Signal Transduction
INTRODUCTION
In order for an organ or cell to function properly within an organism, it must respond to cues from
distant cells as well as from its local environment.
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Cells in different organs communicate with one another through extracellular signaling
molecules released by one set of cells and received by the other.
Not all molecules can pass through the lipid bilayer of a cell, and so signal transduction
systems are used in order to relay an external signal to the cell interior.
This exercise presents a general overview, as well as a few well-characterized examples of different
signal transduction pathways. It is important to note that there are a large number of signal
transduction pathways with widely varied mechanisms, many of which are not shown here. Upon
completion of this unit, you will understand the primary features of signal transduction systems, as
well as become familiar with some specific signal transduction systems and the metabolic role they
play in connection with hormones.
SIGNAL PATHWAYS
All signal transduction pathways include the followin:
1. An extracellular ligand that does not penetrate through the cell membrane.
2. A cell surface receptor that is generally a transmembrane protein.
3. A conformational change in the receptor that occurs when the ligand binds to it.
Generally, an extracellular signal activates many different signal transduction pathways that lead to a
number of cellular responses.
AMPLIFICATION CASCADES
The conformational change in the membrane receptor that occurs upon ligand binding often initiates
several types of enzyme-driven effects within a cell, which typically involve enzymes called kinases.
Kinases transfer a phosphoryl group from ATP onto other proteins.
Because the stimulated enzymes are catalytic, and produce multiple products upon activation, a
cascade-type response occurs, which amplifies the signal. Thus, the ligand binding event is greatly
amplified by the time the final target molecules are produced.
However, these enzymatic cascades cannot be left to run indefinitely. Signal transduction cascades
must be controlled or regulated very tightly, or there can be dire consequences for the proper
functioning of the cell. To this end, signal transduction pathways may have a built in "off" switch. The
same signal that initiates a cellular response may also activate a mechanism for shutting down that
response. For example, the activation of a kinase often triggers the activation of a phosphatase, an
enzyme that removes the phosphoryl group and thereby inactivates proteins activated by the kinase.
G PROTEINS
One of the most common types of signal transduction pathways involves modulating adenylyl
cyclase via GTP-binding proteins, more commonly known as G proteins.
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G proteins are heterotrimers.
They are associated with a membrane-bound receptor molecule.
In its inactive form, a GDP is attached to the  subunit.
When the ligand binds to the receptor, the GDP is exchanged for GTP. The  subunit dissociates
from the  and  subunits and the receptor and attaches to a membrane-bound adenylyl cyclase.
The  subunit activates adenylyl cyclase, stimulating the synthesis of cyclic AMP, or cAMP. cAMP
belongs to a group of intracellular signal molecules called second messengers (to distinguish them
from the ligand, or first messenger). cAMP activates protein kinases in many hormonally regulated
processes.
The  subunit is a slow GTPase, and thus the GTP is hydrolyzed to GDP after a period of time.
The  subunit then reassociates with the  and  subunits and the adenylyl cyclase is inactivated.
It is important to note that there are two types of signal amplification in this particular system:
1. A single ligand/cell membrane receptor complex can activate many G proteins.
2. Until the GTP is hydrolyzed on the  subunit of the G protein, leading to dissociation, a
single activated adenylyl cyclase can continue to synthesize many molecules of cAMP.
ADENYLYL CYCLASE
Catecholamine hormones such as epinephrine and norepinephrine are produced by the adrenal
gland, and are released during times of stress. These hormones bind to - and -adrenergic
receptors, thereby generating stimulatory signals. Although these hormones act differently on
different tissues, the transduction pathways occurs via adenylyl cyclase stimulation.
Opioid drugs such as codeine, on the other hand, act to inhibit adenylyl cyclase via the 2adrenergic receptor. This inhibitory G-protein trimer functions in the same manner as the stimulatory
G-protein trimer, with the exception that it blocks adenylyl cyclase from producing cAMP.
The 2 receptor can also indirectly inhibit the activity of adenylyl cyclase. After the inhibitory G
protein is activated and its - subunit disassociates, the remaining  and  subunits can compete for
the  subunits of the stimulatory G proteins.
PROTEIN KINASE A
One of the proteins activated by cAMP is a kinase called protein kinase A. In the absence of cAMP,
this kinase is an inactive tetramer made up of two regulatory (R) subunits and two catalytic (C)
subunits.
When cAMP binds to the regulatory subunits, the tetramer releases its two active catalytic subunits.
Consequently, the level of cAMP determines the level of activity of protein kinase A.
Protein kinase A phosphorylates many enzymes. One of the targets of protein kinase A is
phosphorylase kinase.
