1. Amino Acids:
Disposal of Nitrogen

2. Overview Amino acids are not stored in the body
So, Amino Acids must be obtained from :
1-Diet
2-Synthesized de novo
3-produced from normal protein degradation (turnover)
Amino Acids in excess of biosynthetic needs of the cell
Not stored
But: rapidly degraded
first step: removal of the a-amino group a-ketoacid
(of a.a.)
ammonia
exc. in urine UREA

3. Synthesis of proteins & other compounds Amino Acid
Diet
Degrad. of Proteins
De novo synthesis
Synthesis of proteins & other compounds Amino Acid
Degradation
a-Ketoacid Ammonia
(carbon skeleton) (nitrogen of a.a.)
Glucose Fatty acids other exc. In urine UREA
ketone Bodies compounds
Glycogen exc. in urine
ENERGY

5. Fate of amino acids obtained by tissue protein hydrolysis
Amino Acid Pool
100 grams of a.a.
Collected from:
Dietary Proteins (by hydrolysis)
Degradation of Tissue Proteins
De novo synthesis of a.a.
Fate of amino acids obtained by tissue protein hydrolysis
75% of amino acids used to synthesize new proteins
25% of amino acids metabolized
precursor for other compounds
compensated by dietary proteins ---- amino acids

6. Protein Turnover
Protein turnover results from the simultaneous synthesis & degradation of tissue proteins
The total amounts of protein in the body is constant because the rate of protein synthesis is just sufficient to replace the degraded protein
Protein turnover leads to hydrolysis & synthesis of grams of body protein each day

7. Rate of Protein Turnover
Short-lived proteins: minutes – hours half-life
(as many regulatory proteins & misfolded proteins)
Long-lived proteins: days – weeks half-life
(majority of proteins in the cell)
Structural proteins: months – years half-live
(as collagen)

8. By Two Major Enzyme Systems
Protein Degradation
By Two Major Enzyme Systems
1- Ubiquitin-proteasome mechanism
energy-dependent
mainly for endogenous proteins
(proteins synthesized within the cell)
2- Lysosomes
non-energy-dependen
primarily for extracellular proteins as:
- plasma proteins that are taken into cells by endocytosis
- cell surface membrane proteins: for receptor-mediated endocytosis

9. Ubiquitin-proteasome pathway

10. Mechanism of action of ubiquitin-proteasome system
Protein is covalently attached to ubiquitin (small globular protein)
More ubiquitin is added to form polyubiquitin chain (protein is tagged with ubiquitin)
Ubiquitin-tagged protein is recognized by the proteasome (proteolytic molecule)
The proteasome cuts the target protein into fragments
(requires ATP)
Fragments are cut by non-specific proteases to amino acids

11. Chemical Signals for Protein Degradation
Protein degradation is influenced by some structural aspect of the protein
Examples:
The half-life of a protein is influenced by the nature of the N-terminal
residue:
with serine: long-lived proteins (half-life is more than 20 hours)
with aspartate: short-lived (half-life is about 3 minutes)
Proteins rich sequence containing PEST:
rapidly degraded (i.e. with short half-life)
PEST sequence: proline, glutamate, serine & threonine

12. Digestion of Dietary Proteins
Most of the nitrogen in diet is consumed in the form of protein
Dietary protein/day: 70 – 100 grams
Dietary proteins must be hydrolyzed to amino acids by proteolytic
enzymes
Proteolytic enzymes are produced by three different organs
stomach
pancreas
small intestine

14. 1- Digestion of Proteins By Gastric Secretion
Gastric secretions contains:
1- hydrochloric Acid: - pH 2 – 3 (for activation pepsinogen)
- kills some bacteria
- denature proteins
2- Pepsin: secreted as peopsinogen (inactive zymogen)
activated to pepsin by Hcl or autocatalytically by pepsin
product of hydrolysis of proteins: peptides few free amino acids
(oligo- + poly-)

