Download 30_General pathways of amino acids transformation

PROTEIN
METABOLISM:
PROTEIN TURNOVER;
GENERAL WAYS OF
AMINO ACIDS
METABOLISM
Proteins function in the organism.
 All enzymes are proteins.
 Storing amino acids as nutrients and as building blocks
for the growing organism.
 Transport function (proteins transport fatty acids,
bilirubin, ions, hormones, some drugs etc.).
 Proteins are essential elements in contractile and motile
systems (actin, myosin).
 Protective or defensive function (fibrinogen,
antibodies).
 Some hormones are proteins (insulin, somatotropin).
 Structural function (collagen, elastin).
PROTEIN TURNOVER
Protein turnover — the degradation and
resynthesis of proteins
Half-lives of proteins – from several minutes to many
years
Structural proteins – usually stable (lens protein
crystallin lives during the whole life of the organism)
Regulatory proteins - short lived (altering the amounts
of these proteins can rapidly change the rate of
metabolic processes)
How can a cell distinguish proteins that are meant
for degradation?
Ubiquitin - is the tag that
marks proteins for
destruction ("black spot" - the
signal for death)
Ubiquitin - a small (8.5-kd)
protein present in all
eukaryotic cells
Structure:
 extended carboxyl terminus
(glycine) that is linked to other
proteins;
 lysine residues for linking
additional ubiquitin molecules
Ubiquitin covalently
binds to -amino group
of lysine residue on a
protein destined to be
degraded.
Isopeptide bond is
formed.
Mechanism of the binding of ubiquitin to target protein
E1 - ubiquitin-activating enzyme (attachment of ubiquitin
to a sulfhydryl group of E1; ATP-driven reaction)
E2 - ubiquitin-conjugating enzyme (ubiquitin is shuttled
to a sulfhydryl group of E2)
E3 - ubiquitin-protein ligase (transfer of ubiquitin from
E2 to -amino group on the target protein)
Attachment of a single
molecule of ubiquitin - weak
signal for degradation.
Chains of ubiquitin are
generated.
Linkage – between -amino
group of lysine residue of
one ubiquitin to the terminal
carboxylate of another.
Chains of ubiquitin
molecules are more
effective in signaling
degradation.
What determines
ubiquitination of the
protein?
1. The half-life of a protein
is determined by its aminoterminal residue (Nterminal rule).
E3 enzymes are the readers
of N-terminal residues.
2. Cyclin destruction boxes
- specific amino acid
sequences (proline, glutamic
acid, serine, and threonine –
PEST)
Digestion of the Ubiquitin-Tagged Proteins
What is the executioner of the protein death?
A large protease complex proteasome or the
26S proteasome digests the ubiquitinated
proteins.
26S proteasome - ATP-driven multisubunit
protease.
26S proteasome consists of two components:
 20S - catalytic subunit
 19S - regulatory subunit
20S subunit
 resembles a barrel
 is constructed from 28 polipeptide chains which are
arranged in four rings (two  and two )
 active sites are located in  rings on the interior of the
barrel
 degrades proteins to peptides (seven-nine residues)
19S subunit
 made up of 20 polipeptide
chains
 controls the access to interior
of 20S barrel
 binds to both ends of the 20S
proteasome core
 binds to polyubiquitin chains
and cleaves them off
 possesses ATPase activity
 unfold the substrate
 induce conformational changes
in the 20S proteasome (the
substrate can be passed into the
center of the complex)
Overview of Amino Acid Catabolism:
Interorgan Relationships
Overview of Amino Acid Catabolism:
Interorgan Relationships
• Liver
– Synthesis of liver and plasma proteins
– Catabolism of amino acids
•
•
•
•
Gluconeogenesis
Ketogenesis
Branched chain amino acids (BCAA) not catabolized
Urea synthesis
– Amino acids released into general circulation
• Enriched in BCAA (2-3X)
Overview of Amino Acid Catabolism:
Interorgan Relationships
• Skeletal Muscle
– Muscle protein synthesis
– Catabolism of BCAA
• Amino groups transported away as alanine and
glutamine (50% of AA released)
– Alanine to liver for gluconeogenesis
– Glutamine to kidneys
• Kidney
– Glutamine metabolized to a-KG + NH4
• a-KG for gluconeogenesis
• NH4 excreted or used for urea cycle (arginine synthesis)
– Important buffer from acidosis
GENERAL WAYS OF AMINO
ACIDS METABOLISM
The fates of amino acids:
1) for protein synthesis;
2) for synthesis of other nitrogen containing
compounds (creatine, purines, choline,
pyrimidine);
3) as the source of energy;
4) for the gluconeogenesis.
