1. Urea cycle and its defects
Dr.S.Chakravarty MD

2. Learning objectives:
Explain the fate of carbon skeleton and nitrogen group of amino acids.
Explain the ways of transport of nitrogen from various parts of the body to the liver
Describe the urea cycle and the enzymes involved in production of urea in the liver
Define and classify Hyperammonemias . List the enzymes deficient in various hyperammonemias and its clinical features

3. Catabolism of Amino acids
Amino group – NH3Formation of urea
Carbon skeletons – Formation of Glucose and Ketone bodies.

4. FATE OF THE CARBON SKELETONS
Carbon skeletons are used for energy
Glucogenic: TCA cycle intermediates or Pyruvate (Gluconeogenesis)
Ketogenic: Acetyl CoA, Acetoacetyl CoA, or Acetoacetate

5. DIGESTION AND ABSORPTION
PROTEINS
Gastric juice ( acidity denatures proteins )
Intestinal enzymes hydrolyze
AMINO ACIDS
Amino acid transporters Na+Amino acid symporter (can take up di and tri peptides )
UPTAKE
DEFECTS :-
1)Hartnups disease  Long neutral amino acid transporter defect( Trp is not taken up  Pellagra like symptoms as Trp Niacin is not formed )
2)Cystinuria Basic amino acid transporter defect that also transports cysteineUrinary stones .Cystine forms Cystine by disulfide linkage (less soluble Stones )

6. Nitrogen Balance
Difference btw total nitrogen intake and total nitrogen loss in feces
POSITIVE NITROGEN BALANCE – MORE NITROGEN INGESTION THAN LOSS
Growing infants and pregnant women
NORMAL BALANCE – INTAKE MATCHES OUTPUT
Normal Individuals
NEGATIVE BALANCE – NITROGEN OUTPUT EXCEEDS INTAKE
Following surgery , cancer , malnutrition (decreased protein intake )

7. EXCESS AMINO ACIDS ARE DEGRADED NOT STORED
OVERVIEW OF AMINO ACID METABOLISM
ENVIRONMENT ORGANISM
Bio-
synthesis
Protein
Ingested
protein 2 3 1
Recycling a
AMINO
ACIDS b c
Degradation
(required) c
Purines
Pyrimidines
Porphyrins
Nitrogen
NH3
Carbon
skeletons
Urea
(ketogenic)
(glucogenic)
pyruvate
α-ketoglutarate
succinyl-CoA
fumarate
oxaloacetate
Used for
energy
acetoacetate
acetyl CoA

8. NH3 is
TOXIC

9. Why ammonia is toxic ? Affects central nervous system
Alkalization of intracellular compartment
Disrupts oxidative phosphorylation ATP depletion
Increased glutamate in Brain
Decreased Neurotransmitters – GABA – convulsions
Cerebral edema

10. Symptoms of AMMONIA toxicity
Flapping Tremor (Asterixis)
( Correlate flapping tremor later on with Liver failure in Clinical medicine )
Slurred Speech
Blurred Vision
COMA  Death

11. Metabolic fates of ammonia
Ammonotelic – Fishes
Lots of water available
Uricotelic – Reptiles and birds
Birds have to keep minimum body weight for flight
Ureotelic – Mammals

12. Adults degrade 1-2% of body protein daily
Body Proteins
80-85%
Amino acids
25%
Catabolism
UREA

13. Amino acids Sources of Amino Acids : Utilization of Amino acids:
Exogenous – Diet
Endogenous –
Breakdown of muscle protein
Biosynthesis from intermediates of citric acid cycle.
Synthesis of New proteins
Formation of Nucleotides
Formations of Porphyrins and Catecholamines
Production of energy and Ammonia.

14. Fate of Amino group
1) Reutilization: Glutamate and Glutamine are involved in recycling of amino acids.
ATP
ADP
Glutamate + Ammonia
Glutamine
Glutamine Synthase
They are secreted by the peripheral tissues in form of glutamine which is taken up by hepatocytes where the NH3 is re-used for amino acid and nucleotide synthesis
Glutamine
Glutamate + Ammonia
Glutaminase

15. 2. Formation of amino acids from carbon skeletons require ammonia
Two important reactions are involved in fixing ammonia back to amino acids:
Reductive Amination:
Amino Transferases:
All non-essential amino acids except for tyrosine and cysteine are derived and are dependent on transamination from glutamate.

