Protein Metabolism: Protein metabolism is an essential part of metabolism. Since

amino-acid metabolism is closely connected with the metabolism of other nitrogen compounds, protein metabolism

is often included in the more general concept of nitrogen metabolism. In autotrophic organisms—that is, plants (except fungi) and chemo-synthesizing bacteria—protein metabolism

begins with the assimilation of inorganic nitrogen and synthesis of amino acids and amides. In man and animals,

only a portion of the amino acids—the so-called nonessential ones—can be synthesized in the organism from simpler

organic compounds. The other portion—the essential amino acids—must be obtained from food, usually as protein.

Proteins contained in various foods are broken down by cleavage under the action of such proteolytic enzymes as

pepsin, trypsin, and chymotrypsin into amino acids, which are absorbed into the blood and carried to organs and tissues.

Plant tissues also contain proteolytic enzymes that hydrolytically break up proteins. The succeeding

processes of protein metabolism in plants and animals are essentially amino-acid metabolism.

A considerable portion of amino acids are used in the formation and completion of various proteins in the

body, including functionally active proteins (enzymes, hormones, antibodies, and so forth), plastic proteins, structural proteins, and others. At the same time, the

body’s proteins undergo constant breakdown and renewal, replenishing the reserve of free amino acids.

The other portion of the amino acids is used in the formation of a number of low-molecular hormones, biologically active peptides, amines, pigments, and other substances necessary for the maintenance of life. For example, the amino acid glycine is used to

form purine bases, and aspartic acid is used to synthesize pyrimidine bases.

The mutual transformation of amino acids is, in significant measure, produced by a process that is widespread in all organisms—the enzyme process,

involving the transfer of amino groups. This process, called transamination, was discovered by the Soviet scientists A. E. Braunshtein and M. G. Kritsman. Excess amino acids undergo enzyme

processes of decomposition. The most common initial reaction of amino-acid

decomposition is deamination, primarily oxidative deamination, after which the nitrogen-free

remainder of the amino-acid molecule degrades to the end products—carbon dioxide, water, and nitrogen that splits off in the form of ammonia.

In man and animals:

The transformation and fate of food proteins from their ingestion to the elimination of their

excretion products: Proteins are of exceptional importance to

organisms because they are the chief constituents, aside from water, of all the soft

tissue of the body. Special proteins have unique roles as structural and functional

elements of cells and tissues. Examples are keratin of skin, collagen of tendons, actin and

myosin of muscle, the blood proteins, enzymes in all tissues, and protein hormones of the

hypophysis.

Protein is digested to amino acids in the gastrointestinal tract. These are absorbed and distributed among the different tissues, where they form a series of amino acid pools that are kept equilibrated with each other through the

medium of the circulating blood. The needs for protein synthesis of the different organs are

supplied from these pools. Excess amino acids in the tissue pools lose their nitrogen by a

combination of transamination and deamination. The nitrogen is largely converted to urea and excreted in the urine. The residual carbon products are then further metabolized

by pathways common to the other major foodstuffs—carbohydrates and fats.

Protein digestion occurs to a limited extent in the stomach and is completed in the

duodenum of the small intestine. The main proteolytic enzyme of the stomach is pepsin,

which is secreted in an inactive form, pepsinogen. Its transformation to the active pepsin, initiated by the acidity of the gastric juice, involves liberation of a portion of the pepsinogen molecule as a peptide. Pepsin

preferentially hydrolyzes peptide bonds containing an aromatic amino acid, and it

requires an acid medium to function.

The acid chyme is discharged from the stomach, containing partially degraded

proteins, into a slightly alkaline fluid in the small intestine. This fluid is composed of

pancreatic juice and succus entericus, the intestinal secretion. The pancreas secretes

three known proteinases, trypsin, chymotrypsin, and carboxypeptidase. All three are secreted as inactive zymogens. Activation starts with the transformation of the inactive

trypsinogen into the active trypsin. Trypsin, in turn, activates chymotrypsin and

carboxypeptidase.

Trypsin and chymotrypsin are endopeptidases; that is, they cleave internal peptide bonds. The so-called peptidases are exopeptidases; they cleave terminal peptide bonds. Trypsin has a

predilection for those containing the basic amino acid residues of lysine and arginine.

