1

Medicine
Dentistry

February 15

Medicine

February 15

Pyruvate Dehydrogenase

TPP

Lipoic

Pyruvate
Dihydrolipoyl
dehydrogenase dehdrgenase

FAD
Dihydrolipoyl
transacetylase

irreversible;
in mitochodria.

Medicine

February 15

Pyruvate dehydrogenase complex:

E1 pyruvate dehydrogenase
Es

E2 dihydrolipoyl transacetylase

E3 dihydrolipoyl dehydrogenase
mnemonics

Tender
Loving Cofactors
Care
(Vitamins)
For
Nancy

Thiamine pyrophosphate, TPP (VB1)

Lipoic Acid
CoA (pantothenic acid)
FAD (VB2)

NAD+ (VB3)

Medicine

Pyruvate Dehydrogenase
Subunits
Enzyme

Abbreviated

Prosthetic Group

Pyruvate
Dehydrogenase

E1

Thiamine
pyrophosphate (TPP)

Dihydrolipoyl
Transacetylase

E2

Lipoamide

Dihydrolipoyl
Dehydrogenase

E3

FAD

Medicine

Medicine

February 15

Lipoic acid (lipoate) in amide linkage with

a Lys residue

The lipoyllysyl moiety is the prosthetic group of dihydrolipoyl transacetylase (E2 of the
PDH complex). The lipoyl group occurs in oxidized (disulfide) and reduced (dithiol)
forms and acts as a carrier of both hydrogen and an acetyl (or other acyl) group.
8

Medicine

February 15

Localization of pyruvate decarboxylation

The transport of pyruvate into the mitochondria is via a transport
protein and is active, consuming energy. Passive diffusion of
pyruvate into the mitochondria is impossible because it carries a
negative charge.
On entry to the mitochondria the pyruvate decarboxylation occurs,
producing acetyl CoA. This irreversible reaction traps the acetyl CoA
within the mitochondria (the acetyl-CoA can only be transported out
of the mitochondrial matrix under conditions of high oxaloacetate via
the citrate shuttle, a TCA intermediate). The carbon dioxide
produced by this reaction is nonpolar and small, and can diffuse out
of the mitochondria and out of the cell.

Medicine

Citrate Shuttle
Acetyl-CoA can only be transported out of the mitochondrial matrix under conditions of high
oxaloacetate via the citrate shuttle

Medicine

Oxidative decarboxylation of pyruvate to

acetyl-CoA by the PDH complex

The first step is the slowest and therefore limits the rate of the overall reaction.
It is also the point at which the PDH complex exercises its substrate specificity
13

Medicine

February 15

Medicine

Oxidative decarboxylation of
pyruvate.

Regulation of Pyruvate
Dehydrogenase Complex:

NADH competes with NAD+ for

binding to E3.
Acetyl CoA competes with CoA
for binding to E2.

Pyruvate (-)

Pyruvate dehydrogenase is inhibited when one or more of the three

following ratios are increased: ATP/ADP, NADH/NAD+ and acetylCoA/CoA.
In eukaryotes PDC is tightly regulated by its own specific pyruvate
dehydrogenase kinase (PDK) and pyruvate dehydrogenase
phosphatase (PDP), deactivating and activating it respectively.
PDK phosphorylates three specific serine residues on E1 with different
affinities. Phosphorylation of any one of them renders E1 (and in
consequence the entire complex) inactive.
Dephosphorylation of E1 by PDP reinstates complex activity.
Products of the reaction act as allosteric inhibitors of the PDC, because
they activate PDK. Substrates in turn inhibit PDK, and thus, reactivating
PDC.
During starvation, PDK increases in amount in most tissues, including
skeletal muscle, via increased gene transcription. Under the same
conditions, the amount of PDP decreases. The resulting inhibition of
PDC prevents muscle and other tissues from catabolizing glucose and
gluconeogenesis precursors. Metabolism shifts toward fat utilization,
while muscle protein breakdown to supply gluconeogenesis precursors
is minimized, and available glucose is spared for use by the brain.
Calcium ion has a role in regulation of PDC in muscle tissue, because it
activates PDP, stimulating glycolysis on its release into the cytosol during muscle contraction.
Medicine

(1) Pyruvate dehydrogenase

complex
allosteric activators:
AMP, CoA,
NAD+,Ca2+

allosteric inhibitors:
ATP, acetyl CoA,
NADH, FA
Pyruvate dehydrogenase
(active form)
Pi

