Proteins or polypeptides are polymers of amino acids. They perform many essential functions in mammalian body.

These functions are: I - Dynamic functions: II- Structural functions: II-

I - Dynamic functions: a) Transport functions Albumin that transports some drugs, calcium, bile pigments and FFA Hemoglobin that transports oxygen. Transferrin that transports iron. * Lipoproteins that transport lipid.

y b) Metabolic control as they enters in the formation

of enzymes and some hormones, e.g. Insulin and glucagon.

y c) Contraction , e.g. Proteins of muscles

(actin and myosin)

y d) Protection, e.g. Immunoglobulins. y e) Blood clot e.g.Fibrinogen,thromboplastin,

and prothrombin.

II- Structural functions:

y a) Essential component of cell membrane

cytoplasm, cell organells and receptors.

y b) Enter in the structure of collagen,

elastin, keratin, and rhodopsin

y Sources of Dietary Proteins:

1- Animal : as milk, fish, meat and eggs. 2- Plant : as cereals and beans.

Protein Digestion :
It is not digested in the mouth due to absence of proteolytice nzymes.

Proteolytic enzymes (peptidases or proteases)

They are responsible for degradation of proteins

Proteolytic enzymes :
They are produced by three different organs:
1.The stomach, 2.The pancreas 3.Tthe intestine.

proteolytic enzymes:
A- Gastric proteolytic enzymes B- Pancreatic proteolytic enzymes C- intestinal proteolytic enzymes

N.B. : Protein is an antigen i.e. able to stimulate the immune system if it reaches blood in the form of large molecules (e.g. If it is taken intravenously). The digestion of protein to amino acids destroys its antigenicity. If protein is not digested completely and absorbed as polypeptide, immunologic response will occur and manifest itself as allergy in the form of urticaria , bronchial asthma and hay's fever.

Absorption: The end products of protein digestion are amino acids, di-and tripeptides. These are absorbed by epithelial cells via amino acid or peptide transport system. It is an active process against concentration gradient it needs Na+ as a co-transport system. ATP is the source of energy of this active process.

Fate of absorbed amino acids: Anabolic pathway Catabolic pathway

Anabolic pathway:
Amino acids enter in the formation of proteins for wear and tear, plasma proteins, hemoglobin, enzymes, some hormones

also enter in the formation of non protein nitrogenous compounds (NPN) as purines, pyrimidines, creatine and thyroxine.

yCatabolic pathway:
a) Urea: formed in the liver, is considered as the main metabolic endoproduct of protein catabolism. b) Supplying energy: 1 gram protein yields 4.1 K cal, only if there is shortage in carbohydrate and fats.

Nitrogen Balance There is no storage (depot) for protein, there is a certain percentage of protein that undergoes a constant process of breakdown and resynthesis i.e. turnover.

Nitrogen balance: is a comparison between the intake of nitrogen (mainly in the form of dietary protein) and the excretion of nitrogen (mainly in the form of undigested protein in stool and urea and ammonia in urine). Also nitrogen output is through nails, hair and desquamated skin.

Nitrogen equilibrium:
The normal adult human will be in nitrogen equilibrium when N2 lost (in urine, feces and sweat) just balanced by N2 in diet intake.

N2 LOST

N2 INTAKE

y Positive nitrogen balance:

A condition in which there is increase in the N2 intake over the output.

N2 INTAKE

>

N2 LOST

It may occur in growth, pregnancy or convalescence from diseases.

Negative nitrogen balance: A condition in which there is either decreased N2 intake as in : y starvation, poverty, y malnutrition,maldigestion, malabsorption, y severe vomiting, severe diarrhea Or increased N2 output as in y hemorrhage, burns, y old age or debilitating disease.
N2 LOST > N2 INTAKE

N.B. 1- Daily protein needs are One gram protein per kilogram body weight (i.e. about 70-100 gm protein per day). At least part of this protein should be of high biological value. A protein of high biological value should contain all essential amino acids.

