Protein Metabolism

1. Protein Metabolism
Dr. Azad Alam Siddiqui
Assistant Professor
Department of B.Voc (MMDT)
Km. Mayawati Govt.Girls PG College
Badalpur, G.B. Nagar (U.P.)

2. Protein Metabolism
• Protein metabolism denotes the various biochemical processes responsible for the
synthesis of proteins and amino acids, and the breakdown of proteins (and other
large molecules) by catabolism.
• Catabolism of amino acids provides carbon skeleton for gluconeogenesis,
ketogenesis, as also for energy yielding pathways.
• Much of the body is made of protein, and these proteins take on a myriad of forms:
1. Catalyst
2. Transport
3. Mechanical support
4. Protection
5. Regulation and communication
6. Blood buffer

3. Protein Metabolism
• Protein metabolism is effectively the metabolism of amino acids.
• Proteins are also sources of energy but supply only a small fraction ̴ 15%.
• Protein undergo turnover constantly.
• Important role of dietary protein is to supply essential amino acids.The
dietary proteins are denatured on cooking and therefore more easily digested.
• Protein Digestion:
a. Digestion in mouth
b. Digestion in stomach
c. Digestion in intestine
• End product of protein digestion is amino acid, dipeptide and tripeptides.

4. DIGESTION OF PROTEINS
• Proteolytic enzymes are secreted as inactive zymogens which are converted to their active form in the
intestinal lumen. The proteolytic enzymes include:
➢Endopeptidases: They act on peptide bonds inside the protein molecule, so that the protein
becomes successively smaller and smaller units. This group includes Pepsin, Trypsin,
Chymotrypsin and Elastase.
➢Exopeptidases: Which act only on the peptide bond located at the ends of the polypeptide chain.
This group includes:
a) Carboxypeptidase, which acts only on the peptide bond at the carboxy terminal end of the
chain.
b) Aminopeptidase, which acts only on the peptide bond at the amino terminal end of the chain.
• Intracellular Protein Degradation:
1. Cathepsins 2. Ubiquitin 3. Proteasomes
• The proteins on degradation (proteolysis) releases individual amino acids. Each one of the 20
naturally occurring amino acids undergoes its own metabolism and performs specific functions.

5. Absorption of free amino acids
Amino acids, on
absorption from intestine
are carried to Liver
through portal blood.
They are taken up by
Liver cells to some extent
and remainder enters the
systemic circulation and
diffuse throughout the
body fluids and taken up
by tissue cells.

6. AMINO ACID POOL
1. Proteins are the most abundant organic compounds, and constitute a
major part of the body dry weight (10-12 kg in adults).
2. Despite the great complexity of protein synthesis, proteins are made
at exceedingly high rates. A polypeptide of 100 residues is
synthesized in an Escherichia coli cell (at 370C) in about 5 seconds.
3. The main role of amino acids is in the synthesis of structural and
functional proteins.

7. AMINO ACID POOL
1. There is also a continuous synthesis of amino acids (except the
“essential” amino acids). Amino acids from all the sources get mixed up
to constitute what is known as “general amino acid pool” of the body.“
2. An adult has about 100 g of free amino acids which represent the amino
acid pool of the body. Glutamate and glutamine together constitute
about 50%, and essential amino acids about 10% of the body pool (100
g).
3. If a cell takes up as much amino acids as it loses, it is in a state of
“dynamic equilibrium”, if the loss is greater, the cell wastes, and if the
gain is greater the cell grows.

8. Protein turnover
• 300-400 g of protein per day is constantly degraded and synthesized which represents body
protein Turnover.
• There is a wide variation in the turnover of individual proteins. For instance, the plasma
proteins and digestive enzymes are rapidly degraded, their half-lives being in hours or
days. The structural proteins (e.g. collagen) have long half-lives, often in months and
years.(Certain proteins with amino acid sequence proline, glutamine, serine and threonine
rapidly degraded).

9. Tissue Amino acids
➢The amino acids are transported into tissues actively. Pyridoxal-P (B6-P) is one of the requirement for
this active transport. Tissue uptake is also favoured by hormones:
➢Insulin, growth hormone and testosterone favour the uptake of amino acids by tissues (anabolic
hormones).
➢Oestradiol stimulates selectively their uptake by uterus.
➢Epinephrine and glucocorticoids: Stimulate the uptake of amino acids by the Liver.

