aa pptt

Amino acid oxidationMetabolic fates of amino acidsTransamination,DeaminationDecarboxylationDeamidation Transdeamination

A. Sources and Uses of Amino Acids

Sources1.Proteins in the diet provide both essential and non-essential amino acids in contrast to microorganisms that for the most part synthesize their own.2.Turnover of endogenous proteins3.De novo biosynthesis(non-essential amino acids)Uses1.Protein synthesis2.Nitrogen and carbon source of general and special product biosynthesis3.Energy source a.glucogenic(those that can be used for the synthesis of glucose) b.ketogenic(those whose metabolism leads to ketonebodies)

Amino acid roles1) protein monomeric units (primary purpose)2) energy metabolites (about 10% of energy)3) precursors of many biologically important nitrogen containing compounds such as: a) HEME b) physiologically active AMINES [(nor)epinephrine, dopamine, GABA (g-aminobutyric acid), serotonin, histamine] c) GLUTATHIONE e) NUCLEOTIDES f) NUCLEOTIDES COENZYMES

Amino acid met related to nitrogen acquisition

Sources of nitrogen proteins & nucleic acids.

The major form of nitrogen in the atmosphere isN2, an extremely stable compound.

No animals are capable of either N-fixation or nitrate assimilation, So we are totally dependent on plants and microorganisms for the synthesis of organic nitrogenous compounds, such as amino acids and proteins, to provide this essential nutrient.

Essential Amino Acids in Humans

• Required in diet• Humans incapable of forming requisite

carbon skeleton• Arginine*• Histidine*• Isoleucine• Leucine• Valine

• Lysine• Methionine• Threonine• Phenylalanine• Tryptophan

* Essential in children, not in adults

Non-Essential Amino Acids in Humans

• Not required in diet• Can be formed from a-keto acids by

transamination and subsequent reactions• Alanine

• Asparagine• Aspartate• Glutamate• Glutamine

• Glycine• Proline• Serine• Cysteine (from Met*)• Tyrosine (from Phe*)

* Essential amino acids

Functions of Proteins

• Functions: Structural

Catalytic

Transport action

Signaling and hormonal functions

Source of energy (16.7kJ/g)

Nutritional Quality of Proteins

• Non-essential amino acids

synthesized in the body

synthesized by the transamination of a-keto acids

Tyrosine and cysteine

synthesized in the body by using essential amino acids

from phenylalanine and methionine respectively

semi-essential

Proteins in the Body

Proteins provide: • Amino acids for

protein synthesis.• Nitrogen atoms

for nitrogen-containing compounds.

• Energy when carbohydrate and lipid resources are not available.

Negative Nitrogen Balance

1. Stress

2. Decreased Intake

3. Lack of an essential AA

Proteins degradation Ubiquitine-proteasome dependent

require ATP (energy dep)

Dynamics of Protein And Amino Acid Metabolism

Dietary Proteins Digestion to Amino Acids

Transport in Blood to Cells

Protein Synthesis Functional Proteins

Protein Degradation In Proteasomes Following Tagging With Ubiquitin

Amino Acids

Metabolites

OVERVIEW OF AMINO ACID METABOLISM

ENVIRONMENT ORGANISM

Ingested protein

Bio- synthesis Protein

AMINO ACIDS

Nitrogen Carbon

skeletons

Urea

Degradation (required)

1 2 3

a

b

PurinesPyrimidinesPorphyrins

c c

Used for energy

pyruvateα-ketoglutaratesuccinyl-CoAfumarateoxaloacetate

acetoacetateacetyl CoA

(glucogenic)(ketogenic)

Major Functions of Amino Acids Derived from Dietary Protein

OxidationGlycogenic amino acids: --Blood glucose--EnergyKetogenic amino acids: -Acetyl CoA-Stored fat-Energy

Biosynthesis of nitrogen-containing metabolites

Heme Blood cell

Choline Phospho lipid

Glycosamine Sugar

Nucleotides DNA

Protein synthesis Protein

Biogenic amines Neurotransmitter

Carnitine Heart

Creatine phosphate « Energy »

Protein Metabolisma) α keto acids are funneled into the Krebs cycle

(glucogenic/ketogenic)

b) NH4+ is cleared via urea, NH4

+, with uric acid however major product is urea (80%)

c) Creatine (a break-down product of creatine phosphate in

muscle)/creatinine (supply energy to all cells in the body, primarily muscle)

Digestion of ProteinsStomach: Pepsinogen Pepsin (max. act. pH 2)

Small Intestine: Trypsinogen Trypsin

Trypsin cleaves:Chymotrypsinogen to chymotrypsinProelastase to elastaseProcarboxypeptidase to carboxypeptidase

Aminopeptidases (from intestinal epithelia)

Enteropeptidase

Enzymatic digestion of dietary proteins in gastro-intestinal-tract.