15. 2- Digestion of Proteins by Pancreatic Enzymes
On entering the small intestine:
Pancreatic Proteases:
Large polypeptides are further cleaved to oligopeptides & free amino acids
PROTEASES:
Secreted as inactive zymogen from pancreatic cells
Secretion of zymogens
is mediated by the secretion of cholecystokinin & secretin (polypeptide hormones of GIT)
Activation of zymogen;
by enteropeptidase (enterokinase): enzyme present on the luminal surface of intestinal mucosal cells
converts trypsinogen to trypsin (by removal of hexapeptide from trypsinogen)
Trypsin converts other trypsinogen molecules to trypsin
Trypsin activates all other pancreatic zymogens (chymotrypsinogen, proelastase &
procarboxypeptidases)
Specificity:
example
Trypsin: cleaves when the carbonyl group of the peptide bond is
contributed by argenine or lysine

17. Abnormalities in Protein Digestion
Deficiency of pancreatic secretion
occurs due to chronic pancreatitis, cystic fibrosis or
surgical removal of the pancreas
Digestion & absorption of fat & protein is incomplete
Abnormal appearance of lipids (steatorrohea) & undigested protein in feces

18. 3- Digestion of oligopeptides by enzymes of the Small Intestine
Aminopeptidases
- on the luminal surface of the intestine
- is an exopeptidase
- cleaves the N-terminal residue from oligopeptides
to produce free amino acids & smaller peptides

19. Transport of Amino acids into cells
Movement of a.a. to cells is performed by active transport (requires ATP)
Seven different transport systems:
with overlapping specificity for different amino acids
for example: cystine, ornithine, argenine & lysine are transported in kidney tubules
by one transporter
In cystinuria: Inherited disease , one of the most common inherited dis.
1: 7000 individuals
defective carrier system for these 4 amino acids
appearance of all 4 amino acids in the urine
precipitation of cystine to form kidney stones (may block urinary tract)

21. Removal of Nitrogen from Amino Acids
Removing the a-amino group
Essential for producing energy from any amino acid
An obligatory step for the catabolism of all amino acids

22. Amino Acids

23. Amino Acid

24. Amino Acid (deamination) removal of amino group (nitrogen)
Amino group carbon
(nitrogen) skeleton
(a-ketoacid)
incorporated
into
other excreted catabolised synthesis
Compounds of other compounds
(e.g. urea)
energy

25. Deamination Pathways
Amino group (nitrogen) is removed from an amino acid by either
1- Transamination (BY: TRANSAMINASES)
2- Oxidative Deamination (BY:GLUTAMATE DEHYDROGENASE)

26. 1- Transamination Funneling of amino groups to glutamate
a-ketoglutarate accepts the amino group from amino acids to become glutamate
By: aminotransferases (transaminases)
Glutamate:
Oxidat. Deam ammonia urea cycle Or
gives amino group to oxalacetate to
produce aspartate--- urea cycle
gives amino group to carbon skeleton to
produce new amino acid
All amino acids (with the exception of
lysine & threonine) participate in transamination

27. Substrate Specificity of Aminotransferases
Each aminotransferase is specific for one or few group of donors
Aminotransferase is named after the specific amino group donor (amino acid that donates its amino group)
The 2 most important aminotransferases
1- alanine aminotransferase (ALT)
2- aspartate aminotranspeptidase (AST)

28. ALanine AminoTransferase (ALT) & ASpartate AminoTransferase (AST)
amino acid
Carbon skeleton of
alanine
UREA CYCLE

29. Mechanism of Action of Aminotransferase

30. Equlibrium of Transamination Reactions
Equilibrium constant : ~ 1
Allowing the reaction to function in both directions
after protein-rich meal
amino acid degradation
(removal of amino group from amino acid a-keto acid)
supply of amino acids from diet is not adequate
amino acid biosynthesis
(addition of amino group to a-keto acid amino acid)

31. Diagnostic Value of Plasma Aminotransferases
Aminotransferases are normally intracellular enzymes
Plasma contains low levels of aminotransferases
representing release of cellular contents during normal cell turnover
Elevated plasma levels of aminotransferases
indicate damage to cells rich in these enzymes
(as physical trauma or disease to tissue)
Plasma AST & ALT are of particular diagnostic value

