Urea cycle

Amino acid oxidation and

the production of urea

Oxidation

Waste or reuse
Ammonia has to be
eliminated
Ammonia originates in the catabolism of amino
acids that are primarily produced by the
degradation of proteins – dietary as well as
existing within the cell:
digestive enzymes
proteins released by digestion of cells
muscle proteins
hemoglobin
intracellular proteins (damaged, unnecessary)
Ammonia has to be
eliminated
Ammonia (NH3) is a relatively strong base, and at physiological
pH values it is mainly present in the form of the ammonium
ion NH4 + .
NH3 and NH4+ are toxic, and at higher concentrations cause
brain damage in particular.
Because it reacts with -ketoglutarate, thus making it limiting
for the TCA cycle  decrease in the ATP level

Liver damage or metabolic disorders associated with elevated

ammonia can lead to tremor, slurred speech, blurred vision,
coma, and death

Normal conc. of ammonia in blood: 30-60 µM

 2 CHOICES
1. Reuse

2. Urea cycle

Fumarate

Oxaloacetate

Overview of amino acid catabolism in mammals

Nitrogen removal from amino
acids
Transamination Aminotransferase
PLP

Oxidative
deamination

Urea cycle
Nitrogen removal from
amino acids
Step 1: Remove amino group
Step 2: Take amino group to liver for nitrogen excretion
Step 3: Entry into mitochondria
Step 4: Prepare nitrogen to enter urea cycle
Step 5: Urea cycle
 Excretory forms of nitrogen

a) Excess NH4+ is excreted as ammonia (microbes, aquatic

vertebrates or larvae of amphibia),
b) Urea (many terrestrial vertebrates)
c) or uric acid (birds and terrestrial reptiles)
Step 1. Remove amino group
Transfer of the amino group of an amino acid to an -keto
acid  the original Aamino acid is converted to the
corresponding -keto acid
Transamination is catalyzed by transaminases
(aminotransferases) that require participation of
pyridoxalphosphate:

amino acid

pyridoxalphosphate Schiff base

 Step 2: Take amino group to liver for
nitrogen excretion
Glutamate releases its amino group
as ammonia in the liver.

The amino groups from many of

Glutamate the amino acids are collected in the
dehydrogenase
liver in the form of the amino
group of L-glutamate molecules.

The glutamate dehydrogenase of mammalian liver has the

unusual capacity to use either NAD+ or NADP+ as cofactor
 Nitrogen carriers
1. Glutamate
Transfers one amino group WITHIN cells:
Aminotransferase → makes glutamate from a-ketoglutarate
Glutamate dehydrogenase → opposite
2. Glutamine
Transfers two amino group BETWEEN cells → releases its
amino group in the liver
3. Alanine
Transfers amino group from tissue (muscle) into the liver
 Move within cells

Move between cells

In liver
 Glucose-alanine cycle

Alanine plays a special role in transporting

amino groups to liver.

Ala is the carrier of ammonia and of the carbon

skeleton of pyruvate from muscle to liver.
The ammonia is excreted and the pyruvate is used
to produce glucose, which is returned to the
muscle.
 Sources of ammonia for the urea cycle:
Oxidative deamination of Glu, accumulated in the liver by the action of
transaminases and glutaminase

Glutaminase reaction releases NH3 that enters the urea cycle in the liver (in the
kidney, it is excreted into the urine)

Catabolism of Serine, Threonine, and Histidine (nonoxidative deamination) also

releases ammonia:

Serine →→ pyruvate + NH4+

Threonine →→ a-ketobutyrate + NH4+

Bacteria in the gut also produce ammonia.

Step 3: entry of nitrogen to mitochondria
Step 4: prepare nitrogen to enter urea cycle

Regulation
Step 5: Urea cycle aspartate

Ornithine
transcarbamoylase

Argininosuccinate
synthase

Arginase 1

Argininosuccinate
lyase
UREA CYCLE
O

H2N C NH2
urea

Most terrestrial animals convert excess nitrogen to urea, prior to excreting

it.
Urea is less toxic than ammonia.
The Urea Cycle occurs mainly in liver. First 2 reactions in mitochondria
and rest in cytosol.
The 2 nitrogen atoms of urea enter the Urea Cycle as NH3 (produced
mainly via Glutamate Dehydrogenase) and as the amino N of aspartate.
The NH3 and HCO3- (carbonyl C) that will be part of urea are incorporated
first into carbamoyl phosphate.
 O O C NH2
NH3+
Urea Cycle H2N C OPO32 NH
CH2 carbamoyl
Enzymes in phosphate
CH2
mitochondria: CH2
CH2
citrulline
Pi
1.Carbamoyl CH2 2
CH2
phosphate synthase I
HC NH3+ NH3+ COO
2.OrnithineTrans- Urea Cycle HC
carbamylase COO COO CH2
Enzymes in cytosol: ornithine ATP 3 HC NH2
3. Arginino- O 5 H2O
AMP + PPi
COO
Succinate COO
H2N C NH2 aspartate
Synthase H2N NH2+ CH2
urea
4. Arginino C H
4 HC N C NH2+
succinase
5. Arginase.
NH COO NH
CH2
COO CH2 arginino-
arginine CH2 succinate
HC CH2
CH2 CH2
CH
HC NH3+
COO HC NH3+
COO fumarate COO
Step -1 HCO3
ATP
Carbamoyl Phosphate
ADP
Synthase (Type I) catalyzes a
O
3-step reaction, with carbonyl
phosphate and carbamate HO C OPO32
intermediates. NH3 carbonyl phosphate
Ammonia is the N input. Pi
O
The reaction, which involves
cleavage of 2 ~P bonds of H2N C O
ATP, is essentially ATP carbamate
irreversible. ADP
O

