Amino acid catabolism- Part-1

Biochemistry For Medics- Lecture notes By- Professor(Dr.) Namrata Chhabra

www.namrata.co

Introduction

• Amino acid catabolism is part of the whole body catabolism

• Nitrogen enters the body in a variety of compounds present in the food, the most important being amino acids present in the dietary protein.

• Nitrogen leaves the body as urea, ammonia, and other products derived from amino acid metabolism

Protein turn over• The continuous degradation and synthesis of cellular proteins

occur in all forms of life. • Each day, humans turn over 1–2% of their total body protein,

principally muscle protein. • The rate of protein turn over varies widely for individual

proteins• Digestive enzymes and plasma proteins are rapidly degraded• High rates of protein degradation also occur in tissues

undergoing structural rearrangement—eg uterine tissue during pregnancy or skeletal muscle in starvation.

• Structural proteins, such as collagen, are metabolically stable and have half lives measured in months or years

Fate of amino acids

• Of the liberated amino acids, approximately 75% are reutilized.

• The remainder serve as precursors for important biological compounds

• Since excess amino acids are not stored, those not immediately incorporated into new protein or utilized else where are rapidly degraded to amphibolic intermediates.

Fate of amino acids

Body protein

Dietary protein

Synthesis of tissue protein

Synthesis of Biological compounds

Synthesis of glucose, ketone bodies or fatty

acids

Oxidation in TCA cycle to yield energy and

CO2

Amino acid degradation

• The major site of amino acid degradation in mammals is the liver.

• The amino group must be removed, as there are no nitrogenous compounds in energy-transduction pathways.

• The α-ketoacids that result from the deamination of amino acids are metabolized so that the carbon skeletons can enter the metabolic mainstream as precursors to glucose or citric acid cycle intermediates.

Fate of alpha amino group of amino acids

o The presence of the α- amino group keeps amino acids safely locked away from oxidative breakdown.

o Removal of α- amino group is an obligatory step in the catabolism of all amino acids

o Once removed this nitrogen can be incorporated in to other compounds or excreted, with the carbon skeleton being metabolized

o Different animals excrete excess nitrogen as ammonia, uric acid, or urea.

Fate of alpha amino group of amino acids

o The aqueous environment of teleostean fish, which are ammonotelic (excrete ammonia), compels them to excrete water continuously, facilitating excretion of highly toxic ammonia.

o Birds, which must conserve water and maintain low weight, are uricotelic and excrete uric acid as semisolid guano.

o Many land animals, including humans, are ureotelic and excrete nontoxic, water-soluble urea.

Biosynthesis of ureaUrea biosynthesis occurs in stages: (1) Transdeamination (Removal of α-amino group) by –a

coupled process of transamination and deaminationo Transamination forms Glutamate in peripheral cellso Deamination of glutamate forms ammonia in liver(2) Minor pathway of oxidative and non oxidative deamination

produce ammonia in peripheral cells(3) Ammonia transport-Ammonia is transported to liver as

glutamate, glutamine or alanine(4) Detoxification of released ammonia – Ammonia is

detoxified by specific reactions (Urea cycle) forming urea in liver.

Transamination• Transamination interconverts pairs of α-amino acids and α

-keto acids. • During Transamination, the amino group of an amino acid

(amino acid R 1) is transferred to a keto acid (keto acid R 2), this produces a new keto acid while from the original keto acid, a new amino acid is formed

Characteristics of Transamination

• The general process of transamination is reversible and is catalyzed by Transaminases, also called amino transferases that require B6-Phosphate as coenzyme.

• Most of the amino acids act as substrate for the transaminases but the amino acids like lysine, threonine, proline, and hydroxy proline do not participate in transamination reactions.

• Transamination is not restricted to α -amino groups.• The δ -amino group of ornithine and the Ʃ -amino

group of lysine—readily undergoes transamination.

Role of B6 Phosphate in transamination

• The coenzyme pyridoxal phosphate (PLP) is present at the catalytic site of aminotransferases and of many other enzymes that act on amino acids. PLP, a derivative of vitamin B6

• During transamination, bound PLP serves as a carrier of amino groups.

• Rearrangement forms an α-keto acid and enzyme-bound Pyridoxamine phosphate, which forms a Schiff base with a second keto acid

Role of B6 Phosphate in transamination

Role of B6 Phosphate in transamination

• The transfer of α-amino group from donor amino acid to Pyridoxal phosphate forms Pyridoxamine phosphate, and a keto acid.

• The α-amino group is finally passed on to acceptor α-keto acid to form a new amino acid.

Biological Significance of Transamination

• Transamination is used both for the catabolic as well as anabolic processes.

• The resultant α-Keto acid can be completely oxidized to provide energy, glucose, fats or ketone bodies depending upon the cellular requirement.

