1. The Nitrogen Cycle
Nitrite reductase
Nitrate reductase
nitrogenase

2. Nitrogen Metabolism Many nitrogen containing compounds
eg. Amino acids, nucleotides, porphyrins, neurotransmitters
There is no dedicated store for nitrogen or nitrogen compounds in humans

3. Nonprotein nitrogen derivatives
Diet protein
Tissue protein
Carbohydrate
(glucose)
transamination
Nonprotein nitrogen derivatives
Amino nitrogen in glutamate
deamination
NH3
Urea
Acetyl-CoA
Citric
Acid
Cycle
CO2
Ketone bodies
Amino acid pool
Overview of the protein metabolism

4. OVERVIEW OF AMINO ACID METABOLISM
ENVIRONMENT ORGANISM
Bio-
synthesis
Protein
Ingested
protein 2 3 1 a
AMINO
ACIDS b c
Degradation
(required) c
Purines
Pyrimidines
Porphyrins
Nitrogen
Carbon
skeletons
Urea
(ketogenic)
(glucogenic)
pyruvate
α-ketoglutarate
succinyl-CoA
fumarate
oxaloacetate
Used for
energy
acetoacetate
acetyl CoA

5. Amino acids are the major source of dietary N

6. NITROGEN BALANCE

7. Nitrogen intake nitrogen excretion
Nitrogen Balance
An individual’s nitrogen balance is dependent on a combination of:
Nitrogen intake nitrogen excretion
Dietary amino acids, nucleotides etc.
Urine, faeces, hair and skin loss, perspiration
Nitrogen balance status can be:
1) In balance
2) Positive
3) Negative

8. NITROGEN BALANCE
Nitrogen balance = nitrogen ingested nitrogen excreted
(primarily as protein) (primarily as urea)
Nitrogen balance = 0 (nitrogen equilibrium)
protein ingested = protein excretion
Observed in adults
Positive nitrogen balance
protein ingested > protein excretion
during pregnancy, infancy, childhood , body building and recovery from severe illness or surgery
Negative nitrogen balance
protein ingested < protein excretion
starvation, following severe trauma, surgery or infections.
Prolonged periods of negative balance are dangerous and fatal if the loss of body protein reaches about one-third of the total body protein

9. Excess or insufficient dietary amino acid intake leads to the catabolism of amino acids
Excess amino acids can be used for energy
Insufficient dietary amino acids lead to the catabolism of proteins
Insufficient dietary energy leads to the catabolism of proteins
For amino acids to be utilised for energy, they must have their a-amino groups removed

10. Deamination of amino acids
Deamination generates:
a carbon skeleton a free amino group
can be used for anabolic or catabolic reactions
generally excreted

11. Some amino acids can be directly deaminated
Serine, threonine and glutamate can be directly deaminated
Glutamate deamination is catalysed by glutamate dehydrogenase (GDH)

12. Glutamine can be deaminated in a two step process
Glutamate is then deaminated by GDH
glutamine + H2O glutamate + NH3

13. Glutamine can also be synthesised from glutamate
Glutamine synthesis is an energy requiring reaction
The reaction is catalysed by glutamine synthetase (GS)
glutamate + NH4+ + ATP glutamine + ADP + Pi GS

14. TRANSAMINATION

15. Transamination
Those amino acids that can not be directly deaminated have their amino groups transferred to specific substrates
These substrates are keto acids found in intermediary metabolism
a - ketoglutarate
oxaloaceatate
pyruvate
CAC

16. Addition of amino groups to these keto acids generates amino acids
a - ketoglutarate
oxaloacetate
pyruvate
glutamate
aspartate
alanine
Most amino acids are deaminated by donating their
a-amino acids to one of these keto acids
Thus the deamination of most amino acids leads to the production of either glu, asp, ala or gln.

