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Nucleotide Metabolism
Pyrimidine Met.
Purine Met.
Learning Objectives
1. How Are Purines Synthesized?
2. How Are Purines Catabolized?
3. How Are Pyrimidines Synthesized and
Catabolized?
4. How Are Ribonucleotides Converted to
Deoxyribonucleotides?
5. How Is dUTP Converted to dTTP?
6. Abnormal metabolism of uric acid.
7. Anticancer drugs targets
Two types of pathways lead to nucleotides: the
de novo pathways and the salvage pathways.
De novo synthesis of nucleotides begins with
their metabolic precursors:
amino acids, ribose 5-phosphate, CO2, and NH3.
Salvage pathways recycle the free bases and
nucleosides released from nucleic acid
breakdown.
Salvage and de Novo Pathways
The purine ring structure is built up one or a few
atoms at a time, attached to ribose
throughout the process.
The pyrimidine ring is synthesized as orotate,
attached to ribose phosphate, and then
converted to the common pyrimidine
nucleotides required in nucleic acid synthesis.
Although the free bases are not intermediates in
the de novo pathways, they are intermediates
in some of the salvage pathways.
de Novo Pathway for Purine
Nucleotide Synthesis
The synthesis of the purine ring is more complex.
The only major component is glycine, which
donates C-4 and C-5, as well as N-7. All of the
other atoms in the ring are incorporated
individually. C-6 comes from HCO3–. Amide
groups from glutamine provide the atoms N-3
and N-9. The amino group donor for the
inclusion of N-1 is aspartate, which is
converted into fumarate in the process, in the
same way as in the urea cycle . Finally, the
carbon atoms C-2 and C-8 are derived from
formyl groups in N10- formyl-tetrahydrofolate
The Purine Ring System Is Assembled
on Ribose Phosphate
Glutamine phosphoribosyl amidotransferase catalyzes
this reaction.
De novo purine biosynthesis, like pyrimidine
biosynthesis, requires PRPP, but for purines, PRPP
provides the foundation on which the bases are
constructed step by step. The initial committed
step is the displacement of pyrophosphate by
ammonia, rather than by a preassembled base,
to produce 5-phosphoribosyl-1-amine, with the
amine in the β configuration.
de Novo Purine Biosynthesis
Inosinate Formation
Generating AMP and GMP
Salvage Pathways Economize Intracellular
Energy Expenditure
Two salvage enzymes with different specificities recover
purine bases. Adenine phosphoribosyltransferase catalyzes
the formation of adenylate
whereas hypoxanthine-guanine
phosphoribosyltransferase (HGPRT) catalyzes the
formation of guanylate as well as inosinate
(inosine monophosphate, IMP), a precursor of
guanylate and adenylate
Pyrimidine Nucleotides Are Made from
Aspartate, PRPP, and Carbamoyl Phosphate
The common pyrimidine ribonucleotides are
cytidine 5-monophosphate (CMP; cytidylate) and
uridine 5-monophosphate (UMP; uridylate),
which contain the pyrimidines cytosine and
uracil. De novo pyrimidine nucleotide
Biosynthesis proceeds in a somewhat different
manner from purine nucleotide synthesis; the sixmembered pyrimidine ring is made first and then
attached to ribose 5-phosphate.
In the first step of the carbamoyl phosphate synthesis
pathway, bicarbonate is phosphorylated by ATP to form
carboxyphosphate and ADP. Ammonia then reacts with
carboxyphosphate to form carbamic acid and inorganic
phosphate.
Carbamoyl phosphate reacts with aspartate to form
carbamoylaspartate in a reaction catalyzed by aspartate
Transcarbamoylase . Carbamoylaspartate then cyclizes to
form dihydroorotate which is then oxidized by NAD+ to
form orotate.
Orotate reacts with PRPP to form orotidylate, a
pyrimidine nucleotide. This reaction is driven by
the hydrolysis of pyrophosphate. The enzyme
that catalyzes this addition, pyrimidine
phosphoribosyltransferase,
Pyrimidine synthesis
Carbamoyl phosphate reacts with aspartate to
yield N-carbamoylaspartate in the first
committed step of pyrimidine biosynthesis .
This reaction is catalyzed by aspartate
transcarbamoylase
By removal of water from
N-carbamoylaspartate, a reaction catalyzed
by dihydroorotase, the pyrimidine ring is
closed to form L-dihydroorotate.
This compound is oxidized to the pyrimidine
derivative orotate, a reaction in which NAD is
the ultimate electron acceptor.
Once orotate is formed, the ribose 5-phosphate
side chain, provided once again by PRPP, is
attached to yield orotidylate .
Orotidylate is then decarboxylated to uridylate,
which is phosphorylated to UTP. CTP is formed
from UTP by the action of cytidylate
synthetase.
de Novo Pathway for Pyrimidine Nucleotide
Synthesis.
The C-2 and N-3 atoms in the pyrimidine ring come from
carbamoyl phosphate, where as the other atoms of the ring come
from aspartate.