2- essential amino acids are amino acids that are not synthesized in our body and hence, must be supplied in food. They are arginine histidine, valine, leucine, isoleucine, lysine, methionine, threonine, phenylalanine and tryptophan. If nine of the ten essential amino acids are present in certain type of food, this food is considered to contain protein of low biological value.

General Metabolism of Proteins : Complete breakdown of proteins and amino acids give rise to

Urea + Co2 + H2O + Energy.

y The major pathway for amino acids excess after protein

synthesis is the removal of the amino group and its conversion to ammonia (as there is no amino acid storage).
y The liver is the major site of removal of amino group from

amino acids..

The amino group is removed by different mechanisms:

y 1. Transamination y 2. Oxidative deamination y 3. Non-oxidative deamination y 4. Transdeamination

I . Transamination :
y It transfers the amino group from an amino acid to

keto acid.
y All the amino acids participate in the reaction of

transamination except threonine and lysine.

y Vitamin B6 is required as a coenzyme. y Its enzymes are termed transaminases

a) Aspartate transaminase: (AST) or(GOT)

COOH CHNH2 + CH2 CH2 COOH

COOH C O CH2 COOH

GOT

B6

COOH C O CH2 + CH2 COOH

COOH CHNH2 CH2 COOH

Glutamic acid Oxaloacetic acid

-Ketoglutaric acid Aspartic acid

b) Alanine transaminase: (ALT)or(GPT)

COOH COOH CHNH2 CH2 CH2 COOH
Glutamic acid Pyruvic acid
+

COOH C O C O CH3
GPT B6

COOH CHNH2
+

CH2 CH2 COOH

CH3

-Ketoglutaric acid

Alanine

Transaminases are cytosolic and mitochondrial enzymes. It is a freely reversible process.

Biological importance of Transamination

1- Synthesis of new non-essential amino acids. 2- Degradation of most amino acids except lysine and threonine. 3- Formation of components of citric acid cycle (filling up reaction of citric acid cycle). 4-Transaminase enzymes are used in diagnosis and prognosis of the diseases.

y N.B. Transaminase enzymes are present inside the cells

and small traces are present in the blood (5-40 IU/L).

y The increase in their level denote cell damage with the

release of enzymes from the destructed cells.

y E.g.

in cardiac infarction SGOT is increased in hepatic infection, SGPT is increased above the normal levels.

II. Oxidative deamination:

y It is catalyzed by : Amino acid oxidases Occur in liver and

kidney. y It includes removal of hydrogen (oxidation) and removal of NH3 (deamination). y There are D- and L-amino acid oxidases that oxidizes Dand L-amino acids respectively, to the corresponding -keto acids and the amino group is released as ammonia (NH3).

Oxidative deamination
NH2 R-CH-COOH
Aminoacid Flavin

NH
Amino acid oxidase

1
Flavin-H2

R-C-COOH
Iminoacid

H2O
2

NH3
H2O2 Catalase
1/2 O2

O2

O R-C-COOH
E -Ketoacid

H2O

y D-amino acid oxidase uses FAD as coenzyme which is of

limited natural occurance in mammals and of high activity,

y L-amino acid oxidase uses FMN as coenzyme which is of

natural occurrance in mammals, but of low activity.

COOH CHNH2 CH2 CH2 COOH

Oxidative
L-glutamic acid dehydrogenase

COOH

deamination

COOH C O CH2

C NH CH2 CH2 COOH Iminoacid H2 O NH3

CH2 COOH

NADP (NAD)

NADPH+H NADH+H+

Glutamic acid

-ketoglutaric acid

The reaction is both mitochondrial and cytoplasmic, occurs mainly in the liver and kidney. ATP and GTP are allosteric inhibitors while ADP and GDP activate the enzyme. It is a reversible reaction.