ATP
pyruvate dehydrogenase
kinase

pyruvate dehydrogenase
phosphatase
H2O

2+

Ca ,insulin

ADP

pyruvate dehydrogenase P
(inactive form)
Medicine

acetyl CoA,
NADH

ADP,
NAD+

During starvation:
Pyruvate Dehydrogenase Kinase increases in amount
in most tissues, including skeletal muscle, via increased
gene transcription.
Under the same conditions, the amount of Pyruvate
Dehydrogenase Phosphatase decreases.
The resulting inhibition of Pyruvate Dehydrogenase
prevents muscle and other tissues from catabolizing glucose
& gluconeogenesis precursors.
Metabolism shifts toward fat utilization.
Muscle protein breakdown to supply gluconeogenesis
precursors is increased.
Available glucose is spared for use by the brain.
Medicine

Advantages of multienzyme complex:

The PDH complex is a classic multienzyme complex in which a

series of chemical intermediates remain bound to the enzyme
molecules as a substrate is transformed into the final product.
Five cofactors, four derived from vitamins, participate in the
reaction mechanism. The regulation of this enzyme complex also
illustrates how a combination of covalent modification and
allosteric regulation results in precisely regulated flux through a
metabolic step

Higher rate of reaction: Because product of one enzyme acts as a

substrate of other, and is available for the active site of next
enzyme without much diffusion.

Minimum side reaction.

Coordinated control.

Medicine

Treatment
Use of a ketogenic diet has been described.
The ketogenic diet is a high-fat, adequate-protein, low-carbohydrate
diet. The diet mimics aspects of starvation by forcing the body to
burn fats rather than carbohydrates.
Current research is being conducted on the viability
of Dichloroacetic acid to treat the lactic acidosis commonly
accompanied by this disorder. Salts of DCA have been studied as
potential drugs because they stimulates the activity of
the enzyme pyruvate dehydrogenase by inhibiting the enzyme
pyruvate dehydrogenase kinase. Thus, it
decreases lactate production by shifting the metabolism of pyruvate
from glycolysis towards oxidation in the mitochondria.

Additionally, there is research being conducted on the viability of

gene therapy for sufferers of this condition as well as many other
mitochondrial defects.

Medicine

Thiamine is involved in a vast array of functions:

Carbohydrate metabolism
Production of the neurotransmitters glutamic
acid and GABA, through the citric acid cycle
Lipid metabolism, necessary for myelin production
Amino acid metabolism
Neuromodulation - physiological process by which a
given neuron uses one or more neurotransmitters to
regulate diverse populations of neurons.

Medicine

Thiamin (Vitamin B1) deficiency causes Beriberi

Thiamine pyrophosphate (TPP) is an important cofactor of pyruvate dehydrogenase complex, or
PDC a critical enzyme in glucose metabolism. Thiamine is neither synthesized nor stored in
good amounts by most vertebrates. It is required in the diets of most vertebrates.
The body cannot produce thiamine and can only store up to 30 mg of it in tissues. Thiamine is
mostly concentrated in the skeletal muscles. Other organs in which it is found are the brain,
heart, liver, and kidneys. The half-life of thiamine is 9-18 days. It is excreted by the kidney
Thiamine deficiency ultimately causes a fatal disease called Beriberi characterized by
neurological disturbances, paralysis, atrophy of limbs and cardiac failure. Note that brain
exclusively uses aerobic glucose catabolism for energy and PDC is very critical for aerobic
catabolism. Therefore thiamine deficiency causes severe neurological symptoms.
The main types of beriberi are:
Dry beriberi and Wernicke-Korsakoff syndrome affect the peripheral and central nervous system
respectively.
Wet (edematous) beriberi affects the cardiovascular system, as well as other bodily systems.
Infantile beriberi affects mostly children in developing countries
Prevalence
Beriberi is rare in developed countries because most foods are now vitamin-enriched. Excluding
the presence of arsenic in the environment (e.g. well water) one can get enough thiamine by
eating a normal, healthy diet. Today, beriberi occurs mostly in patients who abuse alcohol.
Drinking heavily can lead to poor nutrition, and excess alcohol makes it harder for the body to
absorb and store thiamine.
Medicine

Wernicke-Korsakoff Syndrome (WKS)