• Since it is a reversible process, it is also used for the synthesis of non essential amino acids.

• In addition to equilibrating amino groups among available α-keto acids, the process of transamination funnels amino groups from excess dietary amino acids to those amino acids (e.g., glutamate) that can be deaminated.

Clinical significance of Transaminases

Serum aminotransferases such as serum glutamate-oxaloacetate-aminotransferase (SGOT) (also called Aspartate aminotransferase, AST) and serum glutamate-pyruvate aminotransferase (SGPT) (also called alanine transaminase, ALT) have been used as clinical markers of tissue damage, with increasing serum levels indicating an increased extent of damage.

AST-Serum glutamate-oxaloacetate-aminotransferase (SGOT)

• AST is found in the liver, cardiac muscle, skeletal muscle, kidneys, brain, pancreas, lungs, leukocytes, and erythrocytes

• Normal serum activity is 0-41 IU/L. The concentration of the enzyme is very high in myocardium.

• The enzyme is both cytoplasmic as well as mitochondrial in nature

AST-Serum glutamate-oxaloacetate-aminotransferase (SGOT)

Reaction catalyzed can be represented as follows-

ALT- Serum glutamate Alanine transferase

• ALT is found primarily in the liver. • The normal serum activity ranges between 0-45

IU/L• Reaction catalyzed can be represented as follows-

Diagnostic significance of amino transferases

I) Liver Diseaseso The aminotransferases are normally present in the serum

in low concentrations.o These enzymes are released into the blood in greater

amounts when there is damage to the liver cell membrane resulting in increased permeability.

o These are sensitive indicators of liver cell injury and are most helpful in recognizing acute hepatocellular diseases such as hepatitis.

o Any type of liver cell injury can cause modest elevations in the serum aminotransferases

Diagnostic significance of amino transferases

2) Acute myocardial infarction- In acute MI the serum activity rises sharply within the first 12 hours, with a peak level of 24 hours or over and returns to normal within 3 to 5 days.

3) Extra cardiac and extra hepatic conditions• Elevation of AST can also be seen in Muscle disorders like

muscular dystrophies- myositis etc.• Increase activity of AST is also observed in acute

pancreatitis, leukemias and acute hemolytic anemias• In normal health slight rise of AST level can be observed

after prolonged exercise

Oxidative deamination of Glutamate

• The nitrogen atom that is transferred to α-ketoglutarate in the transamination reaction (forming Glutamate)is converted into free ammonium ion by oxidative deamination.

• This reaction is catalyzed by glutamate dehydrogenase. • This enzyme is unusual in being able to utilize either

NAD+ or NADP+• Glutamate dehydrogenase is located in mitochondria,

as are some of the other enzymes required for the production of urea.

Regulation of Glutamate dehydrogenase

o The activity of glutamate dehydrogenase is allosterically regulated.

o The enzyme consists of six identical subunits.o Guanosine triphosphate (GTP) and adenosine triphosphate

(ATP) are allosteric inhibitors, whereas Guanosine diphosphate (GDP) and adenosine diphosphate (ADP) are allosteric activators.

o Hence, a lowering of the energy charge (more of ADP and GDP) accelerates the oxidation of amino acids favoring formation of alpha keto glutarate that can be channeled towards TCA cycle for complete oxidation to provide energy.

Role of Glutamate

• Glutamate occupies a central place in the amino acid metabolism.

• Basically it acts as a collector of amino group of the amino acids.

• All the amino nitrogen from amino acids that undergo transamination can be concentrated in glutamate.

• L-glutamate is the only amino acid that undergoes oxidative deamination at an appreciable rate in mammalian tissues.

• The formation of ammonia from α -amino groups thus occurs mainly via the α -amino nitrogen of L-glutamate.

Role of Glutamate and Glutamate dehydrogenase

• Since in majority of the transamination reactions alpha keto glutarate is the acceptor keto acid forming Glutamate, that is oxidatively deaminated in the liver by Glutamate dehydrogenase to form alpha keto glutarate and ammonia.

• Conversion of α-amino nitrogen to ammonia by the concerted action of glutamate aminotransferase and GDH is often termed "transdeamination."

• Thus Transamination and deamination are coupled processes though they occur at distant places

Transdeamination

Minor pathways of deamination

A) Oxidative deamination• L-amino acid oxidases of liver and kidney

convert amino acids to an α -imino acid that decomposes to an α -keto acid with release of ammonium ion.

• The reduced flavin is reoxidized by molecular oxygen, forming hydrogen peroxide (H2O2), which then is split to O2 and H2O by Catalase.

Minor pathways of deamination

L- amino acid oxidases use FMN as a coenzyme whereas, FAD is used as a coenzyme by D-Amino acid oxidases.

