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The oxidation and utilization of

amino acids

Dr. Solomon Genet


MUHAS

Topics to be covered
Removal of -amino groups from amino acids
(transamination and deamination)
Enzymes and coenzymes involved during
transamination
Entry of the carbon skeletons to the TCA cycle
Amino group transport
Removal of ammonia in humans (urea cycle)

Lecture objectives
To describe the oxidation of amino acids
To explain the different ways of removal of
amino groups from amino acids
To understand the reactions and enzymes
involved in the oxidation of amino acids
To discuss the way of ammonia removal
from the body
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Nitrogen balance
In normal adults, nitrogen intake matches
nitrogen excreted.
Positive nitrogen balance, an excess of
ingested over excreted nitrogen, accompanies
growth and pregnancy.
Negative nitrogen balance, where output
exceeds intake, may follow surgery, advanced
cancer, and kwashiorkor or marasmus.
While ammonia, derived mainly from the amino nitrogen of amino acids, is highly toxic,
tissues convert ammonia to the amide nitrogen
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of nontoxic glutamine.

Removal of -amino groups of


amino acids
Subsequent deamination of glutamine in
the liver releases ammonia, which is then
converted to nontoxic urea.
If liver function is compromised, as in
cirrhosis or hepatitis, elevated blood
ammonia levels generate clinical signs
and symptoms.

Metabolism of proteins
The continuous degradation and synthesis of
cellular proteins occur in all forms of life.
Each day humans turn over 12% of their total
body protein, principally muscle protein.
High rates of protein degradation occur in
tissues undergoing structural rearrangement
eg, uterine tissue during pregnancy, tadpole tail
tissue during metamorphosis, or skeletal
muscle in starvation.
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Removal of -amino groups of


amino acids
Since excess amino acids are not stored, those
not immediately incorporated into new protein
are rapidly degraded.
Different animals excrete excess nitrogen as
ammonia, uric acid, or urea.
The aqueous environment of teleostean fish,
which are ammonotelic (excrete ammonia).
Birds, which must conserve water and maintain
low weight, are uricotelic and excrete uric acid.
Land animals, including humans, are ureotelic
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and excrete nontoxic, water-soluble urea.

Removal of -amino groups of


amino acids
Main way of removal of -amino groups is by a
process called transamination.
Transamination Transfers -Amino Nitrogen to
-Ketoacids (pyruvate, oxaloacetate and ketoglutarate)
-ketoglutarate is the final collectror of -a,mino
groups of amino acids.
The only amino acids that can not be
transaminated are lysine, threonine, and proline
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Removal of -amino groups of amino


acids
Transamination is readily reversible, and it is
catalyzed by transaminases or aminotransferases.
Aminotransferases also function in amino acid
biosynthesis.
The coenzyme pyridoxal phosphate (PLP) is present
at the catalytic site of aminotransferases.
The coenzyme is derivative of vitamin B6.
Alanine-pyruvate aminotransferase (alanine
aminotransferase) and glutamate--ketoglutarate
aminotransferase (glutamate aminotransferase)
catalyze the transfer of amino groups.
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Removal of -amino groups of amino


acids
Each aminotransferase is specific for one pair of
substrates but nonspecific for the other pair.
Alanine is also a substrate for glutamate
aminotransferase, all the amino nitrogen from amino
acids that undergo transamination can be
concentrated in glutamate.
This is important because L-glutamate is the only
amino acid that undergoes oxidative deamination at
an appreciable rate in mammalian tissues.
Transamination is not restricted to -amino groups.
The -amino group of ornithine-but not the -amino
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group of lysine-readily undergoes transamination.