10. NITROGEN BALANCE
• In an adult to maintain constant weight, the amount of intake of N in food will
be balanced by an excretion of an equal amount of N in urine and in faeces. The
individual is then said to be in nitrogen balance or nitrogenous equilibrium.
• A subject whose intake of N is greater than the output, e.g. in growth, is said to
have a +ve nitrogen balance. In the growing period and also during
convalescence from illness or when anabolic hormones are given, the body puts
on weight.
• A subject whose intake of N is less than the output of N, (e.g. in losing weight),
is said to have a –ve nitrogen balance. In old age and during illness and
starvation weight is lost and results in –ve nitrogen balance.

11. DISSIMILATION OF AMINO ACIDS
• α-NH2 group of amino acids, derived either from the diet or breakdown of tissue proteins,
ultimately is converted first to NH3 and then to urea and is excreted in the urine.
• Formation of NH3 and urea can be discussed under the following heads:
1. Transamination
2. Deamination Oxidative deamination
Non-oxidative deamination
3. Transdeamination
4. NH3 transport, and
5. Formation of urea
• Urea is the characteristic end-product of amino acid catabolism in human beings and
ureotelic organisms.

12. TRANSAMINATION
• Transamination is a reversible reaction in which α-NH2 group of one amino acid is
transferred to a α-keto acid resulting in formation of a new amino acid and a new keto acid.
• Important for the production of non essential amino acids.
• The process represents only an intermolecular transfer of NH2 group without the splitting
out of NH3. Ammonia formation does not take place by transamination reaction.

14. • There are two transaminases of clinical importance in the body in that they use specific
amino acid and specific ketoacid. These two specific transaminases are:
• Aspartate transaminase (S-GOT): Aspartic acid is the donor amino acid and α-
oxoglutarate is the recipient ketoacid. New amino acid formed is always glutamic acid.
✓Concentration of the enzyme is high in myocardium and also in liver cells. Helpful in
acute myocardial infarction.
✓Increases in Liver diseases, but it is less than Alanine transaminase (S-GPT).
✓Increase in muscular dystrophies—myositis.
✓Increased activity seen in acute pancreatitis, leukaemia's, in acute haemolytic anaemia.
✓In normal persons, after prolonged severe exercise.
✓A rise has been seen in therapy with certain antibiotics.

15. • Alanine transaminase (S-GPT): Alanine is the donor amino acid and α-
oxoglutarate is the recipient ketoacid. New amino acid formed is again always
glutamic acid.
✓The enzyme is found mainly in Liver.
✓Increases in both transaminases are common finding in hepatic diseases but always
S-GPT > than S-GOT, though in normal healthy persons, S-GOT is slightly more
than S-GPT.
✓It is most useful in assessing severity and progress of the disease in acute viral
hepatitis.
✓Highest values of enzyme activity seen in acute viral hepatitis, peak values 250 to
1500 IU/L or more seen at the time of maximum illness.

16. DEAMINATION
• Deamination is the process by which N– of amino acid is removed as NH3.
A. Oxidative deamination
B. Non-oxidative deamination.
(A)
• L-amino acid oxidase, a flavoprotein is restricted to liver and kidney only and thus does not fulfil
a major role in mammalian amino acid catabolism and formation of NH3.

17. • D-amino acids are found in plant and micro-organism, and not present in
mammalian tissue.
• They are regularly taken in diet.
• D-amino acid oxidase convert D-amino acids into respective α-keto acids
by oxidative deamination.
• α-keto acids undergo transamination to be converted to L-amino acids
which participate in various metabolism.
• D-amino acid oxidase is important as it initiates the step for the conversion
of unnatural D-amino acids to L-amino acids in the body.

18. • Purpose of Deamination is to provide NH3 for urea synthesis and α-keto acids for
variety of reactions including energy generation.
• L-glutamic acid is not deaminated by L-amino acid oxidase but by a specific
enzyme called L-glutamate dehydrogenase (Zn++-containing metalloenzyme).
• It is widely distributed in tissues in humans and has high activity, and is specific for
L-Glutamate.
• Glutamate serves as collection centre for amino groups.
• GDH involved in both catabolic and anabolic reactions.