Phosphorylase kinase phosphorylates glycogen phosphorylase, activating it. Glycogen
phosphorylase is responsible for glycogenolysis—the breakdown of glycogen.
Phosphorylase kinase also phosphorylates glycogen synthase, deactivating it. Glycogen synthase is
responsible for glycogen synthesis. By deactivating glycogen synthase and activating glycogen
phosphorylase, phosphorylase kinase prevents the cell from synthesizing and degrading glycogen at
the same time.
CONCLUSION
In signal transduction, compounds that cannot pass through the membrane have intracellular effects.
Signal transduction cascades can lead to a large amplification of signal from a single binding event.
Citric Acid Cycle
INTRODUCTION
The citric acid cycle is a central metabolic pathway that completes the oxidative degradation of fatty
acids, amino acids, and monosaccharides. During aerobic catabolism, these biomolecules are
broken down to smaller molecules that ultimately contribute to a cell’s energetic or molecular needs.
Early metabolic steps, including glycolysis and the activity of the pyruvate dehydrogenase complex,
yield a two-carbon fragment called an acetyl group, which is linked to a large cofactor known as
coenzyme A (or CoA). It is during the citric acid cycle that acetyl-CoA is oxidized to the waste
product, carbon dioxide, along with the reduction of the cofactors NAD+ and ubiquinone.
The citric acid cycle serves two main purposes:
1. To increase the cell’s ATP-producing potential by generating a reduced electron carriers
such as NADH and reduced ubiquinone; and
2. To provide the cell with a variety of metabolic precursors.
Upon completion of this exercise, you should:
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Be able to describe the sources of acetyl groups that enter the citric acid cycle;
Trace the conversion of substrates to products through each of the citric acid cycle’s eight
reactions and understand how flux through the cycle is regulated;
Understand the energetic output of the citric acid cycle;
Describe the role of the reduced electron carriers and their role in coupling the citric acid
cycle to downstream reactions that produce ATP;
Describe the amphibolic character of the citric acid cycle; and
Understand the reactions that replenish citric acid cycle intermediates.
CELLULAR LOCATION
Both prokaryotic and eukaryotic cells use the citric acid cycle to help meet their energetic and
molecular needs. In respiring prokaryotes, the citric acid cycle takes place in the cytosol. In
eukaryotic cells, such as the cells of the human body, the cycle takes place within the mitochondrial
matrix.
CATABOLISM
The reactions of the citric acid cycle oxidize acetyl-CoA’s acetyl group to two molecules of carbon
dioxide. During the reaction cycle, electrons are transferred from acetyl-CoA to electron carriers.
Once an electron carrier accepts an electron, it is referred to as “reduced.” Ultimately, reduced
electron carriers participate in downstream reaction pathways that generate ATP, the energy
currency of the cell.
Note that one high-energy nucleoside triphosphate is generated directly from the reaction cycle.
Because acetyl-CoA is broken down to smaller molecules during the citric acid cycle, the citric acid
cycle is described as catabolic.
ANABOLISM AND CATABOLISM
In addition to catabolizing molecules to meet cellular energetic needs, the citric acid cycle is key in
supplying various biochemical pathways with precursors needed for synthesizing molecules.
Reactions that involve “building” molecules from smaller parts are referred to as anabolic. Anabolic
reactions use citric acid cycle intermediates as precursors for fatty acid, amino acid, and
carbohydrate synthesis. These anabolic processes may also require reduced cofactors.
Many citric acid cycle intermediates serve the cell as both reaction precursors and reaction products.
For example, -ketoglutarate may act as a precursor for amino acids in an anabolic pathway, or it
may be catabolized to carbon dioxide during the reactions of the citric acid cycle. As such, the citric
acid cycle is neither purely anabolic nor purely catabolic. Reactions that possess this dual character
of building and degrading molecules are considered amphibolic.Amphi is a Greek prefix
meaning both.
SOURCES OF ACETYL-COA
The skeleton drawings of the monosaccharide glucose, the fatty acid palmitic acid, and the amino
acids lysine and glutamate are depicted. These molecules are degraded to a common compound
called acetyl-CoA, the initiator of the citric acid cycle. Select the various molecules to learn how each
compound ultimately enters the citric acid cycle as acetyl-CoA. Then consider how the efficiency of
metabolism would change if a common product of carbohydrate, fatty acid, and amino acid
catabolism did not exist.
Fatty Acids
Many different fatty acids exist, although their structures can be generalized as a carboxylic acid with
a long, hydrocarbon tail. Palmitate is an example of a sixteen-carbon fatty acid.