Glutamate Releases Its Amino Group


as Ammonia in the Liver
The amino groups from many of the amino acids are
collected in the liver in the form of the amino group of
L-glutamate molecules.
In hepatocytes, glutamate is transported from the
cytosol into mitochondria, where it undergoes
oxidative deamination catalyzed by L-glutamate
dehydrogenase
It is an allosteric enzyme with six identical subunits, its
activity is influenced by allosteric modulators.
The best-studied of these are the positive modulator
ADP and the negative modulator GTP.
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Glutamine Transports Ammonia in the


Bloodstream

Two mechanisms are available in humans for the


transport of ammonia from the peripheral tissues to
the liver to be excreted as urea.
1. Synthesis of glutamine from glutamate by glutamine
synthase and glutamate acts as nontoxic ammonia
carrier in blood.
2. In muscle pyruvate is transaminated to alanine,
which delivers ammonia to the liver where it gets
transaminated by ALT and reconverts to pyruvate,
this is again used for glucose synthesis in liver
(gluconeogenesis) and transported to muscle
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(glucose-alanine cycle).

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Muscle
protein

Amino acids
Glucose
glycolysis

Pyruvate

NH4+
Glutamate

Alanine

GLT

-Ketoglutarate

Blood glucose

Blood
Alanine

Glucose
Gluconeogenesis

Pyruvate

-Ketoglutarate
ALT

Glutamate
NH4+
Urea cycle

The glucose alanine cycle between liver and muscle Urea

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Removal of toxic ammonia


Glutamine collects toxic ammonia from
extrahepatic tissues to the liver for removal.
The glutamine is synthesized by glutamine
synthetase enzyme, which requires ATP for the
synthesis.
Glutamine is changed to glutamate by release
of the amide nitrogen-catalyzed by glutaminase
This ammonia is then converted to urea and
excreted.
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The Urea cycle


Ammonia is toxic to the body and shall be removed
Urea is the major disposal form of amino groups (in
humans) derived from amino acids, and accounts for
about 90% of the nitrogen-containing components of
urine.
One nitrogen of the urea molecule is supplied by free
NH4 and the other nitrogen by aspartate.
The cycle utilizes energy for the synthesis (3 ATPs per
urea molecule)
Occurs mainly in the liver and the kidney to a small
extent.
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The Urea cycle


Occurs partly in the cytosol and partly in the
mitochondria.
The rate limiting reaction is the first reaction,
which is the synthesis of carbamoyl phosphate.
The first two reactions of the cycle take place in
the mitochondria.
Urea cycle utilizes CO2 for urea synthesis,
hence it also helps remove CO2
Urea cycle is connected to TCA cycle at the
level of fumarate (found in both cycles as
intermediate)
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The urea cycle


The urea cycle consists of fiver enzyme
catalyzed reactions producing urea.
Urea produced by the liver is transported in the
blood to the kidneys for excretion in the urine.
The liver changes toxic ammonia to a nontoxic
and soluble form urea for excetion.
Carbamoyl phosphate synthetase enzyme is
the rate limiting enzyme of the cycle.

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The reactions of urea cycle


1. Formation of carbamoyl phosphate: HCO3- 2ATPs
and NH4+ are required by the mitochondrial enzyme
carbamoyl phosphate synthetase I, which
synthesizes carbamoyl phosphate in the
mitochondria of liver cells.
2. Synthesis of Citrulline: Carbamoyl phosphate
condenses with ornithine (basic amino acid not
found in proteins) to form citrulline (also is a
nonprotein amino acid) catalyzed by ornithine
transcarbamoylase. Citrulline comes out to the
cytosol to continue with next reactions.
3. Formation of arginosuccinate: citrulline condenses
with aspartate to form arginosuccinate, catalyzed by
arginosuccinate synthetase, which also requires
ATP. This reaction occurs in the cytosol.
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The reactions of urea cycle


4. Cleavage of argininosuccinate:
Argininosuccinate is cleaved to yield arginine
and fumarate (link with TCA cycle). This
reaction is catalyzed by arginosuccinate
lyase. This takes place in the cytosol.
5. Cleavage of arginine to ornithine and urea:
Arginase cleaves arginine to ornithine and
urea, and occurs almost exclusively in the
liver. Ornithine is thus recycles and another
round continues.
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Urea disposal
The urea produced is transported via blood to the
kidney to be filtered and excreted through urine.
A portion of the urea diffuses from the blood into the
intestine, and is cleaved to CO2 and NH3 by bacterial
urease.
In patients with kidney failure, plasma urea levels are
elevated.
The intestinal action of urease on this urea becomes a
clinically important source of ammonia, contributing to
the hyperammonemia often seen in these patients.
Oral administration of neomycin reduces the number
of intestinal bacteria responsible for excess ammonia.
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Regulation of the urea cycle