16. Glutamate dehydrogenase
1. Reductive Amination :
NAD(P)H
NAD(P)
Alpha keto glutarate + Ammonia
Glutamate
Glutamate dehydrogenase
2. Transamination reaction:
Amino acid-1
Alpha keto acid-1
Transaminase
Alpha keto acid-2
Amino acid-2
PLP

17. UREA
Well balanced polarity (Quite uncharged because of amide nitrogen yet sufficiently soluble in plasma  No transporter required )
Non-Toxic
AMMONIA
ASPARTATE

18. CATABOLISM OF PROTEINS
UREA BIOSYNTHESIS IS DIVIDED INTO 4 STAGES:-
1. TRANSAMINATION
2. OXIDATIVE DEAMINATION
3. AMMONIA TRANSPORT
4. REACTIONS OF THE UREA CYCLE

19. TRANSAMINATION
DEF :- THE TRANSFER OF THE ALPHA-AMINO GROUP FROM ONE AMINO ACID TO A KETO ACID , RESULTING IN FORMATION OF A NEW AMINO ACID AND CORRESPONDING KETO ACID .
E.G :- REACTION CATALYZED BY ALANINE AMINOTRANSFERASE
ALANINE PYRUVATE
(AMINO ACID ) (CORRESPONDING KETO ACID )
α-KETOGLUTARATE GLUTAMATE
(KETO ACID ) (NEW AMINO ACID)
ALT
ALL TRANSAMINASES
REQUIRE PLP (VIT B6)
PLP

20. TRANSAMINATION
Corresponding KA AA
New AA KA

21. USES OF TRANSAMINATION
FIRST STEP OF CATABOLISM OF PROTEINS
SYNTHESIS OF NON-ESSENTIAL AMINO ACIDS and INTERCONVERSION OF AMINO ACIDS
REGENERATION OF TCA CYCLE intermediates

22. OVERALL FLOW OF NITROGEN IN AMINO ACID CATABOLISM

23. Sources of ammonia in the body
Aminoacids:
Transamination
Deamination
Transulfuration
Glutaminase
Gastrointestinal tract bacteria
Degradation of Amino sugars
Monoamine Oxidase
Pyrimidine catabolism

24. Nitrogen metabolism
Nitrogen part is toxic. Excreted in the form of either :
Ammonia – charged and alkaline. Excreted as ammonium ion in urine (3%)
Urea – Neutral molecule – Non toxic –( 80-85%)
Creatinine (3-4%) – constant in urine ( 1% of Creatine every day)
Uric acid – from Purines only !

25. Inter organ exchange of amino acids

26. The Glucose alanine cycle

27. Amino acid catabolism in various tissues:
Intestines:
predominantly use glutamine and asparagine for energy.
Liver :
based on the blood levels of Glutamine (starvation / anabolism) – produces Urea or proteins
Cannot metabolize branched chain amino acids
Muscle:
breaks down branched chain amino acids
Produces Alanine – transport form of ammonia to liver
Produces glutamine – to the kidneys.

28. Kaplan lecture notes USMLE step 1

29. Very important NH3 removal mechanism ( esp BRAIN)
Kaplan lecture notes USMLE step 1
Very important
NH3 removal mechanism
( esp BRAIN)

30. ` Kaplan lecture notes USMLE step 1
WHY ALANINE ? NOT GLUTAMATE DIRECTLY
(ALT)
(AST)
2ND NITROGEN
1ST NITROGEN

31. Sources of ammonia in the body
Aminoacids:
Transamination
Deamination
Transulfuration
Glutaminase
Gastrointestinal tract bacteria
Degradation of Amino sugars
Monoamine Oxidase
Pyrimidine catabolism

32. Nitrogen metabolism Nitrogen is Excreted in the form of either :
Ammonia – charged and alkaline. Excreted as ammonium ion in urine (3%)
Urea – Neutral molecule – Non toxic –( 80-85%)
Creatinine (3-4%) – constant in urine ( 1% of Creatine every day)
Uric acid – from Purines only !