Table 1. Phases of Digestion and Absorption of Protein and its Degradative Products

Phase of Digestion

Location Agents Outcome

5. Cleavage of di-/tripeptides transport to capillaries

epithelial cell – cytoplasm contraluminal membrane

dipeptidasestripeptidasesfacilitated diffusion

free amino acids from di/tripeptides;amino acids transported into capillaries

4. Absorption intestinal epithelial cell brush border membrane

transport systems uptake into epithelial cell

3. Brush Border Surface

brush border surface of intestine

endopeptidases and aminopeptidases

free amino acids anddi/ tripeptides

2. Pancreatic Proteases

lumen of small Intestine

trypsin, chymotrypsin,elastase, and carboxypeptidases

free amino acids andoligopeptides – 2 to 8 amino acids

1. Gastric Digestion

stomach stomach acid pepsin

denaturationlarge peptide fragments + some free amino acids

Amino acid metabolism

• Metabolism of amino acids differs, but 3

common reactions:

– Transamination

– Deamination

– Formation of urea

Amino Acid Catabolism

• Deamination of Amino Acids

Oxidative Deamination

Non-oxidative Deamination• Transamination• Deamidation

3. DEAMIDATION

• Glutamic acid and aspartic acid also occur as their corresponding amides i.e glutamine and asparagine

• These can be hydrolyzed by glutaminase and asparaginase to their respective amino acid and ammonia is released in these reactions.

1. DEAMINATION: Oxidative Deamination

• Only a few amino acids can be deaminated directly. Glutamate Dehydrogenase catalyzes a major reaction that effects net removal of N from the amino acid pool .  Glutamate Dehydrogenase is one of the few enzymes that can utilize either NAD+ or NADP+ as electron acceptor. Oxidation at the a-carbon is followed by hydrolysis, releasing NH4

+.

Non-oxidative Deamination

Serine Dehydratase catalyzes: serine à pyruvate + NH4

2.Transamination

Transaminase enzymes (aminotransferases) catalyze the reversible transfer of an amino group between two a-keto acids.

• Transaminases equilibrate amino groups among available a-keto acids. This permits synthesis of non-essential amino acids, using amino groups derived from other amino acids and carbon skeletons synthesized in the cell. Thus a balance of different amino acids is maintained, as proteins of varied amino acid contents are synthesized

• Transaminases or aminotransferases require pyridoxal-5’-phophate PLP (vitamin B6 derivative) donate amino group to keto acid.

Excretory forms of Nitrogen

Metabolic Classification of the Amino Acids

• Essential and Non-essential

• Glucogenic and Ketogenic

Glucogenic Amino Acids• Metabolized to a-ketoglutarate, pyruvate,

oxaloacetate, fumarate, or succinyl CoAPhosphoenolpyruvate Glucose

• Aspartate• Asparagine• Arginine• Phenylalanine• Tyrosine• Isoleucine

• Methionine• Valine• Glutamine• Glutamate• Proline• Histidine

• Alanine• Serine• Cysteine• Glycine• Threonine• Tryptophan

Ketogenic Amino Acids• Metabolized to acetyl CoA or acetoacetyl

CoA

Animals cannot convert acetyl CoA or acetoacetyl CoA to pyruvate

• Isoleucine• Leucine *• Lysine *• Threonine

• Tryptophan• Phenylalanine• Tyrosine

* Leucine and lysine are only ketogenic

Ammonia intoxication, nitrogen excretion and urea formation, urea cycle

and its regulation, genetic defects of urea cycle

Formation of Urea (Urea Cycle)

•

Urea Cycle

• The urea cycle takes place partly in the cytosol and partly in the mitochondria

Urea Cycle

The urea cycle• Detoxifies ammonium ion from amino

acid degradation.• Converts ammonium ion to urea in the

liver. O ||

H2N—C—NH2 urea

• Provides 25-30 g urea daily for urine formation in the kidneys.

Carbamoyl phosphate synthase-I Reaction

Ammonia released from the oxidative deamination is incorporated in carbamoyl phosphate by using ATP and bicarbonate.

N-acetyl glutamine is a positive regulator of this enzyme.

Carbamoyl phosphate enters the urea cycle in the mitochondria.

Carbamoyl Phosphate

• In the mitochondria, an ammonium ion reacts with CO2 from the citric acid cycle, 2 ATP, and water.

NH4+ + CO2 + 2ATP + H2O

O O || ||

H2N—C—O—P—O- + 2ADP + Pi

| O-

Carbamoyl phosphate

Reaction 1 Transfer of Carbamoyl Group

• The carbamoyl group is transferred to ornithine to form citrulline.

• Citrulline moves across the mitochondrial membrane into the cytosol.

Reaction 2 Condensation with Aspartate

• In the cytosol, citrulline combines with aspartate.

• Hydrolysis of ATP to AMP provides energy.

• The N in aspartate is part of urea.

Cytosol

Reaction 3 Cleavage of Fumarate

Fumarate• Is cleaved from argininosuccinate. • Enters the citric acid cycle.

Reaction 4 Hydrolysis Forms Urea

Hydrolysis of arginine:

• Forms urea.• Forms ornithine,

which returns to the mitochondrion to pick up another carbamoyl group to repeat the urea cycle.