The flux of nitrogen through the urea cycle in an
individual animal varies with diet.
All five enzymes are synthesized at higher rates
in starving animals and in animals on very-highprotein diets than in well-fed animals eating
primarily carbohydrates and fats.
Animals on protein-free diets produce lower
levels of urea cycle enzymes.
Allosteric regulation of at least one key enzyme
adjusts the flux through the urea cycle. The first
enzyme in the pathway, carbamoyl phosphate
synthetase I, is allosterically activated by N27
acetylglutamate

Defects in urea cycle enzymes


People with genetic defects in any enzyme
involved in urea formation cannot tolerate
protein rich diets.
The absence of any one of urea cycle enzymes
can result in hyperammonemia.
In the rare cases of arginase deficiency, arginine,
the substrate of the defective enzyme, must be
excluded from the diet.
The levels of serum ammonia are normally low (5 to 50
mM).

However, when liver function is compromised, due


either to genetic defects of the urea cycle, or liver
disease, blood levels can rise above 1000mM. 28

Defects in urea cycle enzymes


Such hyperammonemia is a medical
emergency, because ammonia has a direct
neurotoxic effect on the CNS.
Elevated concentrations of ammonia in the
blood cause the symptoms of
ammoniaintoxication, which include tremors,
slurring of speech, somnolence, vomiting,
cerebral edema, and blurring of vision.
At high concentrations, ammonia can cause
coma and death.
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Essential and non essential amino acids


Amino acids that can not be synthesized are
essential and non essential otherwise.
Transamination helps produce amino acids like,
glutamate, alanine, and aspartate.
Humans can synthesize 11 out of 20 amino
acids.
Arginine is synthesized but not in enough
amounts, especially for growing children who
constantly grow until adult hood.
But most amino acids are also obtained from
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diet.

Essential and non essential amino


acids
Essential
Arginine*
Histidine
Isoleucine
Leucine
Lysine
Methionone$
Phenylalanine#
Threonine
Tryptophane
Valine

Nonessential
Alanine
Aspartate
Asparagine
Cysteine
Glutamate
Glutamine
Glycine
Proline
Serine
Tyrosine

*Produced in animals but not in enough amount


As reqired hence still essential
# required in large quantity for tyrosine synthesis, if the latter is not availed in diet.
$ required for in bulk for cysteine synthesis
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Entry of carbon skeletons of amino acids to


TCA cycle
Amino acids account 15-20% of the human body
energy production.
Not active as glycolysis or fatty acid oxidation.
Carbons of amino acids enter intermediary
metabolism at one of seven points.
Glucogenic amino acids are metabolized to
pyruvate, 3-phosphoglycerate, -ketoglutarate,
oxaloacetate, fumarate, or succinylCoA.
Ketogenic amino acids produce acetylCoA or
acetoacetate.
Some amino acids produce both intermediates
and hence are both ketogenic and glucogenic. 32

Entry of carbon skeletons of amino


acids to TCA cycle
The carbon skeletons of amino acids can be used for the
generation of energy.
Seven amino acids are degraded to acetylCoA or
acetoacetylCoA (phenylalanine, tyrosine, isoleucine, leucine,
tryptophane, threonine and lysine).
The other amino acids producing pyruvate, -ketoglutarate,
succinylCoA and fumarate and are glucogenic (arginine,
glutamine, histidine, proline, methionine, valine, alanine,
cysteine, glycine, serine, asparagine and aspartate)
Isoleucine, threonine, tryptophanetyrosine and phenyl alanine
also produce pyruvate and TCA cycle intermediates and hence
are both ketogenic and glucogenic.
Lysine and Leucine are purely ketogenic.
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