These two proteinases perform the major share in hydrolyzing proteins to small peptides.

Digestion to amino acids is completed by the exopeptidases. Carboxypeptidase acts on

peptides from the free carboxyl end; aminopeptidases from the free amino end.

Other peptidases act on di- or tripeptides, or peptides containing such special amino acids

as proline.

The amino acid digestion products of the proteins are absorbed by the small

intestine as rapidly as they are liberated. The absorbed amino acids are carried by the portal blood system to the liver, from which they are distributed to the rest of the body. Small amounts of the peptides formed during digestion escape further

hydrolysis and may also enter the circulation from the intestine. This is

shown by a rise in the peptide nitrogen in the blood.

Figure 25.17 The Postabsorptive State

Figure 25.17

PROTEIN IS

• A major component of foods. It is digested firstly in the stomach, and then in the duodenum to dipeptides and amino acid.

• Absorbed using symport active transport with sodium.

• Stored in liver and muscles.

Uses

• Protein synthesis : The synthesis of new proteins is very important during growth. In adults new protein synthesis is directed towards replacement of proteins as they are constantly turned over.

• Synthesis of a variety of other compounds : Examples of compounds synthesized from amino acids include purines and pyrimidines (components of nucleotides), catecholamines (adrenaline and noradrenalin) & neurotransmitters (serotonin)

Amino acid catabolism The other biological fuels discussed (carbohydrates & fats) contain only the

elements carbon, hydrogen and oxygen. Amino acids contain nitrogen as well. The first step in amino acid catabolism is the

removal of the nitrogen (the amino group).

Nitrogen removal from amino acidsNitrogen removal from amino acids

Transamination

Oxidativedeamination

Urea cycle

AminotransferasePLP

Transaminationit is a process of transferring amino groups from one molecule to another. There is no formation and no exceretion of ammonia, thusly no net change in the

nitrogen amount of body. It is a process involved in amino acids in which the amino group is transferred from the amino acid to a certain α-ketoacid with the consequant formation of a second α-ketoacid and amino acid. The

reaction is catalyzed by the enzyme aminotranferase (aka transaminase) which requires pyridoxal phosphate as a

prosthetic group. All transaminases contain this prosthetic group which derives from pyridoxine a water soluble

vitamin also known as vitamin B6. The amino group from amino acids is temporarily uptaken by the pyridoxal

phosphate as pyridoxamine phosphate prior to its donation to an α-ketoacid. All aminoacids except lysine, threonine, proline and hydroxyproline participate in transamination

process.

Deaminationit is a process of removing amino groups from one

molecule in order to reduce the amount of nitrogen of the body through ammonia synthesis and elimination. It is a process occurring in the liver during the metabolism of

amino acids. The amino group is removed from the amino acid and converted to ammonia-NH3 whose toxic activity is

canceled by conversion into urea which is eventually excreted. The glutamate dehydrogenase-GDH enzyme occupies a central role in nitrogen metabolism. Glutamate amino acid is cleaved into α-ketoglutarate and ammonia a

reaction catalyzed by GDH in a process called deamination. Glutamate is the only amino acid that

undergoes oxidative deamination at a relatively high rate. The formation of ammonia from the amino group thusly

occurs mainly via the amino group of glutamate.

Once the amino groups have all been "collected" in the form of the one amino acid, glutamate, this amino acid has its amino group removed (termed "oxidative deamination"). This reaction reforms alpha-ketoglutarate with the other product being

ammonia (NH4 +).

Ammonia is toxic to the nervous system and its accumulation rapidly causes death. Therefore it

must be detoxified to a form which can be readily removed from the body. Ammonia is converted to

urea, which is water soluble and is readily excreted via the kidneys in urine.

Unlike glucose, there is no storage form of amino acids.

Amino acids are degraded into free ammonia (NH4+) and the

carbon skeleton. Living organisms excrete excess

nitrogen as ammonia, uric acid, and urea.