When a cell’s metabolic needs increase, free fatty acids enter the mitochondrion where the
degradative reactions called  oxidation ensue. A fatty acid shortened by two carbon atoms plus a
free acetyl-CoA molecule results from each round of  oxidation. Acetyl-CoA initiates the citric acid
cycle.
Amino Acids
Examine the structures of glutamate and lysine. Recall that an amino acid consists of an amino and
a carboxyl group at opposite ends, plus an attached side chain.
In the case of starvation, protein degradation increases and the free amino acids that result may be
used as a source of metabolic fuel. Alternatively, if an organism’s intake of free amino acids exceeds
its protein-building needs, the free amino acids are metabolized, for there is no storage mechanism
for excess amino acids.
Typically, the amino group of an amino acid is removed in a deamination reaction. The remaining
carbon skeleton is broken down to various products depending on which of the twenty amino acids is
undergoing catabolism. In some cases, the remaining carbon skeleton is broken down to acetyl-CoA
or to pyruvate, which is then converted to acetyl-CoA. Alternatively, a citric acid cycle intermediate
such as -ketoglutarate may result. In all cases, the citric acid cycle plays a large role in breaking
down the amino acid skeleton to carbon dioxide.
For example, catabolism of lysine yeilds carbon dioxide and acetyl-CoA, while glutamate breaks
down to -ketoglutarate, carbon dioxide, and acetyl-CoA. Acetyl-CoA initiates the citric acid cycle.
Monosaccharides
The monosaccharide glucose is a six-carbon sugar. In the case of higher eukaryotes, a cell most
commonly acquires glucose in two ways—by breaking down complex carbohydrates into simple
sugars and by mobilizing glucose from glycogen, the body’s storage system for glucose.
In the cytosol, glucose is broken down to two, 3-carbon molecules during glycolysis. The resulting
three-carbon molecules are called pyruvate. Pyruvate is transported across the mitochondrial
membrane where it is broken down to a 2-carbon compound called acetyl-CoA plus carbon dioxide.
Acetyl-CoA initiates the citric acid cycle
REACTANTS AND PRODUCTS
Acetyl-CoA is further oxidized in the citric acid cycle. As you learn each step in the reaction cycle,
keep in mind that additional substrates are necessary to complete one full turn of the reaction cycle,
including one GDP, one inorganic phosphate, three NAD+, and one ubiquinone, commonly referred
to as, Q.
Products that emerge from one turn of the citric acid cycle are two carbon dioxide molecules, one
CoA, one GTP, three NADH, and one reduced ubiquinone, referred to as QH2.
CYCLICAL REACTION PATHWAY
We will now examine each of the eight reactions that make up the citric acid cycle. Consider this
screen your “home base” for the reaction cycle as you study each of the eight reactions in more
detail. Investigate each reaction by clicking on its reaction number.
FATE OF ACETYL-COA CARBON
You have learned that acetyl-CoA and other reaction intermediates lose electrons in a series of
oxidation reactions during the citric acid cycle. For every acetyl-CoA molecule that enters the citric
acid cycle, a total of four pairs of electrons are lost to electron carriers during the oxidation of two
carbon atoms. These two oxidized carbon atoms are released as two molecules of carbon dioxide.
Two carbon atoms are released as carbon dioxide during one round of the citric acid cycle. These
carbon atoms do not originate from the acetyl-CoA molecule that initiated the reaction cycle. Acetyl
carbons are released during subsequent rounds of the circular pathway.
Trace the fate of acetyl-CoA carbon atoms through two rounds of the citric acid cycle, paying
particular attention to which carbon atoms are oxidized, and when.
REGULATION: INHIBITION
The body functions like a finely tuned machine because its internal activities are coordinated and
regulated. For example, after you finishes a heavy meal, your stomach swells and stretch receptors
lining the stomach send messages to your brain queueing you to, “stop eating!” On a much smaller
scale, after a buildup of citric acid cycle products and intermediates accumulate, these compounds
affect enzyme activity near and far, to greatly decrease cycling of the citric acid reactions.
Progression through the citric acid cycle is illustrated. Notice that with each successive cycle, the
levels of key regulatory compounds increase. The key regulatory compounds that act to decrease
the level of citric acid cycle activity are citrate, NADH, and succinyl-CoA. Collectively, these
regulatory compounds function as inhibitors of the citric acid cycle.
REGULATION: ACTIVATION
In contrast to the inhibitory control previously described, positive regulators, called activators,
function to up-regulate the activity of the citric acid cycle when the cell’s energetic or molecular
needs are not met. Key activators include Ca2+ and ADP, which signal to increase the activity of
isocitrate dehydrogenase and -ketoglutarate dehydrogenase. Ca2+ and ADP generally signify the
need to generate cellular free energy.
