that the energy in the gradient is dissipated. In healthy mitochondria the
gradient is maintained. That is, electron transport keeps up with the utilization of the
energy stored in the gradient. Even in the presence of ADP, which allows ATP synthetase
to exploit the gradient, the chemiosmotic gradient is maintained at a set energy level.
Think of physical causes and effects when you attempt to describe respiratory control.
What drives electron transport? It can't be driven by a "need" to maintain the gradient,
because that implies a sense of purpose. The electron transport system is just a structure,
complex as it is. Electron transport is driven by the increasing affinities of successive
carriers for electrons, and by the availability of substrates to provide electrons and free
energy. It is restricted by the chemiosmotic gradient - electron transport can only go as
fast as energy is lost from the gradient. Anything that increases turnover of energy from
the gradient increases the rate of electron transport proportionally.
Here is an analogy that might help with the concept. Suppose that you are trying to blow
up a balloon. Blowing with all your might you succeed in filling the balloon so that it is
two feet in diameter. You cannot force any more air into it so all you are doing now is
holding pressure. Now some joker comes along and pokes a couple of holes in your
balloon, so that air leaks out at a controlled rate. You have maintained constant pressure,
so now you find yourself moving air into the balloon as it leaks out. The balloon
diameter, proportional to internal pressure, remains constant. If you plug one of the holes
you move less air. Open up more holes and you move more air. The electron transport
system applies constant "pressure," holding the gradient at a constant level. The rate of
electron tranport (analagous to air flow in the balloon example) varies as energy is
drained from the system at different rates. Adding ADP in vitro, for example, opens up
avenues for protons to be forced into the matrix, draining energy from the gradient. Its
addition is the equivalent of poking additional holes in your balloon. Electron transport
spontaneously increases.
Respiratory Control in Exercise
Sometimes in wading through all of the details, one loses sight of the big picture. Just
why is all of this information about electron transport and oxidative phosphorylation so
important, anyhow? Suppose you are vegging in front of the TV - in the prone position
with a bag of cholesterol chips. You are very relaxed and are hardly aware that you are
breathing at all. Suddenly the fire alarm sounds, and a bunch of naked people are running
down the hall. This definititely provokes activity on your part, and you run down the hall
also. You start breathing heavily (due to the exercise, not the sight of the naked people).
Your demand for energy is rather low when you are relaxed. However running is heavy
exercise. Your muscle activity causes an immediate demand for ATP which is met in part
by muscle glycogen. When that runs low you need to replace your reserves with aerobic
metabolism, that is, your mitochondria need to make more ATP. Electron transport is
stimulated when the ratio of ATP to ADP goes down. The rate of binding of ADP to the
ATP synthetase automatically increases as more ADP is transported into the matrix. In
turn, electron transport is allowed to speed up - consuming more oxygen in the process.
Sensors in your cardiovascular system (primarily the carotid bodies of the carotid
arteries) detect an increase in carbon dioxide (indicating an increased need for oxygen).
The sensors send a signal to the brain via the nervous system that you need to speed up
breathing.
So... you breathe primarily to get oxygen into the blood and to the tissues, in order to
deliver oxygen to the mitochondria for cellular respiration - to make ATP. In the process,
carbon dioxide is returned to the lungs and exhaled. The processes are essential to your
well being, in fact, to your being at all. When you think of the effects of some of the
poisons that are discussed on these pages, think of the consequences to an intact organism
such as yourself.
The big picture, including regulatory pathways such as the very simplified description
presented above, are in the realm of integrative disciplines such as physiology. Such
disciplines are increasingly ignored in both research and academic curricula in favor of a
purely molecular approach to science. Please consider the importance of knowing the
consequences of molecular and cellular processes to the whole organism, and the fact that
the regulatory pathways controlling those processes are still incompletely understood.
Uncoupling and the basal metabolic rate
Endotherms like ourselves (formerly called homeotherms) maintain a constant body
temperature. The setpoint for that temperature is determined by a region the
hypothalamus (part of the brain, for you newcomers). But just how is the temperature
regulated? That is, how does the body maintain a balance between heat production and
heat loss?
The answer to that one is very complex. There are many mechanisms for heat dissipation
and retention (sweating, changes in distribution of the circulation, shivering, piloerection
- no, it's not a dirty word, etc.). Heat production results from the loss of some of the free
energy of every chemical reaction as heat - that is, every chemical reaction results in the
loss of some of the total free energy in the universe. That's one of the laws of
thermodynamics - I forgot which. A major source of heat production is electron transport
in mitochondria.
