Metabolism of nucleoproteins.

Metabolism of nucleotides
Olga Tagadiuc
 Digestion of nucleoproteins
Digestion in the stomach

 Human gastric juice is able to digest

nucleoproteins
 The HCl of the gastric juice is denaturing the
proteic part and the nucleic acid to individual
linear strands
 Pepsin digests the proteic part of the
nucleoprotein
 Digestion of nucleoproteins
Digestion in the intestine

 Nucleic acids enter into the small intestine from the

stomach dissolved in the gastric chyme
 Enzymes involved in the digestion of the polynucleotides
in the small intestine:
 endonucleases
 exonucleases
• As chyme enters the duodenum 2 enzymes are secreted:
 Ribonuclease (RNAse)
 Deoxyribonuclease (DNAse)
Digestion of nucleoproteins
Digestion in the intestine
1. Ribonuclease: catalyzes the
breakdown of RNA into
ribonucleotides.
2. Deoxyribonuclease: catalyzes
the breakdown of DNA into
deoxyribonucleotides.
3. The ribo- and
deoxyribonucleotides
produced are still too large
and are required to be broken
down into smaller
components until they can be
absorbed in the small
intestine.
 Digestion of
nucleoproteins
Digestion in the
intestine

 Further digestion occurs at the microvilli in

the small intestine bu brush border enzymes
 Two enzymes are responsible:

1. Phosphatases - catalyze the cleavage of a

phosphate group from a nucleotide to form a
nucleoside and a phosphate ion.
2. Nucleosidases - catalyze the breaking of the
N-glycosidic bond between the nitrogenous
base and the pentose sugar of a nucleoside
Digestion
of
nucleic
acids
 Absorption of the end products
of nucleic acids digestion

Absorption of the end products of nucleic acids

digestion mainly occurs in the intestinal villus
of the duodenum and jejunum.
Are absorbed:
 nitrogenous bases
 pentose sugar
 phosphate ions
 Absorption of the end products
of nucleic acids digestion

• Membrane transport proteins carry the products of

nucleotide digestion into epithelial cells from the
intestine lumen.
• Transport mechanism - active transport
• Through diffusion, the products of nucleotide digestion
are transported from the intestinal epithelial cells:
 across the basolateral membrane →
 into the interstitial fluid →
 into the blood capillaries of the intestinal villi.
 Blood transport of the end products
of nucleic acids digestion and absorption

 The nucleotide digestion products are transported by

blood circulation to the liver and other tissues where
they undergo further degradation.
 Uric acid which is the end product of purine
degradation exist as sodium ureate in plasma.
 Maximum amount of sodium ureate that can dissolve
in the blood plasma is about 7 mg/100 ml. At this point
there will be saturation of blood with sodium ureate.
Metabolism of nucleotides
 Requirements for the nucleotides

The metabolic requirements for the nucleotides

or nitrogenous bases can be met by:
 synthesis de novo from low molecular weight
precursors
 salvage of nitrogenous bases
 Biosynthesis of Nucleotides

Biosynthesis of both – purine and pyrimidine

nucleotides, requires ribose-5-phosphate in activated
form – phosphoribosyl-pyrophosphate (PRPP).
Source of ribose-5phosphate – pentose-phosphate
shuttle
 Biosynthesis of Nucleotides
Biosynthesis of phosphoribosyl-pyrophosphate (PRPP)
from ribose-5phosphate

 Rate-limiting reaction
 Enzyme feed-back inhibition by purine nucleoside
diphosphates – ADP and GDP
 Activation by Pi
De novo Biosynthesis of Purine Nucleotides
Determined by John M. Buchanan using isotopic
tracer experiments in birds (1948).
De novo Biosynthesis of Purine Nucleotides
 De novo Biosynthesis of Purine Nucleotides

 The anomeric carbon of the substrate – PRPP, is in the alpha-

configuration; the product is a beta-glycoside.
 The N atom of this N-glycoside becomes N9 of the purine ring. It
is the first atom added in the construction of this ring.
 Glutamine PRPP amidotransferase is subject to feedback
inhibition by binding ATP, ADP and AMP at one inhibitory site
and GTP, GDP and GMP at another.
 The enzyme is stimulated by substrate – PRPP.
 De novo Biosynthesis of Purine
Nucleotides

A sequence of 9 reactions.
 IMP - the branch point for purine biosynthesis, because it
can be converted into either AMP or GMP through two
distinct reaction pathways.
 The pathway leading to AMP requires energy in the form
of GTP; that leading to GMP requires energy in the form of
ATP.
Biosynthesis of AMP from IMP

l. adenylosuccinate synthetase
2. adenylosuccinate lyase
Biosynthesis of GMP from IMP

1. IMP dehydrogenase
2. XMP-glutamine amidotransferase
 Regulation
of Purine Nucleotide
Biosynthesis
 Purine nucleotide synthesis
is regulated by energy status,
since the process requiers
many ATP mol.
 Formation of PRPP from
ribose-5-phosphate by PRPP
synthetase is inhibited by
ADP and GDP as indicators
of poor energy status.
 Regulation
of Purine Nucleotide
Biosynthesis
 PRPP amido
transferase which
produces 5-phospho-
ribosylamine.
 All phosphorylated
purines - AMP, ADP, ATP,
GMP, GDP, GTP, are
allosteric inhibitors of
the enzyme, controlling
overall purine synthesis
versus other cellular
activity.
 Regulation
of Purine Nucleotide
Biosynthesis
 The branches leading from IMP to
AMP and GMP are reciprocally
regulated: AMP inhibits
adenylosuccinate synthase, while
GMP inhibits IMP dehydrogenase.
 In addition, ATP activates
(chemically) the amido-transfer
producing GMP, and GTP
activates (chemically) the the
adenylosuccinate synthase
producing AMP.
 This regulation tends to
balance the relative level of the
two purines.
 Salvage pathway
or recycling of preformed bases