Summary of Urea Cycle

The urea cycle converts:• Ammonium ion to urea• Aspartate to Fumarate• 3ATP to 2ADP, AMP, 4Pi

NH4+ + CO2 + 3ATP + Aspartate + 2H2O

Urea + 2ADP + AMP + 4Pi + Fumarate

Blood Urea Nitrogen

• Normal range: 7-18 mg/dL• Elevated in amino acid catabolism• Glutamate N-acetylglutamate

CPS-1 activation• Elevated in renal sufficiency• Decreased in hepatic failure

Interaction of Urea Cycle and Citric Acid Cycle via Aspartate-Argininosuccinate shunt

Regulation of urea cycle

1. Enzymes involved in urea cycle are synthesized at higher level when proteins are utilized for energy production (starvation, or availability of fat and carbohydrate-free diet.

2. The carbamoyl phosphate synthase is allosterically activated by N-acetylglutamate.

Hereditary deficiency of any of the Urea Cycle enzymes leads to hyperammonemia - elevated [ammonia] in blood.

Total lack of any Urea Cycle enzyme is lethal.

Elevated ammonia is toxic, especially to the brain.

If not treated immediately after birth, severe mental retardation results.

1. High [NH3] would drive Glutamine Synthase:

glutamate + ATP + NH3 glutamine + ADP + Pi

This would deplete glutamate – a neurotransmitter & precursor for synthesis of the neurotransmitter GABA.

2. Depletion of glutamate & high ammonia level would drive Glutamate Dehydrogenase reaction to reverse:

glutamate + NAD(P)+ a-ketoglutarate + NAD(P)H + NH4

+

The resulting depletion of a-ketoglutarate, an essential Krebs Cycle intermediate, could impair energy metabolism in the brain.

Postulated mechanisms for toxicity of high [ammonia]

GABA Formation

NH3+

-O2CCH2CH2CHCO2-

NH3+

-O2CCH2CH2CH2

Glutamate Gamma-aminobutyrate(GABA)

• GABA is an important neurotransmitter in the brain• Drugs (e.g., benzodiazepines) that enhance the effects of

GABA are useful in treating epilepsy

Glutamatedecarboxylase

CO2

• During prolonged starvation, breakdown of muscle protein supply most metabolic energy, urea prod increases.

• Regulation is via synthesis of four urea cycle enzymes and carbamoyl phosphate synthetase I in liver.

• All five enzymes synth increases in starving and on very-high-protein diets animals in comparison to animals eating carbohydrates & fats.

• Animals on protein-free diets produce lower levels of urea cycle enzymes.

• Carbamoyl phosphate synthetase I (synthesized from acetyl-CoA & glutamate by N-acetylglutamate synthase), is allosterically activated by N-acetylglutamate

Regulation of Urea cycle

• In plants and microorganisms N- acetylglutamate synthase catalyzes the first step in synthesis of arginine from glutamate.

The absence of a urea cycle enzyme can result in hyperammonemia or in the build-up of one or more urea cycle intermediates, depending on the enzyme that is missing

Most urea cycle steps are irreversible, absent enzyme activity can often be identified by intermediate present in elevated concentration in the blood/or urine.

limiting protein intake (amount adequate to supply amino acids for growth), & adding a-keto acid to diet analogs of essential amino acids.

Liver transplantation has also been used, since liver is the organ that carries out Urea Cycle.

Treatment of deficiency of Urea Cycle enzymes

• Treatment for deficiencies in urea cycle enzymes.

• The aromatic acids benzoate & phenylbutyrate, combine with glycine & glutamine. Products are excreted in the urine.

• Subsequent synthesis of glycine & glutamine removes ammonia from bloodstream

Treatment for def in urea cycle enzymes

Deficiency of N-acetylglutamate synthase results in the absence of the normal activator of carbamoyl phosphate synthetase I.This condition can be treated by administering carbamoyl glutamate, (analog of N-acetylglutamate) that is effective in activating carbamoyl phosphate synthetase I.Supplementing the diet with arginine is useful in treating def of ornithine transcarbamoylase, argininosuccinate synthetase, and argininosuccinase.

• The complete Urea Cycle is significantly only in liver. However some enzymes of the pathway are in other cells and tissues where they generate arginine & ornithine, which are precursors for other important molecules.

• e.g., Argininosuccinate Synthase, which catalyzes synthesis of the precursor to arginine, is in most tissues.

• Mitochondrial Arginase II, distinct from the cytosolic urea cycle Arginase, cleaves arginine to yield ornithine.

cytosol mitochondrial matrix

carbamoyl phosphate Pi

ornithine citrulline

ornithine citrulline urea aspartate

arginine argininosuccinate fumarate

The amino acid arginine, in addition to being a constituent of proteins and an intermediate of the Urea Cycle, is precursor for synthesis of creatine & the signal molecule nitric oxide.

H3N+ C COO

CH2

CH2

CH2

NH

C

NH2

NH2

H

arginine (Arg)

H2N C N

NH2+

CH2

CH3

C

O

O

creatine

Functions, pathways of amino acid degradation and genetic

disorders of

individual amino acids

Glucose-alanine cycle. Alanine serves as a carrier of ammonia and of the carbon skeleton of pyruvate from skeletal muscle to liver. The ammonia is excreted and the pyruvate is used to produce glucose, which is returned to the muscle.