19. • Reaction is reversible, and the
equilibrium constant favours
glutamate formation, but the
quick removal of NH3 to form
urea in urea cycle and α-
Ketoglutarate to TCA cycle
favours onward reaction, i.e. NH3
formation

20. (B) There are certain amino acids, which can be nonoxidatively deaminated by
specific enzymes, and can form NH3. These reactions do contribute to NH3
formation, but again they do not fulfil, a major role in NH3 formation.
amino acid dehydrases

21. • Transamination takes place in the cytoplasm
of all the cells of the body; the amino group
is transported to liver as glutamic acid,
which is finally oxidatively deaminated in
the mitochondria of hepatocytes.
• Thus, the two components of the reaction
are physical far away, but physiologically
they are coupled. Hence the term
transdeamination.
• This mechanism seems to be the major
pathway for removal of NH2 group from an
L-amino acid and formation of NH3.
TRANSDEAMINATION

22. NH3 TRANSPORT
• In addition to NH3 formed in the tissues, a considerable quantity of NH3 is produced in
the gut by intestinal bacterial flora, both
• From dietary proteins, and
• From urea present in fluids secreted into the GI tract.

23. • NH3 is absorbed from the Intestine into portal venous blood and thus has more NH3
than systemic blood.
• Liver promptly removes the NH3 from the portal blood, so that blood leaving the
liver is virtually NH3-free. This is essential since even small quantities of NH3 are
toxic to CNS.
• NH3 is not just a waste product of nitrogen metabolism. lt is involved (directly or via
glutamine) for the synthesis of many compounds in the body. These include
nonessential amino acids, purines, pyrimidines, amino sugars, asparagine etc.
Ammonium ions (NH4
+) are very important to maintain acid-base balance of the
body.

24. • The transport of NH3 between various tissues and the liver mostly occurs in the
form of glutamine or alanine and not as free NH3.
• NH3 is always produced by almost all cells, including neurons. The
intracellular NH3 is immediately trapped by glutamic acid to form glutamine,
especially in brain cells. The glutamine is then transported to liver, where the
reaction is reversed by the enzyme glutaminase. The NH3 thus generated is
immediately detoxified into urea.

25. Why NH3 is Toxic?
• Normal range of NH3 in blood is 10-80µg/dl.
• Increased NH3 concentration depressing the TCA cycle, affecting the cellular
respiration.
• Increased NH3 concentration enhances glutamine formation from Glutamate and
thus reduces ‘brain cell’ pool of glutamic acid. Hence there is decreased formation
of inhibitory neurotransmitter GABA.
• Rise in brain glutamine level enhances the outflow of glutamine from brain cells.
Glutamine is carried ‘out’ by the same “transporter” which allows the entry of
‘tryptophan’ into brain cells. Hence ‘tryptophan’ concentration in brain cells
increases which leads to abnormal increases in synthesis of “serotonin”, a
neurotransmitter.

26. UREA FORMATION (KREBS-HENSELEIT CYCLE)
• Steps of urea synthesis have been elucidated by Krebs and Henseleit (1932).
• It is a cyclic process, five reactions which involves ornithine, citrulline, arginine and
aspartic acid.
• Kidneys: Urea cycle operates in a limited extent. Kidney can form up to arginine
but cannot form urea, as enzyme arginase is absent in kidney tissues.
• Brain: Brain can synthesise urea from citrulline, but lacks the enzyme for forming
citrulline from ornithine. Thus, neither the kidneys nor the brain can form urea in
significant amounts.
• Enzyme for Urea cycle is partly mitochondrial and partly cytosolic.
• 1 mol. of NH3 and one mol. of CO2 are converted to 1 mol. of urea for each turn of
the cycle and ornithine is regenerated at the end, which acts as a catalytic agent.
• The overall process in each turn of cycle requires 3 moles of ATP.