ExcretoryExcretory forms of nitrogen forms of nitrogen

a) Excess NH4+ is excreted as ammonia (microbes,

aquatic vertebrates or larvae of amphibia),b) Urea (many terrestrial vertebrates)c) or uric acid (birds and terrestrial reptiles)

Nitrogen removal from amino acidsNitrogen removal from amino acids

Step 1: Remove amino group

Step 2: Take amino group to liver for nitrogen excretion

Step 3: Entry into mitochondria

Step 4: Prepare nitrogen to enter urea cycle

Step 5: Urea cycle

Step 1Step 1. Remove amino group. Remove amino group

• Transfer of the amino group of an amino acid to an α-keto acid ⇒ the original AA is converted to the corresponding α-keto acid and vice versa:

• Transamination is catalyzed by transaminases (aminotransferases) that require participation of pyridoxalphosphate:

amino acid

pyridoxalphosphate Schiff base

Step 2Step 2: Take amino group to liver for : Take amino group to liver for nitrogen excretionnitrogen excretion

Glutamatedehydrogenase

The glutamate dehydrogenase of mammalian liver has the unusual capacity to use either NAD+ or NADP+ as cofactor

Glutamate releases its amino group as ammonia in the liver.

The amino groups from many of the α-amino acids are collected in the liver in the form of the amino group of L-glutamate molecules.

1. Glutamatetransferres one amino group WITHIN cells:Aminotransferase → makes glutamate from α-ketogluta-rateGlutamate dehydrogenase → opposite

2. Glutamine transferres two amino group BETWEEN cells → releases its amino group in the liver

3. Alanine transferres amino group from tissue (muscle) into the liver

Nitrogen carriersNitrogen carriers

Move within cells

SynthAtase = ATP

In liver

Move between cells

Glucose-alanine cycle

Ala is the carrier of ammonia and of the carbon skeleton of pyruvate from muscle to liver.The ammonia is excreted and the pyruvate is used to produce glucose, which is returned to the muscle.

Alanine plays a special role in transporting amino groups to liver.

According to D. L. Nelson, M. M. Cox :LEHNINGER. PRINCIPLES OF BIOCHEMISTRY Fifth edition

Sources of ammonia for the urea cycle:Sources of ammonia for the urea cycle:

• Oxidative deamination of Glu, accumulated in the liver by the action of transaminases and glutaminase

• Glutaminase reaction releases NH3 that enters the urea cycle in the liver (in the kidney, it is excreted into the urine)

• Catabolism of Ser, Thr, and His (nonoxidative deamination) also releases ammonia:

Serine - threonine dehydratase

Serine →→ pyruvate + NH4+

Threonine →→ α-ketobutyrate + NH4+

• Bacteria in the gut also produce ammonia.

Review:

• Nitrogen carriers glutamate, glutamine, alanine• 2 enzymes outside liver, 2 enzymes inside liver:

– Aminotransferase (PLP) → α-ketoglutarate → glutamate

– Glutamate dehydrogenase (no PLP) → glutamate → α-ketoglutarate (in liver)

– Glutamine synthase → glutamate → glutamine

– Glutaminase → glutamine → glutamate (in liver)

Step 3: entry of nitrogen to mitochondria

Step 4: prepare nitrogen to enter urea cycle

Regulation

The urea cycle takes place in the mitochondria and the cytosol.

There are four enzymes involved, three of which are cytosolic and

one is mitochondrial.

Step 5: Urea cycleaspartate

Ornithine Ornithine transcarbamoylasetranscarbamoylase

Argininosuccinate Argininosuccinate synthasesynthase

Argininosuccinate Argininosuccinate lyaselyase

Arginase 1Arginase 1

OOA

Oxaloacetate → aspartate

Urea cycle – reviewUrea cycle – review ((Sequence of reactionsSequence of reactions))

• Carbamoyl phosphate formation in mitochondria is a prerequisite for the urea cycle– (Carbamoyl phosphate synthetase)

• Citrulline formation from carbamoyl phosphate and ornithine – (Ornithine transcarbamoylase)

• Aspartate provides the additional nitrogen to form argininosuccinate in cytosol– (Argininosuccinate synthase)

• Arginine and fumarate formation– (Argininosuccinate lyase)

• Hydrolysis of arginine to urea and ornithine– (Arginase)

