Metabolism of amino acids I

Josef Fontana

EC

Overview of the lecture

• Introduction to protein and amino

acids metabolism

• Metabolic pathways of amino acids

–Transamination

–Conversion glutamate - glutamine

–Oxidative deamination of glutamate

–Urea cycle

Introduction to protein and amino

acids metabolism

Turnover of proteins in the

human body, basic reactions

of amino acids

Proteins

• Very intense metabolism - daily

turnover:

–skeletal muscle - 10 %

– liver - 40 %

–mucosa of the small intestine - 80 %

• Daily intake - 100 g

• Daily oxidation - 100 g = 10-20 % E

Proteins

Proteosynthesis Proteolysis

Proteins in diet Amino acids

pool

Amino acids

degradation

Amino acids

biosynthesis

Purines, pyrimidins, heme

Carbon skeleton

Urea

AAs metabolism

• Sources of AAs:

• 1) diet

• 2) degradation of body proteins

• 3) de novo synthesis

• AAs pool

• Use of AAs:

• 1) proteosynthesis

• 2) degradation (energy, glucose, FA)

• 3) synthesis of N-compounds

Protein turnover is strictly regulated

• AAs surplus can not be stored - no

storage protein!

• AAs serve as a fuel

Nitrogen balance

• Reflects the balance between the intake of

nitrogen in food and nitrogen losses

• Most healthy individuals will present with

the nitrogen balance in equilibrium

N intake = N losses

• Increased amount of protein in the diet - excess

amino acids are catabolized and their amino

group excreted as urea or ammonia

Positive nitrogen balance

• Protein intake in the diet exceeds the

protein losses

• During the recovery after illness,

during periods of growth or during an

administration of anabolic hormones

Negative nitrogen balance

• Nitrogen losses exceed its intake

• During starvation, severe illness or

during an administration of catabolic

hormones

• 1 g N – 6.25 g proteins

Degradation of cell proteins

• Cellular proteins have different half-

life

• Ornithine decarboxylase - 11 minutes

• Hemoglobin survives as long as

erythrocyte

• Υ-Crystallin (protein of the eye lens) -

the whole life

Regulation of cellular proteolysis

• Protein ubiquitin

• Marker for cellular

protein - label for

destruction

• Polyubiquitinisation

- degradation in

proteasomes

The Nobel Prize in Chemistry 2004 Aaron Ciechanover, Avram Hershko, Irwin Rose

"for the discovery of ubiquitin-mediated protein

degradation"

Essential and non-essential

amino acids

• Essential AAs

– branched: Val, Leu, Ile

– aromatic: Phe, Trp

– basic: Lys

– sulfur-containing: Met

– with hydroxy group: Thr

• Conditionally essential:

– Arg, His

• Non-essential AAs

– Gly, Ala, Ser, Pro, Cys, Tyr, Asn, Gln, Asp, Glu

Important reactions of AAs

• Decarboxylation

→ biogenic amines

• Transamination → 2-ketoacids

• Oxidative deamination

→ 2-ketoacids

• Formation of peptide bonds

→ peptides and proteins

Metabolic pathways of amino

acids

Transamination

Transamination

• Transaminases (aminotransferases)

• Specific for one pair of AA and the

corresponding α-keto acid

• Reversible reaction

• Pyridoxal phosphate (vit. B6 derivative)

• Liver enzymes:

• 1) ALT (alanine aminotransferase)

• 2) AST (aspartate aminotransferase)

Alanine aminotransferase (ALT)

Aspartate aminotransferase (AST)

Metabolic pathways of amino

acids

Conversion glutamate -

glutamine

Conversion glutamate - glutamine

• Conversion of the carboxyl group of

glutamate (in the side chain) in the

amide in glutamine

• Glutamine synthetase (cytosol)

• The most important transport form of

amino nitrogen in the blood

• Opposite reaction: glutaminase (MIT -

ammonia from Gln to the urea cycle)

Metabolic pathways of amino

acids

Oxidative deamination of

glutamate

Oxidative deamination of glutamate

• Glutamate dehydrogenase

• Mitochondria, mainly in the liver

• Amino group was previously transferred

to αKG by transamination - glutamate

synthesis

• Oxidative deamination releases -NH2 as

NH3 - restoration of αKG - goes to a new

transamination

Oxidative deamination of glutamate

Formation of ammonium

• α-amino groups are converted to ammonium

by oxidative deamination of glutamate

Fate of amino nitrogen derived

from AAs

• Extrahepatic tissues

• 1) Transamination:

forms mainly Ala

and Glu + 2-

oxoacids

• 2) Amidation:

• Glu + NH3 → Gln

In the liver

1) Same mechanisms

as in extrahepatic

tissues

2) Oxidative

deamination of Glu

(forms NH3 and

αKG): glutamate

dehydrogenase

Metabolic pathways of amino

acids

Urea cycle

Ammonium

• Conversion to urea

• Plasma concentration below 35

µmol/l

• Toxic for brain - nonpolar - freely

crosses the blood brain barrier

• Combines with α-KG - glutamate -

block of KC

Urea cycle

• Substrates: NH3 , CO2 and aspartate

• Liver, excreted in kidneys

• Mitochondria / cytosol

• Carbamoylphosphate synthetase I

• Needs lot of energy

• Connected with KC via fumarate

Synthesis of carbamoylphosphate

• Carbamoylphosphate synthetase I

• Mitochondria

• NH4+ + HCO3

-

• 2 ATP

• Citrulline is transported to cytosol

Synthesis of citrulline

Synthesis of arginosuccinate

Cleavage to arginine and

fumarate

• Arginine hydrolysis → urea and ornithine

• Transport of ornithine to matrix

Urea

Restoration of aspartate

• Close association with KC -

aspartate formation from fumarate

• Each degraded AA gives its amino

group to αKG - glutamate - AST

transfer to OAA - aspartate - urea

cycle - urea

Urea cycle - KC

Regulation of urea cycle

• Carbamoylphosphate synthetase I

• Activated by N-acetylglutamate

• produced in reaction: AcCoA + Glu

• N-acetylglutamate synthetase: activated

by arginine

• Protonproductive reaction - inhibited during

acidosis

• Increased transcription in high-protein diet

Metabolism of amino acids II

Josef Fontana

EC

Overview of the lecture

• Metabolic pathways of amino acids

–Utilization of the amino acids carbon

skeleton

–Formation of nonessential amino acids

• Important derivatives of amino acids

• Organ specifics of amino acids

metabolism

Metabolic pathways of amino

acids

Utilization of the amino acids

carbon skeleton

Carbon skeleton of AAs

• Carbon skeleton of each AA is converted by an

original pathway

• Degradation leads to a formation of 7

intermediates:

• acetyl-CoA

• acetoacetyl-CoA

• pyruvate

• α-ketoglutarate

• succinyl-CoA

• fumarate

• oxaloacetate

Ketogenic AAs

Glucogenic AAs

Aminoacids

• Ketogenic: Lys and Leu (begin with L)

• Glugogenic: serine, threonine,

cysteine, methionine, aspartate,

glutamate, asparagine, glutamine,

glycine, alanine, valine, proline, histidine

and arginine

• Keto- and glucogenic: isoleucine,

phenylalanine, tyrosine and tryptophan

7 degradation products of AAs

• pyruvate Gly, Ala, Ser, Thr, Cys, Trp

• oxaloacetate Asp, Asn

• -ketoglutarate Glu, Gln, Pro, Arg, His

• succinyl-CoA Val, Ile, Met, Thr

• fumarate Phe, Tyr

• acetyl-CoA Ile

• acetoacetyl-CoA Lys, Leu, Phe, Tyr, Trp

glucogenic AAs

ketogenic AAs

It is easy to deduce

• Aspartate and asparagine → OAA

(transamination)

• Glutamine and glutamate → αKG

(glutaminase and transamination)

• Alanine → pyruvate (transamination)

• Lysine and leucine are ketogenic → AcCoA

and acetoacetylCoA

• Glycine, serine and cysteine (small AAs) –

converted to pyruvate

Degradation of branched AAs

• 1st step: transamination

– specific transaminase

– ↑ activity in skeletal muscle and heart, ↓

activity in liver

– product: 2-oxoacids

• 2nd step: dehydrogenation +

decarboxylation

– product: acyl-CoA

Metabolic pathways of amino

acids

Formation of nonessential

amino acids

Synthesis of AAs

• Essential: Phe, Trp, Val, Leu, Ile, Met, Thr, Lys

• Conditionally essential: Arg, His

• Nonessential:

• oxalacetate → Asp, Asn

• 2-ketoglutarate → Glu, Gln, Pro, (Arg)