Pathways of Amino Acid Degradation

• Ammonia transport in the form of glutamine.

• Excess ammonia in tissues is added to glutamate to form glutamine, catalyzed by glutamine synthetase.

• After transport in the bloodstream, glutamine enters liver and NH4 is liberated in mitochondria by glutaminase.

Vitamin-Coenzymes in Amino Acid Metabolism

• Cofactors transfer one-carbon groups in different oxidation states

• Vitamin B-6 (Pyridoxal phosphate) PLP• Folic acid (Tetrahydrofolate). • Vitamin B-12 • Biotin• S-adenosylmethionine (SAM)

Vitamin-Coenzymes in Amino Acid Metabolism

• Vitamin B-6 : pyridoxal phosphate– Enzymes that bind amino

acids use PLP as coenzyme for binding• Transaminases• Amino acid decarboxylases• Amino acid deaminases

Vitamin-Coenzymes in Amino Acid Metabolism

• Folacin: Tetrahydrofolate (THF)– Carrier of single

carbons• Donor & receptor• Glycine and serine• Tryptophan degradation• Histidine degradation• Purine and pyrimidine

synthesis

Vitamin-Coenzymes in Amino Acid Metabolism

• Vitamin B-12– Catabolism of BCAA

• Methyl-malonyl CoA mutase (25-9 &10)

• Glycine is degraded via three pathways, only one of which leads to Pyruvate.

• Glycine is converted to serine by enzymatic addition of a hydroxymethyl group.

• This reaction, catalyzed by serine hydroxymethyl transferase, requires the coenzymes tetrahydrofolate and pyridoxal phosphate

Glycine

Catabolic pathways for alanine, glycine,serine, cysteine, tryptophan, and threonine.

Catabolic pathways for tryptophan, lysine, phenylalanine,tyrosine, leucine & isoleucine. These amino acids donate some of their carbons (red) to acetyl-CoA. Tryptophan, PAL,Tyrosine, isoleucine also contribute carbons (blue) to pyruvate orTCA cycle interm. The nitrogen atoms are transferred to -ketoglutarate to form glutamate

Tryptophan as precursor. The aromatic rings of tryptophangive rise to nicotinate, indoleacetate, and serotonin.

Phenylalanine Catabolism Is Genetically Defective in Some People

• Amino acids are either neurotransmitters or precursors or antagonists of neutrotransmitters.

• Phenylketonuria (PKU) elevated levels of phenylalanine (hyperphenylalaninemia): Phenylalanine hydroxylase (first enzyme in the catabolic pathway for phenylalanine.

• Alkaptonuria, urine may turn brown if collected/exposed to open air, Kidney stones, due to the accumulation of homogentisic acid in tissues. Ear wax exposed to air turns red or black bec of the accumulation of homogentisic acid (toxic tyrosine byproduct, or alkapton).

• Tyrosinemia is a genetic disorder characterized by elevated blood levels of the amino acid tyrosine

• Type I tyrosinemia, most severe caused by a shortage of enzyme fumarylacetoacetate hydrolase. Symptoms include failure to gain weight, diarrhea, vomiting, yellowing of the skin & whites of the eyes (jaundice), increased tendency to bleed (particularly nosebleeds). Lead to liver and kidney failure, problems nervous system, liver cancer.

• Type II tyrosinemia caused by a deficiency of enzyme tyrosine aminotransferase. Symptoms affect eyes, skin, mental development. excessive tearing, abnormal sensitivity to light (photophobia), eye pain and redness, and painful skin lesions on the palms and soles, intellectual disability.

• Type III tyrosinemia is rare disorder caused by a deficiency of the enzyme 4-hydroxyphenylpyruvate dioxygenase. Characteristic features include intellectual disability, seizures, and periodic loss of balance and coordination.

Genetic defects in many of these enzymes cause inheritable human diseases

Alternative pathways for catabolism ofphenylalanine in phenyl ketonuria.In PKU, phenyl Pyruvate accumulates in the tissues, blood, and urine. The urine may also contain phenyl acetate & phenyl lactate.

Catabolic pathways for arginine, histidine, glutamate, glutamine, and proline. These amino acids are converted to -ketoglutarate. The numbered steps in the histidine pathway are catalyzed by 1 histidine ammonia lyase, 2 urocanate hydratase, 3 imidazolonepropionase, 4 glutamate formimino transferase

Catabolic pathways for methionine, isoleucine, threonine & Valine. These amino acids are converted to succinyl-CoA; isoleucine also contributes two of its carbon atoms to acetyl-CoA

• Much of the catabolism of amino acids takes place in the liver, the three amino acids with branched side chains (leucine, isoleucine, and valine) are oxidized as fuels primarily in muscle, adipose, kidney, and brain.

• These extrahepatic tissues contain an aminotransferase, absent in liver, that acts on all three branched-chain amino acids to produce the corresponding -keto acids.

• Branched-chain -keto acid dehydrogenase complex then catalyzes oxidative decarboxylation of all three -keto acids, releasing carboxyl group as CO2 & producing acyl-CoA derivative.