ENERGETICS
During the 1860s, Louis Pasteur conducted a series of experiments to determine if the rate of
glucose metabolism is dependent on the presence or absence of oxygen. His experimental set-up
was similar to that shown on your screen, and his findings were coined the Pasteur effect.
Take a look at the two yeast cells illustrated. One is living in an aerobic environment, the other in an
anaerobic environment.
Anaerobic
When a cell living under anaerobic conditions is working to meet its metabolic needs, the reactions
of glycolysis are turned on. For every glucose molecule consumed, the cell produces a net of two
ATPs and two reduced NADH electron carriers during glycolysis. In the absence of oxygen,
however, oxidative phosphorylation can not take place. Therefore this metabolic pathway largely
responsible for transferring electrons from NADH to molecular oxygen to produce ATP does not
exist. Therefore, under anaerobic conditions, energy is derived solely from ATP produced during
glycolysis.
Aerobic
In a cell living in aerobic conditions, ATP is generated by three metabolic pathways: glycolysis, the
citric acid cycle, and oxidative phosphorylation. Reduced electron carriers that emerge from
glycolysis and the citric acid cycle are funneled to the electron transport chain, where they
participate in a series of oxidation and reduction reactions. This establishes a proton gradient that
spans the inner mitochondrial membrane, which ultimately drives the oxidative phosphorylation of
ADP to ATP. Therefore, under aerobic conditions, NADH and reduced ubiquinone, or QH2, serve the
cell by increasing its ATP-producing potential.
ANAPLEROTIC REACTIONS
Imagine a muscle cell that is beginning to deplete its energy stores due to vigorous exercise. The
rate of glucose consumption increases along with a pooling of the glycolytic product, pyruvate. If
molecular oxygen remains readily available, pyruvate is converted to acetyl-CoA, thereby activating
the citric acid cycle. The activities of citric acid enzymes are up-regulated, in large part, by an
increase in the levels of the activators, calcium ions, and ADP.
Reactions exist to replenish the cell with citric acid cycle intermediates, which is especially important
when metabolic activity increases, as in the case of vigorous exercise. The catabolism of three types
of compounds “feed” the citric acid cycle at different points. Reactions that “feed” the citric acid cycle
with intermediates are called anaplerotic.
DNA Replication
INTRODUCTION
DNA replication is the process whereby an entire double-stranded DNA is copied to produce a
second, identical DNA double helix.
The objectives of this exercise are:
1. To understand the functions of the proteins responsible for DNA replication, including
helicase, SSB protein, primase, the sliding clamp, DNA polymerase, Rnase H and DNA
ligase.
2. to understand why the leading strand is synthesized continuously and the lagging strand is
synthesized discontinuously.
THE REPLICATION FACTORY
DNA replication is an intricate process requiring the concerted action of many different proteins. The
replication proteins are clustered together in particular locations in the cell and may therefore be
regarded as a small “Replication Factory” that manufactures DNA copies. The DNA to be copied is
fed through the factory, much as a reel of film is fed through a movie projector. The incoming DNA
double helix is split into two single strands and each original single strand becomes half of a new
DNA double helix. Because each resulting DNA double helix retains one strand of the original DNA,
DNA replication is said to be semi-conservative.
DNA REPLICATION PROTEINS
DNA replication requires a variety of proteins. Each protein performs a specific function in the
production of the new DNA strands. Helicase, made of six proteins arranged in a ring shape,
unwinds the DNA double helix into two individual strands. Single-strand binding proteins, or SSBs,
are tetramers that coat the single-stranded DNA. This prevents the DNA strands from reannealing to
form double-stranded DNA. Primase is an RNA polymerase that synthesizes the short RNA primers
needed to start the strand replication process. DNA polymerase is a hand-shaped enzyme that
strings nucleotides together to form a DNA strand. The sliding clamp is an accessory protein that
helps hold the DNA polymerase onto the DNA strand during replication. RNAse H removes the RNA
primers that previously began the DNA strand synthesis. DNA ligase links short stretches of DNA
together to create one long continuous DNA strand.
STRAND SEPARATION
Let’s look at the steps of DNA replication in more detail. To begin the process of DNA replication, the
two double helix strands are unwound and separated from each other by the helicase enzyme. The
point where the DNA is separated into single strands, and where new DNA will be synthesized, is
known as the replication fork. Single-strand binding proteins, or SSBs, quickly coat the newly
exposed single strands. SSBs maintain the separated strands during DNA replication. Without the
SSBs, the complementary DNA strands could easily snap back together. SSBs bind loosely to the
DNA, and are displaced when the polymerase enzymes begin synthesizing the new DNA strands.