The overall chemical balance of the The overall chemical balance of the biosynthesis of ureabiosynthesis of urea

NH3 + CO2 + 2ATP → carbamoyl phosphate + 2ADP + Pi

Carbamoyl phosphate + ornithine → citrulline + Pi

Citrulline + ATP + aspartate → argininosuccinate + AMP + PPi

Argininosuccinate → arginine + fumarate

Arginine → urea + ornithine

Sum: 2NH3 + CO2 + 3ATP urea + 2ADP + AMP + PPi + 2Pi

Nitrogen balanceNitrogen balance

Tissue proteins

Dietary proteins

Amino acidpool

Excretion as urea andNH4

+

Purines, heme, etc.Energy

The amount of nitrogen ingested is balanced by the excretion of an equivalent amount of nitrogen. About 80% of excreted nitrogen is in the form of urea.

Ammonia is rendered harmless in animals through the synthesis of urea (which in man, mammals, and several other animals forms in the liver and is

discharged with urine) or uric acid (in birds, reptiles, and insects) and is partially given off in the form of

ammonium salts. In plants and some bacteria, inorganic ammonium nitrogen may be reutilized, that is, used again in the synthesis of amino acids and amides and then of proteins. In these processes the amides of aspartic and glutamic acids play an important role, being the most important reserve compounds of nitrogen in plants. These compounds play an important role in animal organisms as well.

Urea is also found in a number of plants; its essential role in rendering ammonia harmless in fungi, bacteria, and higher plants has been established. In contrast to processes in animals, urea in

plants may be used again in the processes of protein synthesis when a sufficient quantity of carbohydrates is formed.

Thus, the principal difference between protein metabolism in animals and plants is that plants

synthesize protein, first forming amino acids and amides from inorganic substances, and

the ammonia that is formed in the deamination of amino acids is again used (through

glutamine, asparagine, and urea) in the resynthesis of protein.

Animals and man synthesize proteins from amino acids that are obtained from food and

that are partially formed as a result of transamination; the cleavage products of amino acids are discharged by the body.

Intermediate stages of protein metabolism in plants and animals have much in common.

The remainder of the amino acid is referred to as the "carbon skeleton". Depending on the particular amino acid being catabolised, its carbon

skeleton will be converted to : acetyl CoA.

Those carbon skeletons which end up as acetyl CoA are committed to energy

production. They will either be immediately oxidised via the citric acid

cycle or they may be converted to ketone bodies. Because the amino acids whose carbon skeletons yield

acetyl CoA are potentially a source of ketone bodies they are referred to as

ketogenic amino acids or pyruvate

or a citric acid cycle intermediate.

Glycine is the principal source for the formation of the pigmented grouping of

hemoglobin. The hormones of the thyroid gland (thyroxin and its derivatives) and of the

adrenal glands (epinephrine and norepinephrine) are formed from the amino

acid tyrosine. Tryptophan serves as the source for the formation of biogenic amines and also (in part) of nicotinic acid and its derivatives. A

number of other nitrogenous substances of the animal organism, such as glutathione,

carnosine, anserine, and creatine, are products of the union or transformation of amino acids. Alkaloids in plants are also formed from amino

acids.

Amino acid synthesisAmino acids are divided into two

classes depending on whether they can be synthesised in the human

body or whether they must be supplied in the diet. The former group are referred to as non-essential and

the latter group as essential. The table below shows which of the

twenty are in each group. Note that there are ten in each of the two

groups

Non-essential amino acids are synthesised from the products of their catabolism - i.e. acetyl CoA,

pyruvate or the relevant Krebs cycle intermediate. The amino group is donated by glutamate and added

by the reverse of the transamination discussed above. The essential amino acids are synthesised in micro-organisms (bacteria and yeasts) and passed

through the food chain until they reach us in our diet. One of the pathways essential to life which is carried

out by bacteria is the "fixation" of atmospheric nitrogen initially as inorganic nitrate and ultimately as

amino groups in amino acids. Higher organisms cannot perform this function.

Overview of amino acid catabolism in mammals

1.Biosynthesis

2.Urea cycle

Fumarate

Oxaloacetate