Five cofactors (thiamine pyrophosphate, FAD, NAD, lipoate, and coenzyme A) participate, and the three proteins in each complex catalyze homologous reactions.

Branched-Chain Amino Acids Are Not Degraded in the Liver

Catabolic pathways for the three branched-chain amino acids: valine, isoleucine, & leucine. Three pathways, in extrahepatic tissues. MSUD, (branched-chain ketoaciduria), autosomal recessive metabolic disorder affecting branched-chain amino acid metabolism.

Catabolic pathway for asparagine and aspartate. Bothamino acids are converted to oxaloacetate.

AMINO ACID BIOSYNTHESIS• ALL ARE SYNTHESIZED FROM COMMON METABOLIC

INTERMEDIATES• NON-ESSENTIAL

– TRANSAMINATION OF -KETOACIDS THAT ARE AVAILABLE AS COMMON INTERMEDIATES

• ESSENTIAL – THEIR -KETOACIDS ARE NOT COMMON INTERMEDIATES

(ENZYMES NEEDED TO FORM THEM ARE LACKING)• SO TRANSAMINATION ISN’T AN OPTION

– BUT THEY ARE PRESENT IN COMMON PATHWAYS OF MICRO-ORGANISMS AND PLANTS

AMINO ACID BIOSYNTHESIS OVERVIEW(USE OF COMMON INTERMEDIATES)

GLUCOSE GLUC-6-PHOSPHATE RIB-5-PHOS→ HIS 3-PHOSPHOGLYCERATE SERINE

GLYCINE E-4-PHOS + PEP CYSTEINE

PHE→TYR PYRUVATE ALA TRP VAL

CITRATE LEU, ILE ↓

OXALOACETATE, -KETOGLUTARATE ASP, ASN, GLU, GLN, PRO, ARG, LYS, THR, MET

SYNTHESIS OF NON-ESSENTIAL AMINO ACIDS

• ALL (EXCEPT TYR) SYNTHESIZED FROM COMMON INTERMEDIATES SYNTHESIZED IN CELL

– PYRUVATE– OXALOACETATE– -KETOGLUTARATE– 3-PHOSPHOGLYCERATE

SYNTHESIS OF NON-ESSENTIAL AMINO ACIDS

• TRANSAMINATION REACTIONS: ONE STEP

• PYRUVATE + AA ALANINE + -KETOACID• OXALOACETATE + AA ASPARTATE + -KETOACID• -KETOGLUTARATE + AA GLUTAMATE + -

KETOACID

• TRANSAMINASES: EQUILIBRATE AMINO GROUPSREQUIRE PYRIDOXAL PHOSPHATE (PLP)

• ALL AAs, EXCEPT LYS, CAN BE TRANSAMINATED

SYNTHESIS OF NONESSENTIAL AMINO ACIDS

• ATP-DEPENDENT AMIDATION OF ASP, GLU– ASN, GLN– GLU + ATP + NH3 GLN + ADP + Pi

• GLUTAMINE SYNTHETASE• NH3 IS TOXIC; IT’S STORED AS GLN

• GLN DONATES AMINO GPS IN MANY REACTIONS– ASP + ATP + GLN ASN + AMP + PPi + GLU

• ASPARAGINE SYNTHETASE

SYNTHESIS OF NONESSENTIAL AMINO ACIDS

NITROGEN METABOLISM IS CONTROLLED BY REGULATION OF GLUTAMINE SYNTHETASE

IN MAMMALS, GLN SYNTHETASES ACTIVATED BY -KG EXCESS AAs TRANSAMINATED TO GLU

OXIDATIVE DEAMINATION OF GLU -KG + NH3

NH3 UREA OR GLN (STORAGE)

-KG IS A SIGNAL THAT ACTIVATES GLN SYNTHETASE

NONESSENTIAL AMINO ACID SYNTHESIS

• CYSTEINE– SER + HOMOCYSTEINE CYSTATHIONINE

• HOMOCYSTEINE IS A BREAKDOWN PRODUCT OF METHIONINE

– CYSTATHIONINE -KETOBUTYRATE + CYS

• NOTE: -SH GROUP COMES FROM MET– SO CYS IS ACTUALLY AN ESSENTIAL AMINO ACID

NONESSENTIAL AMINO ACID SYNTHESIS

SUMMARY POINT:

ALL NONESSENTIALS (EXCEPT TYR) ARE DERIVED FROM ONE OF THE FOLLOWING COMMON INTERMEDIATES:

PYRUVATEOXALOACETATE-KG3-PHOSPHOGLYCERATE

Conversion of Serine to Glycine

N

N

N

NH2N

OH

CH2NHR

H

H

NH

N CH2

NH2C

Folate

Tetrahydrofolate (FH4)

Dihydrofolate reductase

N5, N10-Methylene FH4

NH3+H

CH2OH

CO 2-

C Serine

NH3+H

H

CO 2-

CGlycine

Serine hydroxymethyltransferase (PLP-dep.)