NEW STRAND SYNTHESIS
Now that they are separated, the two single DNA strands can act as templates for the production of
two new, complementary DNA strands. Remember that the double helix consists of two antiparallel
DNA strands with complementary 5’ to 3’ strands running in opposite directions. Polymerase
enzymes can synthesize nucleic acid strands only in the 5’ to 3’ direction, hooking the 5’ phosphate
group of an incoming nucleotide onto the 3’ hydroxyl group at the end of the growing nucleic acid
chain. Because the chain grows by extension off the 3’ hydroxyl group, strand synthesis is said to
proceed in a 5’ to 3’ direction.
Even when the strands are separated, however, DNA polymerase cannot simply begin copying the
DNA. DNA polymerase can only extend a nucleic acid chain but cannot start one from scratch. To
give the DNA polymerase a place to start, an RNA polymerase called primase first copies a short
stretch of the DNA strand. This creates a complementary RNA segment, up to 60 nucleotides long
that is called a primer.
Now DNA polymerase can copy the DNA strand. The DNA polymerase starts at the 3’ end of the
RNA primer, and, using the original DNA strand as a guide, begins to synthesize a new
complementary DNA strand. Two polymerase enzymes are required, one for each parental DNA
strand. Due to the antiparallel nature of the DNA strands, however, the polymerase enzymes on the
two strands start to move in opposite directions.
One polymerase can remain on its DNA template and copy the DNA in one continuous strand.
However, the other polymerase can only copy a short stretch of DNA before it runs into the primer of
the previously sequenced fragment. It is therefore forced to repeatedly release the DNA strand and
slide further upstream to begin extension from another RNA primer. The sliding clamp helps hold this
DNA polymerase onto the DNA as the DNA moves through the replication machinery. The sliding
clamp makes the polymerase processive.
The continuously synthesized strand is known as the leading strand, while the strand that is
synthesized in short pieces is known as the lagging strand. The short stretches of DNA that make up
the lagging strand are known as Okazaki fragments.
THE LAGGING STRAND
Before the lagging-strand DNA exits the replication factory, its RNA primers must be removed and
the Okazaki fragments must be joined together to create a continuous DNA strand. The first step is
the removal of the RNA primer. RNAse H, which recognizes RNA-DNA hybrid helices, degrades the
RNA by hydrolyzing its phosphodiester bonds. Next, the sequence gap created by RNAse H is then
filled in by DNA polymerase which extends the 3’ end of the neighboring Okazaki fragment. Finally,
the Okazaki fragments are joined together by DNA ligase that hooks together the 3’ end of one
fragment to the 5’ phosphate group of the neighboring fragment in an ATP- or NAD+-dependent
reaction.
REPLICATION IN ACTION
We are now ready to review the steps of DNA replication.
1. The process begins when the helicase enzyme unwinds the double helix to expose two
single DNA strands and create two replication forks. DNA replication takes place
simultaneously at each fork. The mechanism of replication is identical at each fork.
Remember that the proteins involved in replication are clustered together and anchored in
the cell. Thus, the replication proteins do not travel down the length of the DNA. Instead, the
DNA helix is fed through a stationary replication factory like film is fed through a projector.
2. Single-strand binding proteins, or SSBs, coat the single DNA strands to prevent them from
snapping back together. SSBs are easily displaced by DNA polymerase.
3. The primase enzyme uses the original DNA sequence as a template to synthesize a short
RNA primer. Primers are necessary because DNA polymerase can only extend a nucleotide
chain, not start one.
4. DNA polymerase begins to synthesize a new DNA strand by extending an RNA primer in the
5' to 3' direction. Each parental DNA strand is copied by one DNA polymerase. Remember,
both template strands move through the replication factory in the same direction, and DNA
polymerase can only synthesize DNA from the 5’ end to the 3’ end. Due to these two factors,
one of the DNA strands must be made discontinuously in short pieces which are later joined
together.
5. As replication proceeds, RNAse H recognizes RNA primers bound to the DNA template and
removes the primers by hydrolyzing the RNA.
6. DNA polymerase can then fill in the gap left by RNase H.
7. The DNA replication process is completed when the ligase enzyme joins the short DNA
pieces together into one continuous strand
Fatty Acid Metabolism
LEARNING OBJECTIVES
Fatty acids are an important energy source, for they yield over twice as much energy as an equal
mass of carbohydrate or protein. In humans, the primary dietary source of fatty acids is
triacylglycerols. This exercise will describe the metabolism of fatty acids. The two main components
of fatty acid metabolism are oxidation and fatty acid synthesis. Upon completion of this exercise,
you will understand that the fatty-acid breakdown reactions of  oxidation result in the formation of
reduced cofactors and acetyl-CoA molecules, which can be further catabolized to release free
energy. You will also understand that the oxidation of unsaturated, odd-chain, and very-long-chain
fatty acids requires additional enzymes, some of them in peroxisomes. In addition, you will
understand how fatty acid synthesis resembles and differs from  oxidation.
