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METABOLISM and

UTILIZATION
OF
AMINO ACIDS
Amino nitrogen Carbon atoms
becomes urea become common
intermediates

AMINO ACIDS

Used in Glutamate as
anabolic a neuro-
pathways transmitter
I. Amino-acid Nitrogen Removal by
Transamination and its Utilization for Urea
Transamination requires:
1) An amino acid and an oxo- (keto-) acid
2) Vitamin pyridoxol
3) An aminotransferase.

NH2 O O O
| || || ||
R1CH– C OH + R2 C– C– OH OH

O O NH2 O
|| || | ||
R1– C – C– O H + R2 –CH – C – OH
The oxoacid can be either

pyruvate, which produces alanine,

oxaloacetate, which produces aspartate

2-oxoglutarate, which produces glutamate


Transamination with 2-oxoglutarate is important
because this reaction allows an amino acid nitrogen to
become the nitrogen of ammonia.

NH2 O
| ||

R– CH– C– O H + 2-OG
O NH2 O
|| | ||
HO– C – CH2 – CH2 –CH – C– OH

NH3
Conversion of glutamate into ammonia (NH3) is catalyzed
by
glutamate dehydrogenase
(uses NAD+)

O NH2 O
|| | ||

HO–C– CH2– CH2– CH - C – OH

2-oxoglutarate + NH3
When the nitrogen of an amino acid is not needed for growth
and maintenance it is excreted in the form of urea.

Urea is synthesized by the Urea Cycle.

The first and rate-determining step requires ammonia.

NH3 + HCO3- + 2 ATP

O O
|| ||

2 H N– C– O – P – OH
||

O
CARBAMOYL PHOSPHATE
First and rate determining step (RDS) of the urea cycle
is catalyzed by

carbamoylphosphate synthetase.

There are, in fact, two enzymes with this name. One

is involved in pyrimidine synthesis (in humans).

The other is the urea cycle enzyme that is designated

carbamoylphosphate synthetase I

or CPS I.
CARBAMOYL PHOSPHATE
+
UREA Ornithine

Arginine UREA Citrulline


CYCLE
Fumarate Aspartate

Argininosuccinate
REGULATION OF THE UREA CYCLE

Acute: N-acetylglutamate, an allosteric effector,

up regulates carbamoylphosphate synthetase

At the level of gene expression: Hepatic nuclear

factors (HNFs) provide tissue specificity


Excessive ammonia is toxic to the central nervous
system!

Treatment
Induce the formation of a compound that contains

the ammonia nitrogen, that is then excreted.

Sodium benzoate induces production of hippuric acid.

Sodium phenylacetylglutamate induces production of

phenylacetylglutamine.
NH3 + CO2 + 5,10-methylenetetra-
hydrofolate
NH2 O
| || Glycine

CH2 – C – OH
Sodium Benzoate

Bz-N-H O
| ||

CH2 – C - OH
Benzoyl glycine or hippuric acid
NH3 + Phenylacetylglutamate

Phenylacetylglutamine
In the presence of excessive ammonia, glutamine is
synthesized in the brain’s astrocytes by the glutamine
synthetase reaction, i.e.,
NH3 + Glutamate + ATP → Glutamine + ADP + Pi .
Water then enters the cell, producing an “intracranial
mass” that is manifested as edema. The astrocytes
become dysfunctional which has ramifications with
respect to the regulation of extracellular potassium
concentrations, among other processes.
II. Catabolic Fates of Amino-acid
Carbon Atoms

Carbons derived from amino acid can become those of

five very common biochemical intermediates.

These are:
pyruvate acetoacetate

acetyl-CoA succinyl-CoA

2-oxoglutarate
A. Amino Acid Degradation Provides Carbon Atoms
of Pyruvate

Examples of carbon atoms of amino acids becoming

those of pyruvate are found in glycine and serine.

Glycine can be converted into serine and then the

carbon atoms of serine can become those of pyruvate.


NH2 O
| ||
CH2 – C – OH + 5,10-methyleneTHF
glycine

OH NH2 O
| | ||

CH2 CH – C – OH + THF (non-ligated)


serine
OH NH2 O O O
| | || || ||
CH2– CH – C –OH CH3– C– C– OH
pyruvate
The third carbon atom is from the 5,10 methylene

group of 5,10 methylenetetrahydrofolate.