The general ways of amino acids
degradation:
 Deamination
 Transamination
 Decarboxilation
The major site of amino acid
degradation - the liver.
Deamination of amino acids
Deamination - elimination of amino group
from amino acid with ammonia formation.
Four types of deamination:
- oxidative (the most important for
higher animals),
- reduction,
- hydrolytic, and
- intramolecular
Reduction deamination:
R-CH(NH2)-COOH + 2H+  R-CH2-COOH + NH3
amino acid
fatty acid
Hydrolytic deamination:
R-CH(NH2)-COOH + H2O  R-CH(OH)-COOH + NH3
amino acid
hydroxyacid
Intramolecular deamination:
R-CH(NH2)-COOH  R-CH-CH-COOH + NH3
amino acid
unsaturated fatty acid
General scheme of oxydative
transamination
R CH COOH
+
NH2
HOOC C CH2CH2COOH
O
2-oxoglutarate
2-oxoglutarát
aminokyselina
amino acid
aminotransferase
aminotransferasa
pyridoxalfosfát
pyridoxal
phosphate
R C
COOH
O
2-oxokyselina
2-oxo acid
+
HOOC CH CH2CH2COOH
NH2
glutamát
glutamate
Glutamate dehydrogenase (GMD, GD, GDH)
• requires pyridine cofactor NAD(P)+
• GMD reaction is reversible: dehydrogenation with NAD+,
hydrogenation with NADPH+H+
• two steps:
• dehydrogenation of CH-NH2 to imino group C=NH
• hydrolysis of imino group to oxo group and ammonia
Oxidative deamination
L-Glutamate dehydrogenase plays a central role in amino acid
deamination
In most organisms glutamate is the only amino acid that has
active dehydrogenase
Present in both the cytosol and mitochondria of the liver
Transamination of amino acids
Transamination - transfer of an amino group from
an -amino acid to an -keto acid (usually to
-ketoglutarate)
Enzymes: aminotransferases (transaminases).
-amino acid
-keto acid
-keto acid
-amino acid
There are different transaminases
The most common:
alanine aminotransferase
alanine + -ketoglutarate  pyruvate + glutamate
aspartate aminotransferase
aspartate + -ketoglutarate  oxaloacetate +
glutamate
Aminotransferases funnel -amino groups from a
variety of amino acids to -ketoglutarate with
glutamate formation
Glutamate can be deaminated with NH4+ release
Mechanism of transamination
All aminotransferases require the
prosthetic group pyridoxal
phosphate (PLP), which is derived
from pyridoxine (vitamin B6).
Ping-pong kinetic mechanism
First step: the amino group of
amino acid is transferred to
pyridoxal phosphate, forming
pyridoxamine phosphate and
releasing ketoacid.
Second step: -ketoglutarate
reacts with pyridoxamine
phosphate forming glutamate
Ping-pong kinetic mechanism of aspartate transaminase
aspartate + -ketoglutarate  oxaloacetate + glutamate
Decarboxylation of amino acids
Decarboxylation – removal of carbon dioxide from
amino acid with formation of amines.
amine
Usually amines have high physiological activity
(hormones, neurotransmitters etc).
Enzyme: decarboxylases
Coenzyme – pyrydoxalphosphate
Significance of amino acid decarboxylation
1. Formation of physiologically active compounds
GABA –
mediator of
nervous
system
glutamate
histidine
gamma-aminobutyric acid (GABA)
histamine
Histamine – mediator of inflammation, allergic reaction.
1) A lot of histamine is formed in inflamatory place;
It has vasodilator action;
Mediator of inflamation, mediator of pain;
Responsible for the allergy development;
Stimulate HCI secretion in stomach.
-CO2
2) Tryptophan  Serotonin
Vasokonstrictor
Takes part in regulation of arterial pressure, body temperature, respiration,
kidney filtration, mediator of nervous system
3) Tyrosine  Dopamine
It is precursor of epinephrine and norepinephrine. mediator of central
nervous system
4) Glutamate  -aminobutyrate (GABA)
Is is ingibitory mediator of central nervous system. In medicine we use with
anticonvulsion purpose (action).
2. Catabolism of amino acids during the decay
of proteins
Enzymes of microorganisms (in colon; dead organisms)
decarboxylate amino acids with the formation of
diamines.
ornithine
putrescine
lysine
cadaverine