33. Inter organ exchange of amino acids in post absorptive state (FASTING)

34. Inter organ exchange of amino acids in absorptive state (after feeding)

35. The Glucose alanine cycle
ALT (Transamination)
ALT (Transamination)

36. UREA
Well balanced polarity (Quite non polar because of amide nitrogen yet sufficiently soluble in plasma  No transporter required )
Non-Toxic
AMMONIA
ASPARTATE

37. Urea cycle Ammonia + Bicarbonate + ATP CPS -1 NAG – N-acetyl Glutamate
High protein Diet
Ammonia + Bicarbonate + ATP
CPS -1
NAG – N-acetyl Glutamate
activator
Carbomyl Phosphate
Ornithine Transcarbomylase
Ornithine
Citrulline
Mitochondria
Cytoplasm
Arginosuccinate synthase
Aspartate
Oxaloacetate
Arginosuccinate
Arginosuccinate Lyase
Fumarate
TCA cycle
Arginase
Arginine
Urea

39. Production of NAG: Acetyl Co-A + Glutamate N-acetyl Glutamate
N-acetyl glutamate synthase (NAGS)
Arginine
N-acetyl glutamate is the allosteric activator of Carbomyl phosphate synthase-1.

40. Urea cycle disorders Hyperammonemia Encephalopathy
Respiratory alkalosis
VOMITING
AVOIDANCE OF HIGH PROTEIN FOODS
INTERMITTENT ATAXIA
LETHARGY
SEVERE MENTAL RETARDATION

41. Symptoms of AMMONIA toxicity
Tremor
( Correlate flapping tremor later on with Liver failure in Clinical medicine )
Slurred Speech
Blurred Vision
COMA  Death

42. Disorders of UREA cycle
Usmle!
Hyperammonemia type -1
Hyperammonemia type -2
X-linked recessive
Defect in OTC
MC urea cycle defect
Orotic aciduria present
Cerebral oedema , coma and death.
Autosomal recessive
Defect in CPS- 1
1 in 200,000
No orotic aciduria
Cerebral Oedema , coma and death .
CAUSE OF OROTIC ACIDURIA Increased Carbamoyl phosphate  spills out from mitochondia to cytosolPyrimidine synthesis Orotic acid

43. 3. Citrullinemia :
Defect in arginosuccinate synthase
Citrullinuria
Autosomal recessive
4. Arginosuccinic aciduria:
Defect in arginosuccinate lyase
Arginosuccinic acid  blood, CSF, Urine
5. Hyperargininemia :
Diet without arginine
Defect in arginase enzyme

44. Treatment of Hyperammonemia:
Limit protein intake
Decrease bacterial source of ammonia – Antibiotics (Like Neomycin, Azithromycin)and Lactulose (purgative )
Replace intermediates of urea cycle – Arginine Citrulline, Aspartate
Remove excess ammonia – Hemodialysis, sodium benzoate, phenyl acetate
Very Important
Very Important

45. Drugs Combination of Sodium benzoate and Phenylacetate/Phenylbutyrate
Lactulose – Acidification – conversion to NH4+ and induction of Purgation
Mainstay
Gut sterilization :-Neomycin/Azithromycin other antibiotics
Very Important
Combination of Sodium benzoate and Phenylacetate/Phenylbutyrate
Sodium benzoate + Glycine  Hippuric acid  excreted
Phenylacetate  phenylacetyl glutamine  excreted. (Phenylacetate conjugates with glutamine to form phenylacetylglutamine, which is excreted by the kidneys)
Rarely used

47. Source http://biocadmin.otago.ac.nz

48. MCQ 1 Select the CORRECT answer .
The first reaction in the degradation of the majority of common amino –acids involves participation of :
NAD +
Pyridoxal Phosphate
Thiamine Pyrophosphate(TPP)
FAD
NAD and TPP

49. MCQ 2
After thorough investigations a man is diagnosed with orotic aciduria . To find out the cause of orotic aciduria which of the following investigations will you prefer?
A. ALP levels
B. vitamin b12 assay
C. FIGLU excretion assay
D. Peripheral smear
E. serum bilirubin

50. Thank you