Insufficient thiamine significantly impairs glucose oxidation, causing highly aerobic tissues,
such as brain and cardiac muscle, to fall first. In addition, branched-chain amino acids are
sources of energy in brain and muscle
Wernicke-Korsakoff syndrome is a brain disorder due to thiamine deficiency.
Wernickes Encephalopathy (peripheral neuropathy ) and Korsakoffs Psychosis are the acute
and chronic phases, respectively, of the same disease.
Korsakoff syndrome, or Korsakoff psychosis, tends to develop as Wernicke's symptoms go
away. Wernicke's encephalopathy causes brain damage in lower parts of the brain called the
thalamus and hypothalamus. Korsakoff psychosis results from damage to areas of the brain
involved with memory.
The disease is typically associated with chronic alcoholism, but may be associated with
malnutrition or other conditions which cause nutritional deficiencies. Alcohol interferes with
thiamine absorption from the intestine,

Congestive heart failure may be a complication (wet beri-beri) owing to inadequate ATP and
accumulation of ketoacids in the cardiac muscles. (Peripheral vasodilation leading to a
high cardiac output state. This leads to salt and water retention and Edema.)
Two other enzyme complexes similar to pyruvate dehydrogenase that use thiamine are:
-Ketoglutarate dehydrogenase (citric acid cycle)
Branched-chain ketoacid dehydrogenase (metabolism of branched-chain amino acids)
.

Medicine

Arsenic compounds in low doses are very toxic to microorganisms, therefore

these compounds were used for the treatment of syphilis and other diseases in
earlier days. Arsenicals were first antibiotics, but with a terrible side effects as
they are eventually very toxic to humans.
Unfortunately and ignorantly, a common nineteenth century tonic, the Fowlers
solution contained 10 mg/ml arsenite. This tonic must have been responsible for
many deaths, including the death of the famous evolution scientist Charlse
Darwin.

Arsenic Poisoning: Arsenic compounds such as arsenite (AsO3---) organic

arsenicals are poisonous because they covalently bind to sulfhydryl compounds
(SH- groups of proteins and cofactors). Dihydrolipoamide is a critical cofactor of
PDC, and it has two-SH groups, which are important for the PDC reaction. These
SH groups are covalently inactivated by arsenic compounds as shown below;
OH
-O

As

HS

S
-O

+
OH

HS

As

+ 2H2O
S

R
Medicine

Glycolysis in disease
Genetic diseases
Glycolytic mutations are generally rare due to
importance of the metabolic pathway, this
means that the majority of occurring mutations
result in an inability for the cell to respire, and
therefore cause the death of the cell at an early
stage. However, some mutations are seen with
one notable example being Pyruvate kinase
deficiency, leading to chronic hemolytic anemia.
Medical Students

Pyruvate Kinase Deficiency

Genetic defects of this enzyme cause the disease known as pyruvate kinase
deficiency.
In this condition, a lack of pyruvate kinase slows down the process of glycolysis.

This effect is especially devastating in cells that lack mitochondria, because these cells
must use anaerobic glycolysis as their sole source of energy because the TCA cycle is
not available.
One example is red blood cells, which in a state of pyruvate kinase deficiency rapidly
become deficient in ATP and can undergo hemolysis. Therefore, pyruvate kinase
deficiency can cause hemolytic anemia and an increase in plasma bilirubin.
A discrepancy between red blood cell energy requirements and ATP generating
capacity produces irreversible membrane injury resulting in cellular distortion, rigidity,
and dehydration. This leads to premature erythrocyte destruction by the spleen and
liver.

Medical Students

Pyruvate Kinase Deficiency

The buildup of reaction intermediates can also increase the level
of 2,3-bisphosphoglycerate in the cells and affect tissue oxygenation.
This will cause a "right shift" in the hemoglobin oxygen saturation
curve, implying a decreased oxygen affinity for the hemoglobin and
earlier oxygen unloading than under normal conditions.
Low prevalence, No Heinz bodies (inclusions or aggregates
within red blood cells composed of denatured hemoglobin )
Second cause of hemolytic anemia after G6Pdase deficiency.
Treatment can include a blood transfusion or removal of the spleen.
Treatment is usually effective in reducing the severity of the
symptoms.

Medical Students

In RBC

2-3-BPG BIND TO Hb, DECREASE ITS AFFINITY TO O2, INCREASE O2

AVALABILITY TO TISSUE.
Medical Students

No BPG

BPG Lowers the binding affinity of

Hb for O2
[BPG] = 0, Hb P50 = 1 torr
[BPG] = 4000mM, Hb P50 = 26
torr
Without BPG, Hb couldnt
unload O2 in cells

With BPG

Hb
SATURATION

0
10

50

o2O2 PRESSURE (torr)

Medical Students

Glycolysis as an indicator of
disease
The glycolytic rates in malignant, rapidly-growing tumor cells are up
to 200 times higher than those of their normal tissues of origin,
despite the ample availability ofoxygen. A classical explanation
holds that the local depletion of oxygen within the tumor is the cause
of increased glycolysis in these cells. However, there is also strong
experimental evidence that attributes these high rates to an overexpressed form of the enzyme hexokinase (Bustamante and
Pedersen 2005),which is responsible for driving the high glycolytic
activity when oxygen is not necessarily depleted. This finding
currently has an important medical application: aerobic glycolysis by
malignant tumors is utilized clinically to diagnose and monitor
treatment responses of cancers using medical imaging techniques
(Pauwels et al. 2000, PETNET Solutions 2006).