• pyruvate → Ala

• 3-phosphoglycerate → Ser, Cys, Gly

• Phe → Tyr

Tyrosine from Phenylalanine

Phenylketonuria

• AR, absence or reduced activity

of phenylalanine hydroxylase

• Degradation of Phe:

phenylpyruvate (urine) →

phenyllactate, phenylacetate

• Degradation of Phe:

phenylethylamine H5C

6-CH

2-

CH2-NH

2 (brain damage)

• Screening in newborns

Metabolic pathways of amino

acids

Important derivatives of amino

acids

Decarboxylation of AAs gives

monoamines (= biogenic amines)

• Tyr → catecholamines

• Trp → serotonin (5-hydroxytryptamine)

• His → histamine

• Ser → etanolamine → choline → acetylcholine

• Cys → cysteamine

• Asp → β-alanine

• Glu → γ-aminobutyrate (GABA)

coenzyme A

Nitric oxide

Nitric oxide

• NO-synthase (NOS)

– in neurons: NOS-I: neurotransmission

– in macrophages: NOS-II: kills bacteria

– endothelial: NOS-III: vasodilation

• Clinical correlation:

– nitrates in the treatment of angina

pectoris

– hypotension during septic shock

Thyroid hormones

OH CH

2

CH

CO

NH

OH CH

2

CH

CO

NH

I

OH CH

2

CH

CO

NH

I

I

I

OH O CH

2

CH

NH2

COOH

I

I

I

OH O CH

2

CH

NH2

COOH

I

I

I

Thyreoglobulin

Thyreoglobulin

Tyr

MIT

Thyreoglobulin

DIT

Trijodthyronin (T3)

Thyroxin (T4)

Melanin

• Pigment derived from tyrosine (its

oxidation and polymerization)

• There are two types:

–oculocutaneous - skin melanocytes

–neuromelanin - in substantia nigra of

the midbrain (mesencephalon)

Formation of activated methionine

= S-adenosylmethionine (SAM)

SAM is used as -CH3 group donor

in metabolic methylations Figure is found on http://themedicalbiochemistrypage.org/amino-acid-metabolism.html#cysteine

Synthesis

of creatine

Organ specifics of amino acids

metabolism

Blood

• Total blood concentration of AAs: 2.3-4.0

mM

• Glutamine: 0.6 mmol/l (main transport

form of ammonia) and alanine: 0.3 mmol/l

• Ammonia: 6-35 µmol/l

• Urea: 2.5-8.3 mmol/l

• Creatinine: 50-120 µmol/l

Liver

• Main organ of AAs metabolism

• Removal of amino group from Aas

• Detoxification of ammonia - urea

cycle and systhesis of glutamine

• C-skeleton metabolism - glucose, FA

or ketone bodies

• Synthesis of non-essential AAs

Intestine - enterocytes

• Change spectrum of ingested AAs - concentration

in the portal blood vary (e.g. more proline and

citrulline - formed from Glu / Gln)

• Glutamine is an essential energy substrate for

rapidly dividing cells (e.g. immune cells and

enterocytes)

• Gln amino group enters the formation of purines,

oxidation of C-skeleton gives energy

• Skeletal muscle is a major source of glutamine

during starvation and stress

Skeletal muscle

• The main "reservoir" of proteins - use during

stress and starvation

• Muscle changes spectrum of AAs released

into the blood also (in comparison with AAs

obtained by proteolysis of muscle proteins)

• Branched AAs transaminated in the muscle -

their α-ketoanalogues are released into the

blood (or are oxidized to gain E), amino groups

transferred to glutamine or alanine - released

to the blood

Kidneys

• Main place of N-catabolites

excretion: urea, ammonia, creatinine,

uric acid etc.

• Tubular cells: conversion of Gln - Glu -

α-KG, ammonia excreted in the

urine

• Gluconeogenesis

• Conversion citrulline - arginine

Marasmus Kwashiorkor

• Inadequate intake of

carbohydrates, fats and

proteins → not covered

energy requirements of the

organism

• Patients in the Hospice Unit

→ appearance of "skin and

bones"

• Treatment: nutritional

intervention (enteral or

parenteral), treatment of the

disease

Inadequate protein

intake (and essential

AA) with adequate

energy intake

Symptoms: retarded

growth, loss of skin

and hair

pigmentation, ascites,

mental apathy