FATTY ACID ACTIVATION
Triacylglycerols are carried by lipoproteins to tissues, where hydrolysis releases their fatty acids from
the glycerol backbone. Fatty acids enter the cell and are activated in the cytosol. This activation
costs two ATP equivalents per fatty acid. Most of the activated fatty acids are then shuttled into the
mitochondria for  oxidation, but a small percentage are carried to the peroxisomes.
STEPS OF  OXIDATION
The activated fatty acid is called a fatty acyl-coenzyme A, or fatty acyl-CoA. In the first step
of  oxidation, an acyl-CoA dehydrogenase catalyzes the oxidation of the acyl group, resulting in the
formation of a double bond between carbons two and three. The two electrons removed from the
acyl group are transferred to an FAD prosthetic group. These electrons are transferred to ubiquinone
through a series of electron transfer reactions. In the second step of  oxidation, a hydratase adds a
molecule of water across the double bond produced in the first step. In the third step of  oxidation,
another dehydrogenase catalyzes the oxidation of the hydroxyacyl group. In this case, NAD+ is the
cofactor. The fourth and final step of  oxidation is called thiolysis. In this step, a thiolase catalyzes
the release of acetyl-CoA from the ketoacyl-CoA.
ENERGY YIELD OF OXIDATION
One round of  oxidation yields three products—one ubiquinol cofactor, one NADH cofactor, and
one molecule of acetyl-CoA. During the citric acid cycle, the acetyl-CoA is used to produce three
NADH cofactors, one ubiquinol cofactor, and one molecule of GTP. During oxidative
phosphorylation, each ubiquinol cofactor is used to produce two ATP molecules, and each NADH
cofactor is used to produce three ATP molecules. The GTP molecule is equivalent to one ATP
molecule. In all, one round of  oxidation produces the equivalent of 17 molecules of ATP. Since two
ATP equivalents were used for the activation step, the net yield is 15 molecules of ATP.
OXIDATION OF PALMITATE
The fatty acid we started with was palmitate. Let’s determine the energy yield for the
complete  oxidation of this 16-carbon fatty acid. Palmitate goes through seven rounds
of  oxidation, each of which yields products equivalent to seventeen molecules of ATP. The final
product of complete  oxidation is an additional molecule of acetyl-CoA, which is equivalent to
twelve molecules of ATP. In all, the complete  oxidation of palmitate produces 131 molecules of
ATP. Subtracting the initial ATP investment for activation yields 129 molecules of ATP from a single
molecule of palmitate.
UNSATURATED FATTY ACIDS
Many common fatty acids contain cis double bonds. These double bonds present an obstacle to the
enzymes of  oxidation. Let’s follow the oxidation of linoleate to see how these metabolic obstacles
are removed.
The first three rounds proceed normally. However, the enoyl-CoA that begins the fourth round has a
double bond between the third and fourth carbon atoms and is not recognized by acyl-CoA
dehydrogenase. Instead, an enoyl-CoA isomerase converts the cis 3-4 double bond to a trans 2-3
double bond so that oxidation can continue. Since the enoyl-CoA isomerase reaction bypasses the
ubiquinol-producing step of this round of  oxidation, the energy yield for this round is 15 ATP
molecules, rather than 17.
Another problem arises in the fifth round. Step one proceeds as normal, but the resulting molecule
has two double bonds: one at the 2-3 position, and one at the 4-5 position. The enoyl-CoA hydratase
of step two cannot recognize this dienoyl-CoA. This problem is overcome by reducing the dienoyl
group, but the reaction requires an investment of one NADPH cofactor, which is equivalent to three
ATP molecules. After enoyl-CoA isomerase acts, the acyl group can continue through the pathway.
Linoleate goes through eight rounds of  oxidation. If linoleate did not have double bonds, this would
result in a total of 146 molecules of ATP. However, because of the corrections for the double bonds,
the total yield is 141 molecules of ATP per molecule of linoleate. In general, double bonds that begin
at odd-numbered positions cost the equivalent of two ATP molecules, and double bonds beginning
at even-numbered positions cost the equivalent of three ATP molecules.