Tetrahydrofolate is a vehicle for specific

“one-carbon units.” These include formyl,

formimmino, methenyl, methylene, and methyl.


Amino-acid carbon atoms that become those of

pyruvate can, in turn, become those of

glucose (gluconeogenesis)

and also those of

palmitic acid (pyruvate  acetyl-CoA)

cholesterol (pyruvate  acetyl-CoA)

steroids (from cholesterol)


B. Amino Acid Degradation Provides Carbon Atoms

of Acetyl-CoA

One example is found in the case of the branched

chain amino acids leucine and isoleucine.

Leucine is degraded to yield acetyl-CoA and acetoacetate

(another of the five key biochemical intermediates).

Isoleucine yields acetyl-CoA and propionyl-CoA

(precursor of another of the five).


The rate-determining reaction in the degradation

of leucine or isoleucine is catalyzed by a branched-

chain oxoacid dehydrogenase (BCOD)

complex that is similar to the pyruvate dehydrogenase


complex.

The activity of the BCOD complex is tightly regulated

by nutritional state and by hormones.


These factors exert their effects by controlling the
activity and/or the expression of a specific protein kinase
that modulates the activity of the BCOD complex. With
respect to acute regulation, phosphorylation of the BCOD
complex by its kinase (the BCOD kinase) down-regulates
the activity of the complex.
Activity of the BCOD complex is also regulated by

controlling the gene expression of the dehydrogenase

subunit. This is appropriate because it is the activity of

the dehydrogenase subunit that is rate-determining for

the degradative process.


Starvation (>24 hours) leads to decreased BCOD
kinase activity in the liver and, therefore, increased
protein degradation.
Diabetes, which can associate with increased
protein degradation, also associates with decreased
BCOD kinase activity. Since insulin promotes the
activity of the BCOD kinase, it would follow that its
lack, as in diabetes, would lead to (decreased activity of
the kinase and) increased protein degradation.
STARVATION DIABETES INSULIN
__________________________________________________
BCOD kinase kinase kinase

Kinase BCOD

BCOD protein
degradation

Degradation Degradation Degradation


Amino-acid carbon atoms that become those of

acetyl-CoA can, in turn, become those of

citrate (by entering the TCA cycle)

palmitic acid (fatty acid synthesis)

cholesterol (cholesterol synthesis uses 3)

steroids (from cholesterol)


C. Amino Acid Degradation Provides Carbon Atoms

of Acetoacetate.

Examples are found in the case of phenylalanine

and tyrosine. Phenylalanine can be converted

into tyrosine and tyrosine can be further degraded

(many steps) into acetoacetate and fumarate.


Acetoacetate is one of the “ketone bodies” and

the precursor of the other two.

ACETOACETATE

-CO2 REDUCTION

ACETONE 3-HYDROXYBUTYRATE
“Ketone bodies” are produced in the liver in order to

provide substrates to be utilized by the brain in times

of glucose deprivation. However, excessive “ketone

body” production characterizes uncontrolled diabetes.

In fact, acetoacetic acid and 3-hydroxybutyric acid

contribute to the metabolic acidosis that frequently

accompanies diabetes.
Acetone and acetoacetate are each ketones and

Contribute to the ketosis that frequently

accompanies diabetes.

Diabetes not only associates with increased protein

degradation, as mentioned above, it also associates with

increased circulating branched chain amino acids. The

latter contribute to the production of “ketone bodies.”


D. Amino Acid Degradation can Provide Carbon

atoms of Succinyl-CoA.

Isoleucine, valine, threonine, and methionine

(so-called “essential” amino acids) are degraded to yield

propionyl-CoA that is, in turn, converted into

succinyl-CoA by a vitamin B12-dependent reaction.

Succinyl-CoA can enter the TCA cycle.


E. Amino Acid Degradation can Provide Carbon

Atoms of 2-Oxoglutarate (-ketoglutarate).