Key intermediatein biosynthesis ofpurines andformation ofthymine Important in

biosynthesis of heme,porphyrins, and purines

Sulfur-Containing Amino Acids

NH3+

CH3SCH2CH2CHCO 2-

NH3+

HSCH2CH2CHCO 2-

NH3+

CH2CHCO2-

NH3+

SCH2CH2CHCO 2-NH3

+

HSCH2CHCO 2-

OH

CH3CHCH 2CO 2-

Methionine(Essential)

L-Homocysteine

MethionineSynthase(Vit. B12-dep.)

+ FH4

+ 5-Methyl FH4

NH3+H

CH2OH

CO 2-

C Serine

Cystathionine

Cystathionineb-synthase(PLP-dep.)

Cystathioninelyase

Cysteine(Non-essential)

+

b-Hydroxy-butyrate

Homocysteine

Homocysteinuria• Rare; deficiency of cystathionine b-synthase• Dislocated optical lenses• Mental retardation• Osteoporosis• Cardiovascular disease death

High blood levels of homocysteine associated withcardiovascular disease

• May be related to dietary folate deficiency• Folate enhances conversion of homocysteine to

methionine

Methionine Metabolism: Methyl Donation

N

N N

N

O

OHOH

-O2CCHCH2CH2-S-H2C

NH2

NH3+ CH3

+NH3

+

CH3SCH2CH2CHCO 2-

N

N N

N

O

OHOH

-O2CCHCH2CH2-S-H2C

NH2

NH3+

N

N N

N

O

OHOH

H3NCH2CH2CH2-S-H2C

NH2

CH3

+

S-Adenosyl methioninesynthase

ATP

S-Adenosyl Methionine(SAM)

S-Adenosyl homocysteine

Methyl-transferases

Decarboxylated SAM

SAM Decarboxylase

CO2

Methionine

R-H

R-CH3

+

Polyamine Biosynthesis

NH3+

H3NCH2CH2CH2CHCO2-

+

H3NH

H

HN

H

N NH3

++++

H3NNH3

++

Ornithine(from urea cycle)

Putrescine

CO2

Ornithinedecarboxylase(ODC)(PLP-dep.)

DecarboxylatedSAM

Spermidine synthase

5’-Methylthio-adenosine

H3NNH

NH3

H

+++

Spermidine

Spermine

DecarboxylatedSAM

Spermine synthase

5’-Methylthio-adenosine

Polyamines

• Spermidine and spermine found in virtually all procaryotic and eucaryotic cells

• Precise role undefined• Bind to nucleic acids

• Inhibition of biosynthetic pathway:

H2NNH2

CO2H

CHF2

a-Difluoromethyl-ornithine (DFMO)(Eflornithine) - inhibits ODC;used to treatPneumocystis carinii infectons

Creatine and Creatinine

NH3+NH2

+H2N=C-HNCH2CH2CH2CHCO 2

-

Arginine Glycine Ornithine

Arginine-glycinetransamidinase

(Kidney)NH2

H2N=C-HNCH2CO 2-

+

Guanidoacetate

NHPO3-2

CH3

+H2N=C-NCH2CO 2

-

GuanidoacetateMethyltransferase

(Liver)

SAM + ATP

S-Adenosyl-homocysteine + ADP

Phosphocreatine

N

NH

CH3

HN

O

Creatinine(Urine) Non-enzymatic

(Muscle)

NH2

CH3

H2N=C-NCH2CO 2-

+

Creatine kinase(Muscle)

ATP

Creatine ADP + Pi

Creatine and Creatinine Creatine:

• Dietary supplement• Used to improve athletic performance

Creatinine:• Urinary excretion generally constant;

proportional to muscle mass

Creatinine Clearance Test:• Compares the level of creatinine in urine (24 hrs.)

with the creatinine level in the blood• Used to assess kidney function• Important determinant in dosing of several drugs

in patients with impaired renal function

Histidine Metabolism: Histamine Formation

N

NH

CH2CHCO2-

NH3

+

N

NH

CH2CH2NH2

Histidine Histamine

Histidinedecarboxylase

CO2

Histamine:• Synthesized in and released by mast cells• Mediator of allergic response: vasodilation, bronchoconstriction

(H1 receptors)• H1 blockers: Diphenhydramine (Benadryl)

Loratidine (Claritin)• Stimulates secretion of gastric acid (H2 receptors)

• H2 blockers: Cimetidine (Tagamet); ranitidine (Zantac)

Phenylalanine and Tyrosine

CH2CHCO2-

NH3+

CH2CHCO2-

NH3+

HO

HN

N

NH

NH

H2N

O

H

H

CHCHCH3

HO OH

HN

N

NH

NH2N

O

CHCHCH3

HO OH

Phenylalanine(Essential)

Tyrosine(Non-essential)

Phenylalanine-4-Monooxygenase(Phenylalaninehydroxylase)

O2

H2O

+

+

NADPH + H+

NADP+

Tetrahydrobiopterin (BH4)

Dihydrobiopterin

Phenylketonuria (PKU) Disease• Deficiency of Phe hydroxylase• Occurs in 1:20,000 live births in U.S.• Seizures, mental retardation, brain

damage• Treatment: limit phenylalanine intake• Screening of all newborns mandated

in all states

CH2CCO2-

O

Phe

Tyr

Transamination

Phenylpyruvate(urine)

