The document summarizes the urea cycle, which occurs in the liver to convert ammonia into urea for excretion. It involves several steps spanning the mitochondria and cytosol. Carbamoyl phosphate synthetase activates ammonia and CO2 to initiate the cycle. Ornithine, aspartate, and several other compounds join the cycle through condensation reactions requiring ATP. Arginase produces urea and ornithine at the end of the cycle. The cycle is connected to the Krebs cycle and regulated by factors like dietary protein and N-acetyl glutamate. Deficiencies in cycle enzymes can cause diseases with high ammonia levels like hyperammonemia.

1. UREA CYCLE
NAUFA BACKER AND Dr. KAYEEN VADAKKAN
DEPARTMENT of BIOTECHNOLOGY
ST.MARY’S COLLEGE, THRISSUR

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Urea cycle

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Mechanism
● Urea cycle begins inside liver mitochondria but
three of the subsequent steps takes place in
the cytosol thus it spans two cellular
compartments.
● Ammonia combines with CO2 (as HCO3)
produced by mitochondrial respiration, to form
carbamoyl phosphate in the matrix.
● It is an ATP dependent reaction - catalyzed by
carbamoyl phosphate synthetase(CPS) 1- a
mitochondrial regulatory enzyme.
● CPS 2 - cytosolic form-separate function in
pyramidine biosynthesis.
● Carbamoyl phosphate - an activated
carbamoyl group donor - enters into urea
cycle.

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Major steps
● First, carbamoyl phosphate donates it's carbamoyl group to ornithine to
form citrulline, with the release of Pi.
● Reaction is catalysed by ornithine transcarbamoylase.
● Citrulline passes from mitochondria to the cytosol.

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● Second, the next amino group enter from aspartate (generated in
mitochondria by transamination and transported into the cytosol) by a
condensation reaction between the amino group of aspartate and carbamoyl
group of citrulline, forming argininosuccinate.
● It is a cytosolic reaction - catalysed by argininosuccinate
synthetase,requires ATP & proceeds through a citrullyl - AMP intermediate.
● Third, the argininosuccinate is then cleaved by argininosuccinase to form
free arginine & fumarate.
● Fumarate can enter mitochondria and join into citric acid cycle. It is the only
reversible step in urea cycle.
● Fourth, the cytosolic enzyme arginase cleaves arginine to yield urea and
ornithine.
● Ornithine is transported into the mitochondria to initiate another round of
urea cycle.

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Krebs bicycle
● Urea cycle and Krebs cycle are inter
connected - Krebs bicycle
● The linking pathways are known as
aspartate - argininosuccinate shunt.
● Citric acid cycle enzymes have both
cytosolic & mitochondrial isoenzymes.
● Fumarate produced in cytosol (by urea
cycle, purine biosynthesis or other
processes) can be converted to cytosolic
malate, which is used in the cytosol or
transported into mitochondria (via malate -
aspartate shuttle) to enter the citric acid
cycle.

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Regulation of urea cycle
Coarse Regulation
● The enzyme levels change with the protein
content of diet.
● During starvation,the activity of urea cycle
enzymes are elevated to meet the increased
rate of protein catabolism.
Fine Regulation
● The major regulatory step is catalysed by
CPS-1 where the positive effector is N-
acetyl glutamate (NAG). It is formed from
glutamate and acetyl CoA.
● Arginine is an activator of NAG synthase.

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Compartmentalization
● The urea cycle enzymes are located in such a way that the first two
enzymes are in the mitochondrial matrix.
● The inhibitory effect of fumarate on its own formation is minimized
because argininosuccinase is in the cytoplasm, while fumarase is in
mitochondria.

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Energetics of urea cycle
● Two ATP moleculs are required to make carbamoyl phosphate, and one
ATP to make argininosuccinate.
● Overall equation of urea cycle :
NH3 + CO2 + aspartate + 3 ATP + 2 H2O → urea + fumarate + 2 ADP + 2 Pi +
AMP + PPi
● Oxaloacetate - - - > fumarate (via aspartate) and the regeneration of
oxaloacetate produces NADH in the malate dehydrogenase reaction.
● Each NADH generate 2.5 ATP during mitochondrial respiration, greatly
reducing the overall energetic cost of urea synthesis.

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DISEASES ENZYME DEFICIT FEATURES
HYPERAMMONEMIA TYPE 1 Carbamoyl phosphate
synthetase - 1
Very high NH3 levels in blood,
mental retardation.
HYPERAMMONEMIA TYPE 2 Ornithine transcarbamoylase High level ammonia in blood,
increased glutamine in blood.
HYPERORNITHINEMIA Defective ornithine transporter
protein
Failure to import ornithine from
cytoplasm to mitochondria.
CITRULLINEMIA Argininosuccinate synthetase High levels of ammonia and
citrulline in blood.
ARGININOSUCCINIC
ACIDURIA
Argininosuccinase Argininosuccinate in blood and
urine.
HYPERARGININEMIA Arginase Arginine increased in blood and
CSF.

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