These amino acids, likewise, provide carbon atoms

that can be catabolized by the TCA cycle to produce

energy.
aspartate alanine PYRUVATE (I) serine glycine
tryptophan cysteine

ACETYL CoA (II)


tryptophan leucine lysine isoleucine

Acetoacetate (III)
tyrosinefumarate

phenylalanine
TCA
2-OG (V)
Cycle 
glutamate  histidine

proline arginine

methionine isoleucine SUCCINYL CoA (IV)


threonine valine propionyl CoA
III. ANABOLIC PATHWAYS THAT
UTILIZE AMINO ACIDS

A. Biosynthesis of creatine

arginine + glycine guanidino acetic acid

S-adenosyl-L-methionine

creatine
B. Phenylalanine and tyrosine are precursors of the
catecholamines, dopamine,
norepinephrine, and epinephrine.
1) Clinical ramifications involving dopamine:
Dopamine has a role in the pathogenic mechanisms
that cause Parkinson’s disease. Parkinson’s disease
is characterized by a loss of dopaminergic neurons
that results in diminished levels of dopamine in the
striatum.
It has recently been suggested that diminished uptake of
dopamine into its storage vesicles is a causative factor in
Parkinson’s. A protein called α-synuclein may have
a role in this process since it promotes the formation of
these vesicles. Dopamine that is not sequestered intra-
cellularly can have deleterious effects upon the cell.
Dopamine is synthesized in the cytoplasm and
(normally) immediately sequestered in intracellular
storage vesicles through the agency of α-synuclein.
2. Biosynthesis of the catecholamines

phenylalanine

tyrosine
Tyrosine 3-monooxygenase*
dihydroxyphenylalanine (DOPA)

dopamine

norepinephrine
S-AM

epinephrine
*Rate-determining step
C. Tryptophan is the precursor of 5-hydroxy-
tryptamine (serotonin).
1. Clinical ramifications
The broad actions of the neurotransmitter 5-
hydroxytryptamine involve effects upon emotion,
mood, and reward. Suicide and depression have been
associated with reduced serotonergic transmission.
One recent postmortem study examined the capacity
to synthesize 5-hydroxytryptamine and the capacity
to bind 5-hydroxytryptamine. Their findings indicated
that the brains of suicide victims had had reduced
serotonergic function.
2. Biosynthesis of 5-hydroxytryptamine
In the conversion of tryptophan into 5-hydroxy-
tryptamine, tryptophan undergoes hydroxylation to
yield 5-hydroxytryptophan that is then decarboxylated
to yield 5-hydroxytryptamine.
Reactions that convert tyrosine or tryptophan into
neurotransmitters.
It has been shown that that the intensity and
duration of catecholamine signaling at the synapse is
governed by the efficiency of the re-uptake of released
neurotransmitter. Re-uptake occurs by means of high-
affinity membrane transporters. The finding that a
dopamine “re-uptake” transporter can serve as a
cocaine “receptor” has provided new insight into
mechanisms of addiction.
Cartoon depicting the binding of cocaine (and other
agents) by the membrane transporters that facilitate
reuptake.
D. Arginine is the precursor of the second
messenger nitric oxide. This a second messenger
that appears to have distinctly different functions in
different cell types. For example, the nitric oxide
synthesized by platelets is seen to inhibit platelet
aggregation and adherence. In this way it
contributes to the anti-thrombogenic properties of
the endothelium.
In the kidney (in experimental animals), a diminution in
nitric oxide was associated with glucocorticoid-induced
hypertension. Such studies were carried out because it
had been suggested that decreased nitric oxide
contributes to the impaired endothelium-dependent
vasodilatation seen in essential hypertension.
On the other hand, cortical cells from patients with
Alzheimer’s disease showed increased levels of nitric
oxide mRNA and protein. Hence, nitric oxide has been
associated with the progression of Alzheimer’s disease.

From an overview however, nitric oxide is generally


considered to be a vasodilator.
As a second messenger, nitric oxide up regulates the
activity of the enzyme that synthesizes cyclic GMP, i.e.,
guanylyl cyclase. The cyclic GMP then up regulates a
cyclic GMP-dependent protein kinase. The protein
kinase, in turn, phosphorylates specific substrates in
order to bring about its effects.
For their pioneering work that lead to the discovery of
nitric oxide (NO) and its signaling in humans,

Dr. Robert Furchgott (SUNY, Brooklyn, NY)

Dr. Louis J. Ignarro (UCLA, Los Angeles, CA)

Dr. Ferid Murad (Univ. Texas Med. Sch.


Houston, TX)

received the 1998 Nobel Prize (Physiology or Medicine).