Catecholamine Biosynthesis

CH2CHCO2-

NH3+

HO

CH2CHCO2-

NH3+

HO

HO

CH2CH2NH2

HO

HO

CHCH2NH2

HO

HO

OH

CHCH2NHCH3

HO

HO

OH

Tyr hydroxylase

O2

Tyrosine Dihydroxyphenylalanine (DOPA)

Dopamine

DOPAdecarboxylase CO2

Dopaminehydroxylase

Norepinephrine

Catechol

Epinephrine(Adrenaline)

SAM

S-Adenosyl-homocysteine

Methyl transferase

DOPA, dopamine, norepinephrine,and epinephrine are all neurotransmitters

L-DOPA in Parkinsonism

Blood Brain

Blood Brain Barrier

L-DOPA L-DOPA Dopamine

Dopamine

HO

HO CH2-C-CO2H

CH3

NHNH2Carbidopa

Blocks

Parkinsonism associated with dopamine in brain through loss ofneurons in basal ganglia.Carbidopa + L-DOPA

Homogentisic Acid Formation

CH2CHCO2-

NH3+

HO

OH

OH

CH2CO2-

Transamination

Tyrosine p-Hydroxyphenyl-pyruvate

Homogentisate

p-Hydroxyphenyl-pyruvatedioxygenase(ascorbate-dep.)

O2

CO2

CH2CCO2-

O

HO

Homogentisatedioxygenase

O2

Cleavage of aromatic ring

Fumarate + acetoacetate

Deficient in alkaptonuria

Alkaptonuria• Deficiency of homogentisate dioxygenase

• Urine turns dark on standing• Oxidation of homogentisic acid

• Asymptomatic in childhood

• Tendency toward arthritis in adulthood

Melanin Formation

CH2CHCO2-

NH3

O

O

+

CH2CHCO2-

NH3+

HO

HO

Highly colored polymeric

intermediates

Melanin(Black polymer)

Tyr hydroxylase

DOPA

Dopaquinone

CH2CHCO2-

NH3+

HO

Tyrosine

Tyrosinase

Melanin formed in skin (melanocytes), eyes, and hairIn skin, protects against sunlightAlbinism: genetic deficiency of tyrosinase

O2

Tryptophan Metabolism: Serotonin Formation

NH

CH2CHCO2-

NH3

+

NH

CH2CHCO2-

NH3

HO

+

NH

CH2CH2NH2

HO

Tryptophan(Trp)

Indole ring

Trphydroxylase

O2

5-Hydroxy-tryptophan

Decarboxylase

CO2 5-Hydroxy-tryptamine (5-HT);Serotonin

Serotonin• Serotonin formed in:

• Brain (neurotransmitter; regulation of sleep, mood, appetite) • Platelets (platelet aggregation, vasoconstriction)• Smooth muscle (contraction) • Gastrointestinal tract (enterochromaffin cells - major storage site)

• Drugs affecting serotonin actions used to treat: • Depression

• Serotonin-selective reuptake inhibitors (SSRI) • Migraine• Schizophrenia• Obsessive-compulsive disorders • Chemotherapy-induced emesis

• Some hallucinogens (e.g., LSD) act as serotonin agonists

• Food supplement promoted for serotonin effects• L-Tryptophan disaster (1989):

• Eosinophilia-myalgia syndrome (EMS) • Severe muscle and joint pain • Weakness • Swelling of the arms and legs • Fever• Skin rash • Eosinophilia• Many hundreds of cases; several deaths• Traced to impurities

L-Tryptophan

Serotonin Metabolism: 5-HIAA

NH

CH2CH2NH2

HO

NH

CH2CHO

HO

NH

CH2CO2H

HO

Serotonin

MAO

Dehydrogenase

5-Hydroxyindole acetic acid (5-HIAA) (Urine)

Carcinoid tumors: • Malignant GI tumor type• Excretion of large amounts of 5-HIAA

Serotonin Metabolism: Melatonin

NH

CH2CH2NHCOCH3

H3CO

NH

CH2CH2NH2

HO2 Steps

Serotonin Melatonin

Melatonin:• Formed principally in pineal gland• Synthesis controlled by light, among other factors• Induces skin lightening• Suppresses ovarian function• Possible use in sleep disorders

Tryptophan Metabolism:Biosynthesis of Nicotinic Acid

NH

CH2CHCO2-

NH3

+

Tryptophan

N

CO2H

Nicotinic acid (Niacin)

Several steps

Nicotinamide adenine dinucleotide (NAD)

Transport of amino group, role of pyridoxal phosphate, glutamate, glutamine, alanine

Specialized Amino Acid Roles1. Certain NEAA continue being synthesized even when

adequate levels are supplied in diet because of a specialized role

2. ARG → urea synthesis

ASP → urea synthesis

GLU → conduit for disposal of N

3. ALA & GLN → key role in exchange between tissues (liver & skeletal muscle)

4. Liver: major site gluconeogenesis (AA → Glucose)

major site urea synthesis (kidneys to a lesser extent)

5. Skeletal Muscle: 60% total body protein, 50% total body AA pool and is the major source to provide AA carbons → hepatic gluconeogenesis

AA are released from muscle during the post- absorptive state (O/N fast). Of the AA released by muscle ALA= 30% & GLN= 25% (total> 50%)

But output (ALA+GLN) > abundance in muscle proteins which contain 7-10% ALA & 6% GLN

Where does this ALA & GLN come from?