Medical Students

Ethanol Metabolism and Gluconeogenesis

Ethanol strongly inhibits gluconeogenesis and can bring about
hypoglycemia, a potentially dangerous decrease in blood glucose
levels. Ethanol metabolism occurs primarily in the liver. The
reaction, catalyzed by alcohol dehydrogenase is shown as
follows:
Ethanol + NAD+ <=> Acetaldehyde + NADH + H+
The NADH produced in this reaction shifts the equilibrium in the liver
cytosol of the lactate dehydrogenase from pyruvate formation to
lactate synthesis.
The NADH also favors reduction of oxaloacetate to malate in the
reaction catalyzed by malate dehydrogenase, making less
oxalacetate available forgluconeogenesis. The resulting
hypoglycemia can affect the part of the brain concerned with
temperature regulation and the body temperature can fall by as
much as 2 C. Thus, feeding alcohol to people suffering from
hypothermia is counterproductive. Metabolically
speaking, glucose would be far more effective in raising body
temperature.
Medical Students

FRUCTOSE METABOLISM

Hereditary fructose1,6-bisphosphatase
deficiency results in
severely impaired
hepatic
gluconeogenesis and
leads to episodes of
hypoglycemia, apnea,
hyperventillation,
ketosis and lactic
acidosis.

Sequestering ATP

Medical Students

GALACTOSE METABOLISM
Sequestering ATP

SORBITOL/
Cataracts early in life

Medical Students

Gal-1-P-uridyltransferase deficiency:
Cataracts early in life.
Vomiting, diarrhea following lactose ingestion.
Lethargy.
Liver damage, hyperbilirubinemia.
Mental retardation.

Figure 12.4. Sorbitol metabolism

Medical Students

Conversion of glucose to fructose via sorbitol

Synthesis of sorbitol:
~Aldose reductase reduces glucose, producing sorbitol (glucitol).
~This enzyme is found, in lens, retina, Schwann cells, liver, kidney,
placenta, red blood cells, and in cells of the ovaries and seminal
vesicles
~In cells of the liver, ovaries, sperm, and seminal vesicles, there is a
second enzyme, sorbitol dehydrogenase, that can oxidize the
sorbitol to produce fructose sperm cells use fructose .

~pathway from sorbitol to fructose in the liver provides a mechanism

by which any available sorbitol is converted into a substrate that can
enter glycolysis or gluconeogenesis.

Medical Students

The effect of hyperglycemia on sorbitol metabolism:

Because insulin is not required for the entry of glucose into the
cells where sorbitol syn. occurs. large amounts of glucose may
enter during hyperglycemia (uncontrolled diabetes).
Elevated intracellular glucose produce a significant increase in the
amount of sorbitol, which cannot pass efficiently through cell
membranes and, therefore, remains trapped inside the cell.
This is exacerbated when sorbitol dehydrogenase is low or
absent(in retina, lens, kidney, and nerve cells).
result, sorbitol accumulates in these cells(strong osmotic effects
(cataract formation, peripheral neuropathy, and vascular problems
leading to nephropathy and retinopathy)
Medical Students

2. The effect of hyperglycemia on sorbitol metabolism

- Because insulin is not required for entry of gluc into cells
listed in previous paragraph, large amounts of gluc may enter
these cells during times of hyperglycemia, e.g., in
uncontrolled diabetes.
- Elevated intracellular gluc concs & an adequate supply of
NADPH cause aldose reductase to produce a sufficient
increase in the amount of sorbitol, which cant pass efficiently
through CMs &, therefore, remains trapped inside cell.
- This is exacerbated when sorbitol dehydrogenase is low or
absent, e.g., in retina, lens, kidney & nerve cells. As a result,
sorbitol accumulates in these cells, causing strong osmotic
effects &, therefore, cell swelling as a result of water retention
- Some of the pathologic alterations associated with diabetes
can be attributed, in part, to this phenomenon, including
cataract formation, peripheral neuropathy, & vascular
problems leading to nephropathy, & retinopathy
Medical Students