1. BY
Dr A.VEERENDRA KUMAR REDDY MBBS, MD

2. Amino acid Catabolism
There are 20 different amino acid, they are monomeric
constituents of proteins.
In mammalian tissues, α-NH2 group of amino acids,
derived from the diet or breakdown tissue proteins,
ultimately is converted first to NH3 and then urea
and is excreted in the urine.
Urea is the end product of the amino acid catabolism.

Urea Cycle: Conversion of NH3 to urea for
excretion

3. Urea biosynthesis occurs in four stages:
(1) Transamination
(2) Oxidative deamination of glutamate
(3) Ammonia transport
(4) reactions of the urea cycle.
Also called as Krebs-Henseleit cycle or Ornithine
cycle
First metabolic pathway to be elucidated(1932).

5. 
Transamination: Transfer of amino group to αketoglutarate. There are several aminotransferases
specific to different amino acids. In this step amino
group from all the amino acids are transferred to aketoglutarate and they exist as glutamate.

Transaminases or aminotransferases require pyridoxal5’-phophate PLP (vitamin B6 derivative)

PLP is very important cofactor for many enzymatic
reactions.

7. 2.Oxidative deamination:
The amino group of glutamate is released as ammonia, regenerating α ketoglutarate, by an enzyme glutamate dehydrogenase.
Glutamate dehydrogenase requires NAD+ or NADP+ as cofactor. This is the
only enzyme known that has specificity for both type of cofactors.
This enzyme is allosterically inhibited by GTP and activated by ADP.

9. Sources of Ammonia:
1. Ammonia is produced in the body from the different tissues
by Amino acid catabolism.
2. Purine and Pyramidine catabolism.
3. The other source is from the dietary proteins and from
urea present in fluids secreted into the GI tract.
Glutamine Synthetase
GLUTAMINE is the important plasma transport form of
nitrogen from muscle.

10. Transport of excess
ammonia by glutamine:
Excess ammonia is toxic to animal
tissues.
Glutamine synthetase catalyses the
synthesis of glutamine by adding the
ammonia to glutamate at the expense
of ATP hydrolysis.
Glutamine is a non-toxic carrier of
ammonia. It is transported to liver or
kidney via blood.
In the liver & kidney, glutamine is
reconverted to glutamate and
ammonia by glutaminase. Ammonia is
incorporated in urea cycle to form
urea and then it will be excreted
through kidneys.

11. Normal blood ammonia levels:15-45 micro gm/dL.
http://www.nlm.nih.gov/medlineplus/ency/article/003506.htm
Hyper Ammonaemia:
1. Acquired hyperammonaemia: result of Cirrhosis
of Liver with development of collaterals
circulation.
2. Inherited hyperammonaemia: results from genetic
defects of the urea cycle enzymes.

12. Postulated mechanisms for toxicity of high ammonia:
1. High [NH3] would drive Glutamine Synthetase:
glutamate + ATP + NH3  glutamine + ADP + Pi
This would deplete glutamate – a neurotransmitter &
precursor for synthesis of the neurotransmitter GABA.
2. Depletion of glutamate & high ammonia level would drive
Glutamate Dehydrogenase reaction to reverse:
glutamate + NAD(P)+  a-ketoglutarate +
NAD(P)H + NH4+
The resulting depletion of a-ketoglutarate, an essential
Krebs Cycle intermediate, could impair energy metabolism
in the brain.

13. Features of Ammonia Toxicity:
A peculiar flapping tremor
Slurring of speech
Blurring of vision
Coma and death.
Metabolic fate of NH3 in the body
1. Mainly NH3 is converted to Urea(through Urea
cycle).
2. Formation of Glutamine
3. Amination of α-ketoacid to form α-amino acid.