Sources of Alanine (from Muscle)

(i)Muscle: Protein → ALA + AA

AA → NH4+ + α keto acids

α keto acids → ALA (“simplest” AA).

Therefore total ALA released > ALA derived from proteins

(ii) Liver: ALA → NH4+ + α keto acids

NH4+ → urea

(iii) As well Glucose → Pyruvate (no N) → ALA (with N)

Therefore ALA serves as a vehicle for transport of NH4+ from

muscle to liver (NH4+ is generated through breakdown of AA

→ energy).

(iv) Because free NH4+ is very toxic even at low levels therefore

Pyruvate + NH4+ → ALA (non-toxic)

(v) In liver: NH4+ → urea for excretion

Glucose-Alanine cycleAmino group from excess glutamate produced in muscle as a result of amino acid catabolism, is transferred to pyruvate resulting in the formation of alanine.

Alanine is another safe way to transport ammonia from muscle to liver via blood.

In liver alanine aminotransferase transfers the amino gp to glutarate and pyruvate regenerated is used in gluconeogenesis.

Glucose produced by gluconeogenesis is transported to muscle where it enters the glycolysis.

Thus the excess puruvate and ammonia generated in muscle are safely transported to liver.

Role of Pyridoxal phosphate • PLP acts as a coenzyme in all transamination reactions,

and in some decarboxylation and deamination reactions of amino acids

• The aldehyde group of PLP forms a Schiff-base linkage (internal aldimine) with the ε-amino group of a specific lysine group of the aminotransferase enzyme

• The α-amino group of the amino acid substrate displaces the ε-amino group of the active-site lysine residue

• The resulting external aldimine becomes deprotonated to become a quinoid intermediate, which in turn accepts a proton at a different position to become a ketimine

• The resulting ketimine is hydrolysed so that the amino group remains on the complex

• In addition, PLP is used by aminotransferases (or transaminases) that act upon unusual sugars such as perosamine and desosamine

• In these reactions, the PLP reacts with glutamate, which transfers its alpha-amino group to PLP to make pyridoxamine phosphate (PMP)

• PMP then transfers its nitrogen to the sugar, making an amino sugar.

• It is also active in the condensation reaction in heme synthesis.

• Pyridoxal phosphate is not required in the transaminase reaction of lysine catabolism.

• PLP plays a role in the conversion of Dopa into Dopamine

• PLP allows the conversion of the excitatory neurotransmitter Glutamate to the inhibitory neurotransmitter GABA.

• PLP also allows SAM to be decarboxylated to form propylamine which is a precursor to polyamines.

• PLP allows the conversion of histidine to histamine via decarboxylation.

Sources of Glutamine (from Muscle) (i) Extra GLN released is also made from

other AA & serves as a non-toxic transport of NH4

+ from muscle → kidneys & gut (previous fig)

(ii) Kidneys: GLN → ALA (to the Liver )

& GLN → glucose (blood) +NH4+ (Urine)

(iii) Gut: GLN → ALA (to the liver)

Transport of excess ammonia by glutamine: Excess ammonia is toxic to animal tissues. Other than amino acid catabolism in tissues ammonia is also produced as a result of nucleic acid degradation.

Glutamine synthase catalyses the synthesis of glutamine by adding the ammonia to glutamate at the expense of ATP hydrolysis.

Glutamine is a non-toxic carrier of ammonia. It is transported to liver or kidney via blood.

In liver or kidney mitochondria, the glutamine is converted to glutamate and ammonia. Ammonia is incorporated in urea cycle in liver to be excreted.

Incorporation of NH4+ Into

Organic Compounds

1) NH4+ + HCO3

- + 2 ATP NH2CO2PO3

-2 + 2 ADP +

Carbamoyl Phosphate Pi + 2 H+

2) NH4+ +

Carbamoyl PhosphateSynthase I

(CPS-I)

Glutamate dehydrogenase

O-O2CCH2CH2CCO 2

-

a-Ketoglutarate Glutamate

NH3+

-O2CCH2CH2CHCO2-

TCA Cycle

NADP+NADPH + H+

mitochondria

Incorporation of NH4+ Into

Organic Compounds

NH3+

-O2CCH2CH2CHCO 2- + NH4

+ + 2 ATP

NH3+O

H2NCCH2CH2CHCO 2-

Glutamine

Glutamate Glutamine Synthase Mg++

N of glutamine donated to other compoundsin synthesis of purines, pyrimidines, and other amino acids

3)

Response to Food Deprivation

(i) For the first 7 days, maintain blood glucose (brain use 65% of glucose 400 - 600 Cal)

(ii) > 7 days: Protein proteolysis decreases (protect essential proteins) therefore use over a prolonged period compromises organism.

(iii) → Switch to Ketone bodies