III - Non-oxidative deamination (direct deamination):

y The - amino group of serine and threonine

( amino acids containing hydroxyl group) can be directly converted to NH3 without removal of hydrogen. y This reaction is catalyzed by serine and threonine dehydratase which need pyriodoxal phosphate as coenzyme.

Non-oxidative deamination (direct deamination)

NH2
Serine

OH NH2 CH2-CH-COOH
L-serine

H2O

NH CH3-C-COOH H2O NH3 CH3-CO-COOH pyruvic acid

dehydratase

CH2=C-COOH

PLP

NonNon-oxidative deamination

IV .Transdeamination (L-Glutamate dehydrogenase):

NAD

Vit B6

NADP

Metabolism of ammonia

Sources of blood ammonia Fates of ammonia (Removal of ammonia)

Sources of blood ammonia: 1.From amino acids :

y Transdeamination y Oxidative deamination y Non-oxidative deamination .

2.From glutamine :

y Renal glutaminase y Intestinal glutaminase

Glutamine synthesis and ammonia formation

Glutamine synthetase

+
ATP ADP+p +H2O
Glutamic acid

glutaminase

H2O

Glutamine

Glutamic acid

3.From amines : whether dietary amine or monoamine hormones by amine oxidase. 4. From catabolism of purines and pyrimidines . 5.From bacterial action in the intestine either from dietary protein residue or from urea diffuses into the intestine

Fates of ammonia (Removal of ammonia):

y Amination of

-ketoacid to form non-essential amino acids and other biosynthetic reactions.

y Glutamine synthesis in the brain, liver, muscle and renal

tissues (4%).
y The majority of NH3 (90%) will produce urea in the liver

by urea cycle.
y Excretion in urine upto 1 gm /24 hours urine. y Traces in blood (up to 100 ug / dl).

Oxidative Deamination

Non Oxidative Deamination

Transdeamination

From bacterial action in the intestine on protein

Glutamine

4%

NH3

Purine and pyrimidine

1%

90 %
New aminoacid
Excretion in urine upto 1 gm /24 hours urine

Urea Traces in the blood up to 100 ug / dl

Sources and Fates of ammonia

y urea is released into the blood with a

level of

20 - 40 mg/dL
y It is the major end product of nitrogen

catabolism in humans representing 8090% of the nitrogen excreted.

Urea formation
NH2 3 ATP CO2 +2 NH3 CO NH2
urea

+ H 2O

y Five reactions each of them utilises

specific enzyme in urea cycle.

y The first 2 reactions of urea cycle are

mitochondrial and the rest 3 reactions are cytoplasmic.

mitochondria

cytoplasm

Five enzymes of urea cycle:

y Carbamoyl phosphate synthase 1 y Ornithine transcarbamoylase (citrulline synthase) y Argininosuccinate synthetase. y Argininosuccinase. y Arginase.

cytoplasm

mitochondria

Urea Cycle

Urea Cycle

Reactions (steps) of the urea cycle: Carbamoylphosphate formation: Using active CO2 , NH3 , 2 ATP and carabmoylphosphate synthase I, which is a mitochondrial enzyme active in presence of N-acetylglutamic acid.
carabmoylphosphate synthase I

CO2 + NH3 + 2ATP

H2N.CO. P + 2 ADP + P
Carbamoyl phosphate

UREA CYCLE
arginase
Carbamoyl phosphate synthase 1

Ornithine transcarbamoylase

argininosuccinase.

argininosuccinate synthetase

LINK BETWEEN KREBS' UREA CYCLE AND KREBS' 1 TRICARBOXYLIC ACID CYCLE:

2ATP CO2 + NH3 2

LINK BETWEEN KREBS' UREA CYCLE AND KREBS' TRICARBOXYLIC ACID CYCLE:

Malate

Oxalacetate

-ketoglutarate

Glutamate

1. The fumarate resulting from reaction number 4 (in Krebs urea cycle), under the influence of argininosuccinase, undergoes conversion to malate by fumarase enzyme. This malate forms oxaloacetate by malate dehydrogenase. The oxaloacetate undergoes Transamination by SGOT to form aspartate. This aspartate is needed in urea cycle at argininosuccinic synthase enzyme.