27. Biosynthesis of urea or ornithine—urea cycle

28. Energetics of Urea Cycle
• The overall reaction may be summarized as:
NH3 + CO2 + Aspartate → Urea + Fumarate
• 2 ATPs are used in the first reaction, another ATP is converted to AMP and PPi.
So the urea cycle consumes 4 high energy phosphate bonds.
• Fumarate formed in the 4th step may be converted to malate. Malate when
oxidized to oxaloacetate produces 1 NADH equivalent to 2.5 ATP.
• So, net energy expenditure is only 1.5 high energy phosphates. The urea cycle
and TCA cycle are interlinked, and so, it is called as "urea bicycle".

29. Significance of Urea Cycle
1. Detoxification of NH3: Major biological role of this pathway is the
detoxification of NH3. Toxic ammonia is converted into a nontoxic substance
urea and excreted in urine.
2. Biosynthesis of arginine: The urea cycle also serves for the biosynthesis of
arginine from ornithine in liver, kidney and intestinal mucosa. Kidney and
intestinal mucosa probably contribute most of the body arginine because they
possess all the urea cycle enzymes except arginase. Hence they can form upto
arginine and cannot form urea. The arginine is used for protein synthesis.
3. The urea cycle is linked to the TCA cycle through the production of fumarate.
Amino acid catabolism is therefore directly coupled to energy production.

30. DECARBOXYLATION REACTION AND BIOGENIC AMINES
• Decarboxylation is the reaction by which CO2 is removed from the COOH group of an
amino acid as a result an amine is formed.
• The reaction is catalysed by the enzyme decarboxylase, which requires pyridoxal-P (B6-
PO4) as coenzyme.
• Tissues like liver, kidney, brain possess the enzyme decarboxylase and also by
microorganisms of intestinal tract.

31. • Histamine:
• Acts as neurotransmitter in hypothalamus and is a mediator of anaphylactic shock and
inflammation.
• Actions are mediated by both histamine H1 and H2 type of receptors.
• Drugs that blocks both receptors are known as antihistamines.(Promethazine and
mepyramine are H1 blocker and cimetidine is H2 blocker.)
• GABA:
• Decarboxylation of glutamic acid by glutamate-a-decarboxylase produces GABA.
(coenzyme B6-P in presence of Mg++)
• It is inhibitory neurotransmitter of CNS and retina.
• GABA binds to 2 distinct receptors GABA-A and GABA-B.

32. METABOLISM OF INDIVIDUAL AMINO ACIDS
• GLYCINE:
• Simplest amino acid. non-essential. Though it is non-essential but it is an
important amino acid as it forms many biologically important compounds in the
body.
• Neutral, aliphatic, Optically inactive and glucogenic.
• Most abundant amino acid normally excreted into urine.

33. Glycine is formed from:
1. Serine:
2. Threonine:

34. Aromatic Amino Acids
➢PHENYLALANINE AND TYROSINE:
• Phenylalanine and tyrosine are structurally related aromatic amino acids.
• Phenylalanine is an essential amino acid while tyrosine is non-essential amino
acid.
• Only function of phenylalanine is its conversion to tyrosine.
• Phenylalanine will be required in minimal, if adequate tyrosine is supplied in the
food. This is called the sparing action of tyrosine on phenylalanine.
• Tyrosine is incorporated into proteins and is involved in the synthesis of a variety
of biologically important compounds-epinephrine, norepinephrine, dopamine,
thyroid hormones and the pigment melanin.
• During the course of degradation, phenylalanine and tyrosine are converted to
metabolites which can serve as precursors for the synthesis of glucose and fat.
Hence it is partly glucogenic and partly ketogenic.

35. Phenylalanine to Tyrosine
• The reaction involves addition of a
hydroxyl group to the aromatic ring, by
phenylalanine hydroxylase.
• Needs NADPH, NADH and
tetrahydrobiopterin as co-enzymes.
• Irreversible reaction, tyrosine cannot
replenish phenylalanine. Hence,
phenylalanine is essential in food.
• One molecule of O2 is needed in this
reaction; out of which one atom is
incorporated in the OH group and the
other is reduced to water.