The pyrimidine ring is made up of three
components:
the nitrogen atom N-1 and carbons C-4 to C-6
are derived from aspartate, carbon C-2 comes
from HCO3-, and the second nitrogen (N-3) is
taken from the amide group of glutamine.
Degradation of Purines and Pyrimidines
Produces
Uric Acid and Urea, Respectively
Purine nucleotides are degraded by a pathway in
which they lose their phosphate through the action
of 5-Nucleotidase . Adenylate yields adenosine,
which is deaminated to inosine by adenosine
deaminase, and inosine is hydrolyzed to
hypoxanthine (its purine base) and D-ribose.
Hypoxanthine is oxidized successively to xanthine
and then uric acid by xanthine oxidase, a
flavoenzyme with an atom of molybdenum and
four iron-sulfur centers in its prosthetic group.
Molecular oxygen is the electron acceptor in this
complex reaction.
Purine Catabolism
Purine bases are converted first into xanthine and then into
urate for excretion.
Xanthine oxidase catalyzes two steps in this process.
Urate Crystals.
Micrograph of sodium urate crystals. Joints and kidneys
are damaged by these crystals in gout.
Uric acid is the excreted end product of purine
catabolism in primates, birds, and some other
animals. A healthy adult human excretes uric
acid at a rate of about 0.6 g/24 h; the excreted
product arises in part from ingested purines
and in part from turnover of the purine
nucleotides of nucleic acids. In most mammals
and many other vertebrates, uric acid is
further degraded to allantoin by the action of
urate oxidase.
Lesch-Nyhan syndrome
A genetic lack of hypoxanthine-guanine
phosphoribosyltransferase activity, seen almost
exclusively in male children, results in a bizarre set
of symptoms .
Children with this genetic disorder, which becomes
manifest by the age of 2 years, are sometimes
poorly coordinated and mentally retarded. In
addition, they are extremely hostile and show
compulsive self-destructive tendencies:
they mutilate themselves by biting off their fingers,
toes, and lips.
Excess Uric Acid Causes Gout
Long thought, erroneously, to be due to “high living,”
gout is a disease of the joints caused by an elevated
concentration of uric acid in the blood and tissues.
The joints become inflamed, painful, and arthritic,
owing to the abnormal deposition of sodium urate
crystals.
The kidneys are also affected, as excess uric acid is
deposited in the kidney tubules.
Gout occurs predominantly in males. Its precise cause
is not known, but it often involves an underexcretion
of urate. A genetic deficiency of one or another
enzyme of purine metabolism may also be a factor
in some cases.
Gout is effectively treated by a combination of
nutritional and drug therapies. Foods especially
rich in nucleotides and nucleic acids, such as liver
or glandular products, are withheld from the
diet. Major alleviation of the symptoms is
provided by the drug allopurinol , which inhibits
xanthine oxidase, the enzyme that catalyzes the
conversion of purines to uric acid.
multiple tophi on the hands (Panel
A), feet, knees, Some of the tophi
exuded a white, chalky material.
Laboratory studies were notable
for a serum uric acid level of 8.5
mg per deciliter (506 µmol per
liter), Xray hand:soft tissue
swelling and pararticular erosions
• The Gout-By James Gilray-1799
Johnson and Rideout NEJM, 350 (11): 1071, Figure 1
March 11, 2004
Control of Purine Biosynthesis.
Feedback inhibition controls both the overall rate of
purine biosynthesis and the balance between AMP and
GMP production
Many Chemotherapeutic Agents Target
Enzymes
in the Nucleotide Biosynthetic Pathways
The first set of agents includes compounds that
inhibit glutamine amidotransferases. Recall
that glutamine is a nitrogen donor in at least
half a dozen separate reactions in nucleotide
biosynthesis. The binding sites for glutamine
and the mechanism by which NH4 is
extracted are quite similar in many of these
enzymes. Most are strongly inhibited by
glutamine analogs such as azaserine and
acivicin
Several Valuable Anticancer Drugs Block the
Synthesis of Thymidylate
One inhibitor that acts on thymidylate synthase,
fluorouracil, is an important chemotherapeutic
agent. Fluorouracil itself is not the enzyme inhibitor.
In the cell, salvage pathways convert it to the
deoxynucleoside monophosphate FdUMP, which
binds to and inactivates the enzyme. Inhibition by
FdUMP is a classic example of mechanism-based
enzyme inactivation. Another prominent
chemotherapeutic agent, methotrexate, is an
inhibitor of dihydrofolate reductase. This folate
analog acts as a competitive inhibitor; the enzyme
binds methotrexate with about 100 times higher
affinity than dihydrofolate.
Methotrexate is a valuable drug in the treatment of
many rapidly growing tumors, such as those in acute
leukemia and choriocarcinoma, a cancer derived from
placental cells. However, methotrexate kills rapidly
replicating cells whether they are malignant or not.
Anticancer Drug Targets
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