2. Biosynthesis of nitric oxide
Nitric oxide is synthesized by the enzyme nitric
oxide synthase (NOS). There are three types of
enzymes, the neuronal nitric oxide synthase (nNOS),
the endothelial nitric oxide synthase (eNOS), and the
inducible nitric oxide synthase (iNOS). They each
catalyze the complex oxidation/reduction reaction that
utilizes one of arginine’s nitrogen atoms in order to
make the “free radical” nitric oxide.
The NOS reaction requires NADPH, FAD, and FMN
along with tetrahydrobiopterin (BH4) and heme (Fe).
An electron transport chain has been formulated for
the reaction.
E. Methionine has multiple anabolic fates.
1. As a precursor of the polyamines,
spermidine and spermine.
While it is generally accepted that the polyamines,
spermidine and spermine, are essential for cell
proliferation in mammalian cells, the critical reaction(s)
have not been identified. Inhibiting ornithine
decarboxylase (ODC) (the rate-determining enzyme)
has been shown to inhibit cell growth, however.
Biosynthesis of the polyamines from methionine.

METHIONINE

S-Adenosyl-L-methionine ORNITHINE
RDS

Decarboxylated S-AM + Putrescine

SPERMIDINE

SPERMINE
2. As a precursor of cysteine
Cysteine is synthesized in humans using the carbons
of serine and the sulfur of methionine.
METHIONINE

SERINE + Homocysteine

CYSTEINE
3. As a precursor of S-adenosyl-L-
methionine (SAM)
The methyl donor S-adenosyl-L-methionine is derived
from methionine.
METHIONINE + ATP

S-ADENOSYL-L-METHIONINE + Pi + PPi
4. As a necessary participant in the
synthesis of deoxyribothymidylic acid and, hence, DNA
The conversion of deoxyribouridylic acid
monophosphate (dUMP) into the corresponding
thymidylic acid derivative (dTMP) depends upon
converting 5-methyltetrahydrofolate into the unligated
tetrahydrofolate. This is accomplished by the enzyme
methionine synthase.
The synthesis of deoxyribothymidylic acid (dTMP)

requires both vitamin B12 and folic acid.


5-methyl THF
methionine synthase homocysteine
vitamin B12 METHIONINE
THF (non-ligated)
serine hydroxymethyl serine
transferase glycine

5,10-methylene THF
thymidylate dUMP
synthase dihydrofolate
dTMP
In summary, methionine provides carbon and
nitrogen atoms that become a part of spermidine and
spermine, the sulfur atom that is required for the
biosynthesis of cysteine, and the “active” methyl
group of the methyl donor, S-adenosyl-L-methionine.
H3C ─S NH O

CH2─CH2─CH──C─OH
METHIONINE
POLYAMINES
THF
5-methylTHF
ODC

Homocysteine SAM

cystathionine
methyl acceptor

S-Adenosylhomocysteine

Cysteine Glutathione
IV. “INBORN ERRORS” OF AMINO
ACID METABOLISM
A. PHENYLKETONURIA

1. Clinical Ramifications:
neurological, mental retardation

2. Critical Reaction:
phenylalanine + tetrahydrobiopterin + O2

tyrosine + dihydrobiopterin + H2O


B. ALBINISM

1. Clinical Ramifications:
usually not severe, sensitivity to sun

2. Critical Reaction:

Melanogenesis entails the oxidation of tyrosine

by a tyrosinase. In vertebrates this enzyme is active

only in specialized organelles including, primarily, the

retinal pigment epithelium and melanocytes.


The oxidation of tyrosine yields dopaquinone that,

subsequently, is converted into the polymer referred to

as melanin. It has recently been shown that

oculocutaneous albinism (OCA), type II, an autosomal

recessive disorder, results from failure of appropriate

intracellular transport of the tyrosinase.


Studies showed that in affected individuals, the

melanocytes contained markedly diminished

quantities of tyrosinase. The fault resided in a gene

that encodes a protein that is critical for the processing

and transport of tyrosinase to the melanocyte.