ODD-CHAIN FATTY ACIDS
Most fatty acids have an even number of carbon atoms, since they are built from two-atom acetyl
units, as we’ll see momentarily. However, some plant and bacterial fatty acids have an odd number
of carbon atoms. Such odd-chain fatty acids yield a three-carbon propionyl-CoA after the final round
of  oxidation. This intermediate is further metabolized through a series of reactions, both in the
mitochondria and in the cytosol. Essentially, to get rid of the one extra carbon atom, one ATP
molecule was invested, and the process produced the equivalent of nine additional ATP. Therefore,
to calculate the energy yield of the complete  oxidation of an odd-chain fatty acid, add eight to the
total for a fatty acid with one less carbon atom.
VERY-LONG-CHAIN FATTY ACIDS
Fatty acids with chains that contain twenty-two or more carbon atoms are called very-long-chain fatty
acids. While shorter fatty acids are oxidized in the mitochondria, very-long-chain fatty acids
begin  oxidation in the peroxisomes. This process is almost identical to  oxidation in the
mitochondria, with one key difference. Instead of reducing ubiquinone in the first step, the
peroxisomes produce hydrogen peroxide. This peroxide can be used in other reactions to oxidize
toxic substances in the cell. Each round of  oxidation in the peroxisomes produces the equivalent of
fifteen molecules of ATP—two less than oxidation in the mitochondria. However, peroxisomes
usually do not completely degrade the fatty acids. Because the enzymes in peroxisomes have a low
affinity for short-chain fatty acids, shortened fatty acids are transported to the mitochondria to
finish  oxidation.
SYNTHESIS VS. OXIDATION
At first glance, fatty acid synthesis appears to be the exact reverse of  oxidation—fatty acyl groups
are built and degraded two carbon atoms at a time, and several of the reaction intermediates in the
two pathways are similar or identical. However, the pathway for fatty acid synthesis cannot be the
exact reverse of oxidation; since  oxidation is thermodynamically favorable, the reverse process is
thermodynamically unfavorable. Thus, fatty acid synthesis requires a large investment of energy in
the form of ATP.
STEPS OF SYNTHESIS
Let’s take a closer look at the steps of fatty acid synthesis. Before fatty acid synthesis can begin, an
acetyl group must be transferred from coenzyme-A to an acyl carrier protein, called ACP. The first
step in the cycle adds a two-carbon unit to the growing fatty acid. The two carbon atoms come from
malonyl-CoA, which is produced from acetyl-CoA in a reaction requiring one molecule of ATP. In the
second step, NADPH is used to reduce the ketoacyl-ACP from step one. In the third step,
hydroxyacyl-ACP dehydrase catalyzes the removal of a water molecule from the hydroxyacyl-ACP
produced in step two. In the fourth step, a second NADPH-dependent reduction converts the enoylACP produced in step three to a fatty acyl-ACP two carbon atoms longer than the starting substrate.
In all, adding two carbon atoms to the fatty acid costs the cell one ATP and two NADPH molecules.
PALMITATE SYNTHESIS
Palmitate synthesis requires seven rounds of fatty acid synthesis. In all, this costs the cell 49 ATP
equivalents. After the final round of fatty acid synthesis, a fatty acyl thioesterase catalyzes the
removal of the fatty acid from the acyl carrier protein.
FATTY ACID SYNTHASES
In bacteria and chloroplasts, fatty acid synthesis is carried out by several enzymes. In mammals, the
main reactions of fatty acid synthesis are carried out by one multifunctional enzyme made of two
identical polypeptides. Packaging several enzyme activities into one multifunctional protein like
mammalian fatty acid synthase allows the enzymes to be synthesized and controlled in a
coordinated fashion. Also, the product of one reaction can quickly diffuse to the next active site.
CONCLUSION
Fatty acid metabolism is important to the function of many cells. Note that in fatty acid synthesis, the
chain is extended two carbon atoms at a time, at the expense of ATP. In fatty acid oxidation, the
chain is degraded two carbon atoms at a time, producing ATP. The two pathways are regulated so
that a cell can synthesize energy-storing fatty acids in times of plenty, and oxidize the fatty acids
when the cell needs to generate ATP.
Lipoproteins
INTRODUCTION
Cholesterol has frequently been in the news over the last several years. High levels of "bad
cholesterol" have been linked to atherosclerosis, cardiovascular disease, heart attacks, and
strokes. But what is "bad cholesterol?" When people refer to "bad cholesterol" and "good
cholesterol," they’re actually talking about lipoproteins—the complexes used by the body to
transport cholesterol and other lipids. This exercise will describe the structures of lipoproteins
and help you understand how different types of lipoproteins function in cholesterol metabolism.
MODELS
Before we investigate the differences between the different types of lipoproteins, let’s take a look
at what they have in common. The interiors of all lipoproteins are composed of cholesteryl esters
and triacylglycerols.