2.The CO2 used in urea cycle comes mainly from Krebs' tricarboxylic acid cycle. The first NH2 group comes from L-glutamic acid by L-glutamate dehydrogenase. The second NH2 group comes from amino group of aspartic acid.

y 1. Excess ammonia formation stimulates

REGULATION OF UREA CYCLE:

urea formation.

y 2. High arginine level stimulates N-acetyl

glutamate synthase enzyme, thus increases urea formation.

y 3.

High urea level inhibits carbamoylphosphate synthase (reaction 1), ornithine transcarbamoylase (reaction 2) and arginase enzymes (reaction 5). Carbamoylphosphate synthase is inactive in the absence of activator, N-acetylglutamate.

y 4.

METABOLIC DISORDERS OF UREA CYCLE:

There are five types of congenital hyperammonaemia. They affect children and manifested by 1. Vomiting, 2. Irritability, 3. Ataxia, 4. lethargy coma , 5. Mental retardation,and death

y 1.

Hyperammonaemia type I:

y It may be due to carbamoylphosphate synthase

deficiency.
y It causes increase in blood ammonia level (normally

plasma ammonia is less than 100 g/dL).

y It is a familial disease.

2.

Hyperammonaemia type II:

y It is due to ornithine transcarbamoylase deficiency. y It is X-chromosome linked deficiency. y There is increased glutamine in blood, CSF and urine

due to increased glutamine synthesis as consequence of increased tissue levels of ammonia.

3. Citrullinaemia type III:

y It is due to lack of argininosuccinic synthase . y It is recessive inherited disorder. y There is an increase in citrulline in plasma, CSF and

urine.

4.

Argininosuccinic aciduria type IV:

It is due to argininosuccinase deficiency.

y It is recessive inherited disorder. y There is increase in argininosuccinic in plasma, CSF

and urine.
y It is manifested at age of two years. y It usually ends in death early in life.

5.

Hyperargininaemia type V:

y It is due to arginase deficiency. y There is increase in arginine in blood, CSF and

urine.
y It affects children ( 1: 30,000 ) leading to mental

retardation ,coma and death.

Ammonia intoxication (Ammoniacal encephalopathy)

y

It is defined as toxicity of the brain due to increase in NH3 level in the systemic blood.

y This increased ammonia will be

fixed to - ketoglutaric acid to form glutamic acid then glutamine leading to interference with citric acid cycle so decrease ATP production in the brain cells.

yCauses:
I. Congenital: The 5 types of hyperammonaemia due to enzymes deficiencies in urea cycle.

II.

Acquired: 1. Liver disease as cirrhosis due to failure of urea formation and glutamine synthesis. 2. Portocaval shunt as in bilharziasis. 3. Gastrointestinal bleeding by action of bacterial flora on the blood urea and thus NH3 is released in large amounts.

Manifestations of ammonia intoxication:

y 1. Tremors y 2. Blurred vision y 3. Slurred speech y 4. Vomiting y 5. Confusion followed by coma

and death.

1. Restrict protein diet.

Treatment:

2. Injection of Glutamic acid and E-ketoglutaric acid: They act as a carrier for NH3 and combine with it to form a nontoxic material called glutamine. Glutamine passes to the kidney and by glutaminase yielding glutamic acid and NH3 excreted in urine as ammonium salt. (NH4cl).

3. Sodium benzoate and phenylacetate are given to conjugate with glycine and glutamine and rapidly the conjugates are excreted in urine. 4. Frequent small meals to avoid sudden increase in blood ammonia levels. 5. Removal of excess NH3 by dialysis in acute cases.