C. BRANCHED-CHAIN AMINOACIDURIA

1. Clinical Ramifications:
neurological effects, mental retardation,
ketoacidosis

2. Critical Reaction:
“keto-leucine” + oxidized lipoamide
THIAMINE
DIPHOSPATE

O
||

3HC– CH2– CH2 – C – LIPOAMIDE


|

3HC
This reaction is catalyzed by the BCOD component of
the multi-enzyme complex. (It is rate-determining for
the process.) Several mutations in this enzyme have
been identified and characterized. Attempts are being
made to correlate particular mutations with the severity
of the disease.
D. HOMOCYSTEINURIA

1. Clinical ramifications:

First described in 1962 in relation to mental

retardation in children. Now associated primarily with

vascular pathologies including atherosclerosis and

premature thromboembolic disease. Also associated with

certain CNS disorders and abnormal bone development.


Homocysteine can alter the surface properties of
endothelial cells, causing them to be procoagulant
as a result of inhibiting antithrombin III and inhibiting
the synthesis of an anticoagulant (heparan sulfate). This
has been offered as a rationale for the suggested role of
homocysteine in atherosclerosis.
2. Critical reaction
The most common cause for genetically determined
homocysteinuria is a “loss of function” mutation that
results in decreased activity of the enzyme
cystathionine β-synthase. This is the enzyme that
catalyzes the formation of cysteine using the carbon
atoms of serine and a sulfur from methionine (as
discussed above).
V. Glutamate and its Receptors and
Signaling
The amino acid glutamate is an endogenous excitatory

neurotransmitter, utilized by neurons in the human

central nervous system and the peripheral nervous system.

Glutamate acts by binding glutamate receptors that

allow it to play a pivotal role in synaptic mechanisms

involved in learning and memory.


Learning and memory involve synaptic plasticity, the
phenomenon in which the efficacy, or efficiency, of
synaptic transmission varies in an activity-dependent
manner. The effect can be transient (short-term) or
persistent (long-term). Signal transduction involving
glutamate receptors governs the reactions involved in
these processes..
Cartoon depicting signal transduction involving the
glutamate receptor in the phenomenon of synaptic
plasticity.
Glutamate binds two types of receptors, the
ionotropic glutamate receptors and the metabotropic
receptors. There are three of the former, the NMDA
(N-methyl-D-aspartate), the AMPA (α-amino-3-
hydroxy-5-methyl-4-isoxazolepropionic acid), and the
kainate receptors. They are referred to as ionotropic
because they associate to form cation channels that
facilitate the entry of Ca2+ (or Na+) into the cell.
Glutamate receptor molecules form dimers and then
tetramers that function as ion channels.
These ion channels are ligand-gated channels that
assume an “open” configuration upon glutamate (or
agonist) binding. They are locked in a “closed”
configuration when an antagonist is bound. It should
be remembered that when a protein binds a ligand, the
protein generally undergoes a conformational change
that can alter its properties and its activity.
Effect of an antagonist, a partial agonist, and the full
agonist, glutamate upon the conformation of the
glutamate receptor.
Excessive glutamate binding to glutamate

receptors in the brain can bring about excitotoxic

neuronal cell death, however. Excessively activated

receptors lead to brain damage seen with cerebral

ischemia, traumatic brain injury, and the neuro-

degeneration associated with Huntington’s chorea,

epileptic disorders, and Alzheimer’s disease.


These excitotoxic effects are due, in part, to
signaling downstream of certain glutamate receptors that
leads to in increased intracellular Ca2+. The ionotropic
N-methyl-D-aspartate (NMDA) glutamate receptor is
one of the principal ones that facilitates the entry of
extracellular calcium (Ca2+) into the neuronal cell.
Calcium (Ca2+) that enters the neuronal cell by way of
NMDA channels can, among other actions, up regulate
the synthesis of nitric oxide by the nNOS.
As a result of increasing intracellular Ca2+ NMDA
glutamate receptors have broad influence, modulating
the activities of a variety of protein kinases and
phosphoprotein phosphatases.
Recently, it has been suggested that the NMDA
glutamate receptor and a (D1) dopamine receptor
cooperate in the molecular mechanisms of compulsion
and persistence as related to drug adiction.
TRANSAMINATION CATABOLISM
pyruvate acetyl-CoA
acetoacetate succinyl-CoA
GLUTAMATE 2-oxoglutarate

AMINO ACIDS ANABOLISM


UREA
creatine,
catechol-
INBORN amines, HT,
ERRORS NO, poly-
PKU, amines,
GLUTAMATE
Albinism,
RECEPTOR cysteine,
SIGNALING BCAA uria, SAM, dTMP
HC uria