How do mitochondria produce heat? Much of the energy released by electron transport
does no useful work at all (that law of thermodynamics is at work again). That is, each
exchange of electrons results in a loss of some energy as heat. Furthermore, dissipation of
the chemiosmotic gradient without doing work results in the transformation of a great
deal more energy to heat. The mechanism of respiratory control can be exploited to
increase heat production by dissipating the gradient at a faster rate, or to slow heat
production by making the ETS more efficient. One example of that type of regulation is
the hormone thyroxine (thyroid hormone).
Thyroid hormone has many functions, mostly associated with promotion of anabolic
activity - synthesis of compounds and cellular growth. However thyroxine is also a
mildly effective uncoupling agent. Uncoupling agents dissipate the chemical gradient,
usually by creating a mechanism by which protons can escape the intermembrane space
(or otherwise 'short out' the gradient), allowing an increase in electron transport. In colder
regions of the world a noticible change in thyroid hormone levels takes place
circannually, that is, levels are higher in winter and lower in summer.
Oxidative phosphorylation
The relationship between synthesis (phosphorylation) of ATP and electron transport (the
last part of oxidative metabolism) often confuses students. Here are some facts that may
help dispel misconceptions about oxidative phosphorylation.
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ATP synthase is not part of the electron transport system (ETS)
Oxygen consumption results from electron transport and does not require ATP
synthesis
Protons entering the matrix through ATP synthase do not reduce oxygen
The ETS cannot transport electrons if protons cannot be translocated into the
intermembrane space
The number of protons translocated by proton pumps do not corrrelate directly
with number of ATP molecules synthesized
The ETS does not "want" to maintain or restore a chemiosmotic gradient –
electron transport is driven by the proximity of reduced and oxidized carriers,
facilitating exchange of electrons and free energy
As long as substrate is present a chemiosmotic gradient is maintained (unless
mitochondria are poisoned)
Activation of ATP synthase does not "lower" the gradient – it increases the rate at
which energy is removed from the gradient; the ETS maintains the gradient at a
constant level
ATP synthase consists of two functional units, one that conducts the passage of protons
(F0) and one that catalyzes the phosphorylation of ADP (F1). Both units must be
functional for ATP synthesis to take place. The F0 subunit (actually "F sub-zero," the
zero is a subscript) can be pictured as a rotor while the F1 subunit remains stationary. F0
includes a "gamma" subunit that rotates as protons are driven through the channel created
by the c ring component of F0.
The following is a simplistic description of the Boyer model for proton-driven ATP
synthesis. The F1 subunit includes three "beta" subunits that are identical in structure but
not in form. The conformation of each beta subunit changes as the gamma subunit of F0
rotates. At any given time one beta subunit is in the "loose" (L) conformation, which
binds ADP and inorganic phosphate. In that conformation the subunit is constrained so
that it does not release either molecule. A beta subunit in "tight" (T) conformation binds
ATP with such tenacity that it readily converts ADP and inorganic phosphate to ATP. A
subunit in the T conformation cannot release ATP, however. In the "open" (O)
conformation a beta subunit releases bound nucleotides.
Even if there is no proton gradient one beta subunit will be in the T form with bound
ATP, which forms spontaneously even in the absence of a proton gradient. The role of
the gradient is to cause the release of bound ATP, not to cause its synthesis. Once ATP is
released, binding of ADP and inorganic phosphate is spontaneous. Of course, both
reactants must be present for the system to operate. A complete cycle takes place as
follows. As protons are driven through the c ring their passage causes rotation of the
gamma subunit. As the subunit rotates it causes conversion of the T form (with bound
ATP) to the O form. ATP is then released. Meantime, the subunit in L form (which holds
bound ADP and inorganic phosphate) is converted to the T form which results in
conversion of ADP and phosphate to ATP. The subunit that had been in the O
conformation is converted into the L form, binding and "locking" ADP and inorganic
phosphate in place.
For ATP synthesis to continue ADP and inorganic phosphate must both be available and
the ETS must be capable of conducting electron transport and storing energy as a
chemiosmotic gradient. Each proton pump translocates a specific number of protons
(from 2 to 4) with each passage of an electron pair. A specific number of protons must be
driven through the F0 subunit of ATP synthase to accomplish one complete cycle.
Because some energy stored in the gradient is always lost as heat or exploited for
processes other than ATP synthesis, there is not a one to one correspondence between
number of protons translocated by the ETS and number of protons entering the matrix
through ATP synthase.