36. • Due to a defect in phenylalanine hydroxylase, the conversion of
phenylalanine to tyrosine is blocked resulting in the disorder
phenylketonuria (PKU)
• Accumulated phenylalanine makes phenylpyruvate by transamination and
phenyl lactate by reduction which are excreted in urine.
• Phenylpyruvate is a phenyl ketone, hence the name phenylketonuria.
• Phenylpyruvate and phenyl lactate are excreted in traces in normal person.
• Tyrosine is not formed and so it becomes an essential amino acid.
• If tyrosine intake is not adequate, there is decrease formation of
catecholamines, melanin etc.
PHENYLKETONURIA (PKU)

37. • Due to genetic mutation either the enzyme is not synthesized, or a non-functional
enzyme is synthesized.
• Frequency of PKU was considered to be 1 in 10,000 births; but recent introduction
of better diagnostic facilities showed that the incidence is as high as 1 in 1,500
births.
• There are 5 types of PKU described. Type I is the classical one. It is due to
phenylalanine hydroxylase deficiency. Types II and III are due to deficiency of
dihydrobiopterin reductase. Type IV and V are due to the deficiency of the enzyme
synthesizing biopterin.
• The classical PKU child is mentally retarded with an IQ of 50.
• Agitation, hyperactivity, tremors and convulsions are often manifested.
• The child often has hypopigmentation.

38. Dopamine and Parkinson's disease
• Parkinson's disease is a common disorder in many elderly people, with about 1% of
the population above 60 years being affected. lt is characterized by muscular
rigidity, tremors, expressionless face, lethargy, involuntary movements etc.
• The exact biochemical cause has not been identified. however, Iinked with a
decreased production of dopamine. The disease is due to degeneration of certain
parts of the brain (substantia nigra and locus coeruleus), leading to the impairment
in the synthesis of dopamine.
• Dopamine cannot enter the brain, hence its administration is of no use. DOPA
(levodopa or L-dopa) is used in the treatment of Parkinson's disease. In the brain,
DOPA is decarboxylated to dopamine which alleviates the symptoms of this
disorder.
• Carbidopa and γ-methyl-dopa (dopa analogs) are administered along with dopa for
the treatment of Parkinson's disease.

39. ALKAPTONURIA
• This is based on the observation that the urine becomes black on standing.
Homogentisate in urine is oxidized and polymerized to brownish black color.
• The disease is inherited and it is due to the deficiency of the enzyme required for
further metabolism of homogentisic acid.
• Alkaptonuria is an autosomal recessive condition with an incidence of 1 in 250,000
births.
• The metabolic defect is the deficiency of homogentisic acid oxidase. This results
in excretion of homogentisic acid in urine.
• The homogentisic acid is oxidized by polyphenol oxidase to benzoquinone acetate.
It is then polymerized to black coloured alkaptone bodies.
• No specific treatment is required. But minimal protein intake with phenylalanine
less than 500 mg/day is recommended.
• Ferric chloride test will be positive for urine.
• Benedict’s test is strongly positive.

40. One Carbon Metabolism
• There is a group of biochemical reactions that have a special set of enzymes and
coenzymes. They are involved in amino acid metabolism. This group of reactions is
referred to as one-carbon metabolism.
• Amino acid metabolism is particularly important for the transfer or exchange of one
carbon units.
• Methyl (-CH3)
• Hydroxymethyl (-CH2OH)
• Methylene (=CH2)
• Methenyl (-CH=)
• Formyl (-CH=O)
• Formimino (-CH=NH)
• Tetra hydro folic acid (THF) is a versatile coenzyme that actively participates in one
carbon metabolism. Vitamin B12 is also involved besides THF.

41. Generation of one carbon units
• The formyl released from glycine and tryptophan metabolism combines with THF
to form N10-formyl THF.
• Histidine contributes formimino fragment to produce N5-formimino THF.
• When serine is converted to glycine, N5, N10-methylene THF is formed.
• The different derivatives of THF carrying one carbon units are interconvertible,
and this is metabolically significant for the continuity of one- carbon pool.
• One-carbon fragments from THF are used for the synthesis of a wide variety of
compounds. These include purines, formyl methionine tRNA (required for
initiation of protein synthesis), glycine, pyrimidine nucleotide etc.