32. Diagnostic Value of Plasma Aminotransferases
1- liver disease:
Plasma ALT & AST are elevated in nearly all liver diseases
but, particularly high in conditions that cause cell necrosis as:
viral hepatitis
toxic injury
prolonged circulatory collapse
ALT is more specific for liver disease than AST
AST is more sensitive (as liver contains a large amount of AST)
2- Nonhepatic disease:
as: myocardial infarction
muscle disorders
These disorders can be distinguished clinically from liver disease

33. 2- Oxidative deamination of amino acids By: Glutamate Dehydrogenase
Oxidative Deamination of glutamate by glutamate dehydrogenase results in liberation of the amino group as free ammonia
(i.e. no transfer of amino group)
primarily in the liver & kidney (but can occur in other cells of the body)
Oxi Deamin. Of Glutamate provides:
1- a-ketoglutarate (can be reused for transamination of a.a.)
2- free ammonia urea cycle urea

34. Urea used for tranasmination
GLUTAMATE (from transamination)
Glutamate oxi. deamin. (liver & kidney (mainly) & others)
Dehydrogenase
ammonia a-ketoglutarate
Urea Cycle
Urea used for tranasmination
of a.a.

35. OXIDATIVE DEAMINATION by GLUTAMATE DEHYDROGENASE

36. Glutamate Dehydrogenase
Amino group of most amino acids are funneled to GLUTAMATE
(by transamination with a-ketoglutarate)
GLUTAMATE is the only amino acid that undergoes oxidative deamination (by glutamate dehydrogenase) to give a-ketoglutarate & ammonia
Amino Acids donate their amino group to
a-ketoglutarate to produce glutamate (Transamination)
Glutamate is oxi deamin. to a-ketoglutarate & ammonia
(by Glutamate dehydrogenase)

37. Amino acid + a-ketoglutarate ammonia
TRANS AMINASE GLUTA MATE UREA
CYCLE
DEHYDROGENASE
a-keto acid Glutamate
(carbon Skeleton)
metabolized
UREA
Energy other compounds
(catab.)

38. Removal of Amino Group from an Amino Acid
UREA
CYCLE
Amino
acid
Carbon skelton
of an amino acid
Coenzyme for glutamate dehydrogenase: NAD+

39. Coenzyme for glutamate dehydrogenase: NADPH
Synthesis of an Amino Acid from its carbon skeleton (reductive amination)
Amino
acid
Carbon skelton
of an amino acid
Coenzyme for glutamate dehydrogenase: NADPH

40. Combined Actions of Transaminases & Glutamate Dehydrogense reactions

41. Direction of Reactions
After protein ingestion
Glutamate level in liver is elevated
Reactions proceeds in direction of amino
acid degradation & formation of ammonia

42. Allosteric Regulation of Glutamate Dehydrogenase
ATP & GTP are allosteric inhibitors of glutamate dehydrogenase
ADP & GDP are allosteric activators of glutamate dehydrogenase
Energy in cells
glutamate degradation by glutamate dehydrogenase
Energy production
from the carbon skeleton of amino acids

43. D-amino acid oxidase a-keto acids Energy Reaminated ammonia
D-amino acids are found in plants & cell walls of microorganisms
Not used in synthesis of mammalian proteins
D-amino acids is available in diet (from plants)
Metabolized by D-amino acid oxidase (in liver) FAD-dependent
Oxidative Deamination of D-amino acids
a-keto acids
Energy Reaminated ammonia
L-amino acids UREA

44. Transport of Ammonia to Liver
in most tissues
ammonia + glutamate
Glutamine
synthase
By blood
to liver Glutaminase
Glutamate + ammonia

45. In skeletal muscles ammonia (by transamination)
Transamination of pyruvate to form alanine
Alanine is transported in blood to liver
In liver, alanine is converted to pyruvate &
ammonia (by transamination)
Pyruvate can be converted to glucose
(by gluconeogenesis)
Glucose can enter the blood and used by
sk. Muscles
(GLUCOSE - ALANINE PATHWAY)