H2N C OPO32
carbamoyl phosphate
 HCO3 + NH3 + 2 ATP
Carbamoyl Phosphate
Synthase
O

H2N C OPO32 + 2 ADP + Pi

carbamoyl phosphate

Carbamoyl Phosphate Synthase I is the committed step of the Urea Cycle, and is
subjected to regulation.
Carbamoyl Phosphate Synthase II: present in cytosol of liver cells and involved in
synthesis of pyrimidines.
 glutamate (Glu) N-acetylglutamate
H O H

H3N+ C COO H 3C C N C COO

H
CH2 CH2

CH2 CH2

COO COO

Carbamoyl Phosphate Synthase has an absolute requirement for an allosteric

activator N-acetylglutamate.
This derivative of glutamate is synthesized from acetyl-CoA & glutamate when
cellular [glutamate] is high, signaling an excess of free amino acids due to protein
breakdown or dietary intake.
Step:2 (synthesis of Citrulline)
Ornithine transcarbamoylase – mitochondrial enzyme
Catalyses addition of ornithine to the carbonyl group of
Carbamoyl phosphate.
Step:3 (synthesis of Argininosuccinate)
Argininosuccinate synthase – cytosolic enzyme
Requires ATP
Step:4 (Cleavage of Argininosuccinate)
Argininosuccinase /Argininosuccinate lyase – cytosolic eznyme
Argininosuccinate-- Arginine + Fumarate
Regulation of urea cycle:

1.The reaction catalyzed by carbamoyl phosphate synthase I is the rate

limiting reaction and committed step of Urea cycle
It is allosterically activated by N-acetylglutamate(NAG).
2. Consumption of protein rich meal increases the levels of NAG in liver,
leading to enhanced urea synthesis.
3. Carbamoyl phosphate synthase I and glutamate dehydrogenase are located
in mitochondria. They coordinate each other in the formation of NH3,
and its utilization for the synthesis of carbamoyl phosphate.
The remaining enzymes of urea cycle are mostly controlled by the
concentrations of their respective substrates.
 UREA CYCLE DISORDERS
S.No Disorder Enzyme involved

1. Hyperammonemia type I Carbamoyl phosphate synthase I

2. Hyperammonemia type II Ornithine transcarbamoylase

3. Citrllinemia Arginosuccinate synthase

4. Arginosuccinicaciduria Arginosuccinase

5. Hyperargininemia Arginase
Hyperammonemia
levels of serum ammonia are normally low (5–35 μmol/L).
However, when liver function is compromised, due either to genetic
defects of the urea cycle or liver disease, blood levels can rise above
1,000 μmol/L.
Such hyperammonemia is a medical emergency, because ammonia
has a direct neurotoxic effect on the CNS.
For example, elevated concentrations of ammonia in the blood cause
the symptoms of ammonia intoxication, which include tremors,
slurring of speech, somnolence, vomiting, cerebral edema, and
blurring of vision. At high concentrations, ammonia can cause coma
and death.
Acquired hyperammonemia
Liver disease is a common cause of hyperammonemia in adults, and
may be due, for example, to viral hepatitis or to hepatotoxins such as
alcohol.
The conversion of ammonia to urea is, therefore, severely impaired,
leading to elevated levels of ammonia
Congenital hyperammonemia:
Genetic deficiencies of each of the five enzymes of the urea cycle
have been described, with an overall prevalence estimated to be
1:25,000 live births.
Ornithine transcarbamoylase deficiency, which is X-linked, is the
most common of these disorders, predominantly affecting males,
although female carriers may become symptomatic.
All of the other urea cycle disorders follow an autosomal recessive
inheritance pattern.
In each case, the failure to synthesize urea leads to
hyperammonemia during the first weeks following birth.
Treatment
Administration of compounds that bind covalently to amino acids,
producing nitrogen-containing molecules that are excreted in the urine,
has improved survival.
For example, phenylbutyrate given orally is converted to
phenylacetate. This condenses with glutamine to form phenyl -
acetylglutamine, which is excreted