Triacylglycerols and cholesteryl esters are hydrophobic and are therefore only slightly soluble in
aqueous solutions such as the bloodstream. Lipoproteins solve this problem by coating the
hydrophobic interior with an amphiphilic layer of phospholipids and cholesterol.
The final components of lipoproteins are proteins called apolipoproteins or just apoproteins. At
least nine different apolipoproteins associate with human lipoproteins in substantial amounts, but
the structures of most of them are believed to be similar. Apolipoproteins have a high alpha helix
content. One side of the alpha helix tends to contain nonpolar residues, while the other contains
polar residues. The nonpolar residues associate with the nonpolar tails of the phospholipids in
the lipoprotein, while the polar residues associate with the polar head groups.
DIETARY CHOLESTEROL
Fats and cholesterol from the foods you eat are packaged into lipoproteins in the small intestine.
These lipoproteins are called chylomicrons. Chylomicrons are relatively large and consist mostly
of triacylglycerols. One of the functions of chylomicrons is to deliver dietary cholesterol to the
liver.
VLDL
Lipoproteins are classified by their density. Since the protein components of lipoproteins are
denser than the lipid components, lipoproteins with a smaller percentage of protein are lower in
density.
In the liver, cholesterol and triacylglycerols are packaged into very low-density lipoproteins, or
VLDL, for delivery to other tissues. VLDL are composed of only about 5% to 10% protein, and
about 50% to 65% triacylglycerols. VLDL have five different major apolipoproteins. These
proteins help to target the VLDL to muscle cells and fat cells.
LDL – “BAD CHOLESTEROL"
As VLDL travel throughout the body, they give up triacylglycerols and other lipids to muscle and
fat cells. As they do so, they become denser. Eventually, they lose all but one of their
apolipoproteins, becoming low-density lipoproteins, or LDL. LDL contain a high percentage of
cholesterol and cholesteryl esters. LDL receptors on the surfaces of cells bind the apolipoprotein
of LDL, allowing the cells to take up cholesterol through receptor-mediated endocytosis.
When the apolipoprotein binds to an LDL receptor, the LDL enters the cell in a coated vesicle.
The vesicle fuses with an endosome. The lower pH of the endosome causes the LDL to detach
from the LDL receptors, which are then recycled to the cell surface. The vesicle then merges
with a lysosome. The apolipoprotein is degraded into amino acids, and the cholesteryl esters are
converted to cholesterol. These components can then be used by the cell.
LDL has been linked to the disease atherosclerosis. This link is the reason LDL is often referred
to as "bad cholesterol." If more LDL is present in the blood than can be quickly taken up by cells,
the LDL and its cholesteryl esters accumulate in the walls of large blood vessels and produce
chemical signals that attract white blood cells. The white blood cells produce inflammation,
leading to formation of a plaque. This plaque can become coated in calcium, resulting in the
hardening of the arteries associated with atherosclerosis. Eventually, the blood vessel can
become constricted and can trigger formation of a blood clot. If the blood vessel supplies the
heart with blood, the result is a heart attack. If the blood vessel supplies the brain with blood, the
result is a stroke.
A disease called familial hypercholesterolemia is caused by a genetic defect in the LDL receptor.
The cells of individuals homozygous for this defect are unable to take up LDL, so the
concentration of serum cholesterol is about three times higher than normal. Individuals
homozygous for familial hypercholesterolemia often die of heart attacks as early as age five.
HDL – “GOOD CHOLESTEROL”
A different lipoprotein is responsible for removing excess cholesterol from the tissues and
returning it to the liver for disposal. This lipoprotein contains about 50% protein, and is therefore
much higher density than the other lipoproteins. For this reason, this lipoprotein is called highdensity lipoprotein, or HDL. Since HDL helps clear excess cholesterol from the body, it is often
referred to as "good cholesterol."
The transfer of cholesterol from cells to HDL requires several different cell-surface proteins. One
of these proteins is believed to be a transport protein or flippase that moves cholesterol from the
cytosolic leaflet of the cell membrane to the extracellular leaflet. The other proteins are
responsible for recognizing the HDL and converting the cholesterol to cholesteryl esters.
Defects in the gene for the flippase cause Tangier disease. Individuals with Tangier disease are
unable to efficiently clear excess cholesterol from their cells, resulting in an accumulation of
cholesterol in their tissues and a high risk of heart attack.
SUMMARY
Several different lipoproteins function in cholesterol metabolism. Chylomicrons transfer dietary
cholesterol from the intestine to the liver. VLDL and LDL transfer cholesterol from the liver to the
rest of the body. HDL transfer excess cholesterol from the tissues back to the liver for disposal.