14. O
H 2N
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.

16. O
NH3+
Urea Cycle
CH2
HC
OPO32-
ornithine
5
O
C
urea
CH2
H2O
NH2
COO-
COO-
4
COOHC
CH2
HC
HC
H
N
COO-
CH2
CH
NH3+
COO-
HC
NH2
COO-
aspartate
CH2
NH
arginine
CH2
3
ATP
AMP + PPi
C
CH2
COO-
NH3+
HC
NH2+
H2N
citrulline
CH2
Urea Cycle
COO-
H2N
CH2
2
NH3+
NH2
NH
Pi
CH2
1.Carbamoyl
phosphate
synthase I
2.OrnithineTranscarbamylase
Succinate
Synthase
4. Arginino
succinase
5. Arginase.
C
carbamoyl
phosphate
CH2
Enzymes in
mitochondria:
Enzymes in
cytosol:
3. Arginino-
H2N
C
O
COO-
fumarate
C
NH2+
NH
CH2
CH2
argininosuccinate
CH2
HC
NH3+
COO-

17. HCO 3-
Step -1
Carbamoyl Phosphate
Synthase (Type I) catalyzes a
3-step reaction, with carbonyl
phosphate and carbamate
intermediates.
Ammonia is the N input.
The reaction, which involves
cleavage of 2 ~P bonds of
ATP, is essentially
irreversible.
ATP
ADP
O
HO
OPO32-
C
carbonyl phosphate
NH3
Pi
O
H2N
C
O-
carbamate
ATP
ADP
O
H2N
C
OPO32-
carbamoyl phosphate

18. HCO3- + NH3 + 2 ATP
O
H2N
C
Carbamoyl Phosphate
Synthase
OPO 32- + 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.

19. glutamate (Glu)
N-acetylglutamate
H
H3N+
C
O
COO-
H3C
C
H
N
H
C
COO-
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.

20. Step:2 (synthesis of Citrulline)
Ornithine transcarbamoylase – mitochondrial
Catalyses addition of ornithine to the carbonyl group
of Carbamoyl phosphate.
Step:3 (synthesis of Argininosuccinate)
Argininosuccinate synthase – cytosolic
ATP -- AMP
Step:4 (Cleavage of Argininosuccinate)
Argininosuccinase /Argininosuccinate lyase cytosolic
Argininosuccinate-- Arginine + Fumarate

21. Step:5 ( Cleavage of Arginine to Ornithine and Urea)
Enzyme: Arginase – cytosolic
ENERGETICS
2 ATPs are utilized for the synthesis of carbamoyl
phosphate.
1 ATP is converted to AMP and Ppi to produce
Arginosuccinate which is equals to 2 ATP
Net utilization = 4 ATP

NH4+ + CO2 + aspartate + 3 ATP --->
urea + fumarate + 2 ADP + AMP + 4 Pi

22. 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.

25. 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

26. Hereditary deficiency of any of the Urea Cycle enzymes
leads to hyperammonemia - elevated [ammonia] in
blood.
Elevated ammonia is toxic, especially to the brain.
Other metabolites of urea cycle accumulate depending
on specific enzyme defect.
The clinical symptoms: Vomiting, lethargy, irritability,
ataxia, and metal retardation.
If not treated immediately after birth, severe mental
retardation results.
Total lack of any Urea Cycle enzyme is lethal.

27. Hyperammonemia type I:
Familial disorder, produces hyperammonemia and
symptoms of ammonia toxicity.
Hyperammonemia type II:
X linked, produces symptoms of ammonia toxicity.
Increased levels of glutamine (glutamine synthesis
enhanced in response to elevated NH3)

28. Postulated mechanisms for toxicity of high [ammonia]:
1. High [NH3] would drive Glutamine Synthase:
glutamate + ATP + NH3  glutamine + ADP + Pi
This would deplete glutamate – a neurotransmitter &
precursor for synthesis of the neurotransmitter GABA.
2. Depletion of glutamate & high ammonia level would drive
Glutamate Dehydrogenase reaction to reverse:
glutamate + NAD(P)+  a-ketoglutarate +
NAD(P)H + NH4+
The resulting depletion of a-ketoglutarate, an essential
Krebs Cycle intermediate, could impair energy metabolism
in the brain.

29. Possible therapies for the patients with defect in
urea cycle:
1. Defined diet containing just the minimum amount of essential
amino acids.
2. Feeding the patients with Benzoate or phenylacectate: These
compound react with glycine and glutamine respectively forming
non-toxic compounds that are excreted in urine. Thus the body
runs low in glycine and glutamine and starts synthesizing these
AA using the ammonia available in system. Thus clearing the
system of excess ammonia.
3. In the patients with N-acetylglutamate synthase deficiency, Carbamoyl
glutamate can act as activator of carbamoyl phosphate synthase.
4.Liver transplantation has also been used, since liver is the organ
that carries out Urea Cycle.

30. THANK YOU