The genetic lack of hypoxanthine-guanine

phosphoribosyltransferase activity results in
Lesch-Nyhan syndrome.
 Lesch-Nyhan syndrome

 Michael Lesch and William Nyhan provided the

first detailed clinical description of Lesch-Nyhan
disease in 1964.
 The enzymatic defect associated with Lesch-Nyhan
disease, deficiency of the enzyme hypoxanthine-
guanine phosphoribosyl transferase (HPRT), was
discovered by Seegmiller and colleagues in 1967.
 The gene encoding the human enzyme was cloned
and sequenced by Friedmann and colleagues in
1985.
 Lesch-Nyhan syndrome
1. Cause - the genetic lack of hypoxanthine-guanine
phospho-ribosyl transferase (HGPRT)activity.
2. In the absence of HPRT
– hypoxanthine and guanine cannot be salvaged;
– they are degraded and excreted as uric acid.
3. The synthetic rate for purines is accelerated to
compensate for purines lost by the failure of the salvage
process.
4. The failure of recycling together with the increased
synthesis of purines lead to the overproduction of uric
acid → hyperuricemia.
Lesch-Nyhan syndrome

3 major clinical elements:

– overproduction of uric acid,
– neurologic disability
(dominated by dystonia),
– mental retardation and
behavioral problems
(aggressive and impulsive
behaviors)
Purine catabolism
 Gout

Gout is a disorder that is related to excess

production and deposition of uric acid crystals.
Cause:
 PRPP synthetase hyperactivity
 HGPRPP transferase deficiency
Signs: elevated concentration of uric acid in the
blood and tissues
 Gout simptoms
The joints become inflamed, painful, and arthritic, due
to the abnormal deposition of crystals of sodium urate.
 Gout simptoms
The kidneys are affected, because excess uric acid
is deposited in the kidney tubules.
 Gout treatment

 Diet without foods rich in nucleotides and nucleic

acids, such as liver or glandular products
 Allopurinol – inhibitor of xanthine oxidase, the
enzyme responsible for converting purines into
uric acid
 Gout treatment
Allopurinol Uric acid

 Suicidal inhibition of xanthine oxidase →

 the excreted products of purine metabolism
are xanthine and hypoxanthine, which are
more soluble in water than uric acid →
 less likely to form crystalline deposits.
 Gout treatment

Allopurinol was developed by Gertrude Elion

and George Hitchings, who also developed
acyclovir, used to treat AIDS, and other purine
analogs used in cancer chemotherapy.
 De novo biosynthesis
of pyrimidine nucleotides
 De novo biosynthesis of pyrimidine nucleotides

 Carbamoyl phosphate utilized in pyrimidine nucleotide

synthesis differs from that synthesized in the urea cycle - it is
synthesized from glutamine instead of ammonia and is
synthesized in the cytosol.
 The reaction is catalyzed by the carbamoyl phosphate
synthetase II – CPS-II
 The CPS-II is activated by ATP and inhibited by UDP, UTP,
dUTP, and CTP.
 De novo biosynthesis
of pyrimidine nucleotides
1. E – aspartate
transcarbamoylase (ATCase)
2. ATCase controls the rate of
pyrimidine biosynthesis
3. regulators: cellular levels of
both pyrimidines and purines
 CTP and/or UTP decreases the
catalytic activity
 ATP stimulates the catalytic
activity
 De novo
biosynthesis
of pyrimidine
nucleotides
 De novo
biosynthesis
of pyrimidine
nucleotides

In all higher
eukaryotes DHODH:
 located in the inner
mitochondrial
membrane
 prosthetic group flavin
and ubichinone
De novo biosynthesis of pyrimidine nucleotides

 The decarboxylase activity domain is

competitively inhibited by UMP and by CMP
 De novo biosynthesis
of pyrimidine nucleotides

 UMP is phosphorylated to UTP

 UTP is utilized for the synthesis of CTP
 Uridine nucleotides are the precursors for de
novo synthesis of the thymine nucleotides
 UMP phosphorylation to UTP

 E1 – uridylate kinase
 E2 – nucleoside diphosphate kinase
 Synthesis of CTP from UTP

 UTP is aminated by the action of CTP synthase

 CTP synthase is feedback-inhibited by CTP and
activated by GTP.
Synthesis of the Thymine Nucleotides

The de novo pathway to dTTP synthesis first

requires the synthesis of dUMP
Synthesis of the Thymine Nucleotides

The de novo pathway to dTTP synthesis first

requires the synthesis of dUMP
Synthesis of the Thymine Nucleotides
Synthesis of the Thymine Nucleotides

 The unique property of the action of

thymidylate synthase is that the THF is
converted to dihydrofolate (DHF)
 THF is regenerated from DHF through the
action of dihydrofolate reductase (DHFR)
 THF is converted to N5,N10-THF via the action
of serine hydroxymethyl transferase
 Salvaging Pyrimidines
 The major salvage pathway for the pyrimidines –
uracil and thymine:
 Base + Ribose-1-phosphate → Nucleoside + Pi
E – nucleoside phosphorylase
 Nucleoside + ATP - Nucleotide + ADP
E – nucleoside kinase
 The salvage pathway of the thymidine nucleotide
synthesis is especially important in the preparation
for cell division.
 The activity of thymidine kinase fluctuates with the
cell cycle, rising to peak activity during the phase of
DNA synthesis; it is inhibited by dTTP.
Salvaging Pyrimidines
Catabolism of
pyrimidine
nucleotides