17. An example transamination
glutamate
a-KG
a-amino acid
a-keto acid
glutamate aminotransferase

18. Pyridoxal phosphate Derived from vitamin B6
Takes part in all amino transferase reactions
Forms a Schiff base intermediate with substrates

20. Role of transamination in metabolism
Transamination allows for:
1) the generation of amino acids in short supply
2) the provision of carbon skeletons for energy generation
3) the safe removal of excess amino groups

21. However when ammonia concentrations are high:
Free ammonia is a by-product of brain metabolism
glutamate + NH4+ + ATP glutamine + ADP + Pi
The neurotransmitter GABA is inactivated by deamination GS
GDH
a-ketoglutarate + NH4+ + NADPH
glutamate + NADP+ + H2O
However when ammonia concentrations are high:
Brain requires large amounts of ATP
This must be generated via oxidative phosphorylation
Therefore the CAC must function efficiently
GABA – Gamma Amino Butyric Acid

23. Free ammonia is also produced in muscle
Amino groups can be liberated:
during normal muscle turnover
during starvation
during severe muscle activity
ATP ADP + Pi
2ADP ATP + AMP
AMP IMP + NH4+
AMP deaminase

24. alanine aminotransferase
Pyruvate is usually abundant in active muscle
Muscle uses pyruvate as an acceptor keto acid
glutamate + pyruvate a-ketoglutarate + alanine
alanine aminotransferase
Thus in muscle most amino groups are shuttled to alanine (via glutamate)
Alanine is then exported to the liver where the amino groups can be liberated

25. AMP

26. FATE OF AMINO GROUP DEAMINATION A. Transamination
B. Oxidative deamination
C. purine nucleotide cycle

27. A. Transamination Transamination by Aminotransferase (transaminase)
always involve PLP coenzyme (pyridoxal phosphate)
reaction goes via a Schiff’s base intermediate
all transaminase reactions are reversible

28. Aminotransferases
Aminotransferases can have specificity for the alpha-keto acid or the amino acid
Aminotransferases exist for all amino acids except proline and lysine
The most common compounds involved as a donor/acceptor pair in transamination reactions are glutamate and a-ketoglutarate, which participate in reactions with many different aminotransferases
to an alpha-keto acid  alpha-amino acid

29. Transamination
aminotransferases

30. Glu+pyruvate
glutamate-pyruvate aminotransferase
GPT, ALT
-Ketoglutarate+Ala
Glu+Oxaloacetate
Glutamic oxaloacetictransaminase
GOT, AST
-Ketoglutarate+Asp
*** ALT and AST are components of a "liver function test". Levels increase with damage to liver (cirrhosis, hepatitis) or muscle (trauma)

31. The mechanism of transamination+
PLP AA
–H2O
+H2O
Schiff’s base

32. Schiff’s base isomer
Molecule rearrange
PMP
–H2O
+H2O
-ketoacid +

33. Transamination Interconversion of amino acids Collection of N as glu
Provision of C-skeletons for catabolism

34. B. Oxidative Deamination
L-glutamate dehydrogenase (in mitochondria)
Glu + NAD+ (or NADP+) + H2O  NH4+ + a-ketoglutarate + NAD(P)H +H+
Requires NAD+ or NADP + as a cofactor
Plays a central role in AA metabolism ?

35. It is inhibited by GTP and ATP, and activated by GDP and ADP
urea cycle ?
It is inhibited by GTP and ATP, and activated by GDP and ADP

36. Combined Deamination ?

37. Transamination + Oxidative Deamination
Combined deamination ＝
Transamination + Oxidative Deamination
The major pathway ！！！

38. C. purine nucleotide cycle AMP
NH3 AA
Asp
IMP
-Keto
glutarate
H2O
aminotransferases
AST
C. purine nucleotide cycle
AMP
-Keto
acid
Oxaloacetate
fumarate
malate

39. The metabolism of α-ketoacid
Biosynthesis of nonessential amino acids
TCA cycle member + amino acid α-keto acid + nonessential amino acid
A source of energy (10%) ( CO2+H2O )
Glucogenesis and ketogenesis

40. SUMMARY
Nitrogen balance status depends on the intake and use of N containing compounds
Excess N from amino acids must be excreted
A series of aminotransferase and deamination reactions shuttle nitrogen to appropriate molecules and tissues
Brain and muscle can generate large amounts of excess nitrogen as part of their metabolism
The liver is an important tissue for processing excess nitrogen