Minor pathways of deamination

Non oxidative deaminationo Serine, threonine, cysteine/cystine and

Histidine undergo non oxidative deamination to form corresponding Alpha keto acids

o Deamination of Serine and Threonine is carried out by Dehydratase, whereas deamination of cysteine/cystine is carried out by desulfurase.

o Both the enzymes are B6 dependent.

Minor pathways of deamination

• Deamination takes place in two steps- First an imino acid is formed by the action of dehydratase/desulfurase.• The imino acid undergoes hydration and deamination to produce alpha keto acid.

oDeamination of cysteine and cystine is carried out in a similar wayoDeamination of histidine produces urocanic acid

Transport of Ammonia to the liver

• Two mechanism are available for the transport of ammonia from peripheral cells to liver for detoxification

• The first uses glutamine synthetase to combine glutamate with ammonia

• The second , used primarily by muscle, involves transamination of pyruvate to Alanine

Glutamate and Glutamine relationship

• Ammonia Nitrogen can be transported as glutamine.• This is the first line of defense in brain cells.• Glutamine synthetase catalyzes the synthesis of

glutamine from glutamate and NH4 + in an ATP-dependent reaction

• The nitrogen of glutamine can be converted to urea in liver by the action of glutaminase in liver

• Hydrolytic release of the amide nitrogen of glutamine as ammonia, catalyzed by glutaminase favors glutamate formation.

Glutamate and Glutamine relationship

The concerted action of glutamine synthase and glutaminase thus catalyzes the interconversion of free ammonium ion and glutamine

Glucose Alanine Cycle and role of Glutamate

• The transport of amino group of amino acids also takes place in the form of Alanine.

• Nitrogen is transported from muscle to the liver in two principal transport forms.

• Glutamate is formed by transamination reactions, but the nitrogen is then transferred to pyruvate to form alanine, which is released into the blood.

• The liver takes up the alanine and converts it back into pyruvate by transamination.

• The pyruvate can be used for gluconeogenesis and the amino group eventually appears as urea.

• This transport is referred to as the alanine cycle. It is reminiscent of the Cori cycle and again illustrates the ability of the muscle to shift some of its metabolic burden to the liver

Glucose Alanine cycle

Glutamate in muscle is transaminated to alanine, which is released into the blood stream. In the liver, alanine is taken up and converted into pyruvate for subsequent metabolism.

Ammonia intoxication• The ammonia produced by enteric bacteria and absorbed into

portal venous blood and the ammonia produced by tissues are rapidly removed from circulation by the liver and converted to urea.

• Thus, only traces (10–20 g/dL) normally are present in peripheral blood.

• This is essential, since ammonia is toxic to the central nervous system.

• Should portal blood bypass the liver, systemic blood ammonia levels may rise to toxic levels.

• This occurs in severely impaired hepatic function or the development of collateral links between the portal and systemic veins in cirrhosis.

Ammonia intoxication

• Excess of ammonia depletes glutamate and hence GABA level in brain

• To compensate for glutamate, alpha keto glutarate is used , the decrease concentration of which subsequently depresses TCA and thus deprives brain cells of energy.

• Excess Glutamine is exchanged with Tryptophan , a precursor of Serotonin , resulting in hyper excitation.

• Symptoms of ammonia intoxication include tremor, slurred speech, blurred vision, coma, and ultimately death.

Ammonia intoxication

Ammonia is highly toxic. Normally blood ammonium concentration is < 50 µmol /L, and an increase to only 100 µmol /L can lead to disturbance of consciousness. A blood ammonium concentration of 200 µmol /L is associated with coma and convulsions.

Renal glutaminase activity

• Excretion into urine of ammonia produced by renal tubular cells facilitates cation conservation and regulation of acid-base balance.

• Ammonia production from intracellular renal amino acids, especially glutamine, increases in metabolic acidosis and decreases in metabolic alkalosis.

Sources of Ammonia

• Transamination of amino acids with alpha ketoglutarate to form Glutamate and subsequent deamination of Glutamate by glutamate dehydrogenase (Transdeamination)

• Deamination of amino acids (oxidative and non oxidative)• From Glutamine by the action of Glutaminase• By the action of intestinal bacteria on the unabsorbed

dietary amino acids• From mono amines by the action of mono amine oxidase• From the catabolism of purines and pyrimidines

Fate of ammonia

• Biosynthesis of Glutamate from ammonia and alpha keto glutarate

• Biosynthesis of non essential amino acids through transdeamination

• Biosynthesis of Glutamine by the action of Glutamine synthetase

• Small amounts are excreted in urine• Major fate is formation of urea

Summary

Urea cycle

• To be continued …

• For further reading follow • Biochemistry for medics- Lecture notes• http://www.namrata.co/transamination-and-t

ransaminases/http://www.namrata.co/glutamate-central-role-in-amino-acid-metabolism/• Biochemistry for medics- clinical cases• http://usmle.biochemistryformedics.com/