Download Nucleic Acid metabolism De Novo Synthesis of Purine

Nucleic
Acid
metabolism
• De Novo Synthesis of Purine Nucleotides
• We use for purine nucleotides the entire glycine
molecule (atoms 4, 5,7), the amino nitrogen of
aspartate (atom 1), amide nitrogen of
glutamine (atoms 3, 9), components of the
folate-one-carbon pool(atoms 2, 8), carbon
dioxide, ribose 5-P from glucose and a great
deal of energy in the form of ATP. In de novo
synthesis, IMP is the first nucleotide formed. It
is then converted to either AMP or GMP.
• PRPP
• Since the purines are synthesized as the
ribonucleotides, (not as the free bases) a
necessary prerequisite is the synthesis of the
activated form of ribose 5-phosphate. Ribose
5-phosphate reacts with ATP to form 5Phosphoribosyl-1-pyrophosphate (PRPP).
• This reaction occurs in many tissues because
PRPP has a number of roles - purine and
pyrimidine nucleotide synthesis, salvage
pathways, NAD and NADP formation. The
enzyme is heavily controlled by a variety of
compounds (di- and tri-phosphates, 2,3-DPG),
presumably to try to match the synthesis of
PRPP to a need for the products in which it
ultimately appears
• Commitment Step
• De novo purine nucleotide synthesis occurs
actively in the cytosol of the liver where all of
the necessary enzymes are present as a macromolecular aggregate. The first step is a
replacement of the pyrophosphate of PRPP by
the amide group of glutamine. The product of
this reaction is 5-Phosphoribosylamine. The
amine group that has been placed on carbon 1
of the sugar becomes nitrogen 9 of the ultimate
purine ring. This is the commitment and ratelimiting step of the pathway
• Control of De Novo Synthesis
• Control of purine nucleotide synthesis has two
phases. Control of the synthesis as a whole
occurs at the amidotransferase step by
nucleotide inhibition and/or [PRPP]. The second
phase of control is involved with maintaining
an appropriate balance (not equality) between
ATP and GTP. Each one stimulates the synthesis
of the other by providing the energy. Feedback
inhibition also controls the branched portion as
GMP inhibits the conversion of IMP to XMP and
AMP inhibits the conversion of IMP to
adenylosuccinate.
• De Novo Synthesis of Pyrimidine Nucleotides
• Since pyrimidine molecules are simpler than purines, so is their
synthesis simpler but is still from readily available components.
Glutamine's amide nitrogen and carbon dioxide provide atoms 2
and 3 or the pyrimidine ring. They do so, however, after first
being converted to carbamoyl phosphate. The other four atoms
of the ring are supplied by aspartate. As is true with purine
nucleotides, the sugar phosphate portion of the molecule is
supplied by PRPP.
• Carbamoyl Phosphate
• Pyrimidine synthesis begins with carbamoyl phosphate
synthesized in the cytosol of those tissues capable of making
pyrimidines (highest in spleen, thymus, GItract and testes). This
uses a different enzyme than the one involved in urea synthesis.
Carbamoyl phosphate synthetase II (CPS II) prefers glutamine
to free ammonia and has no requirement for N-Acetylglutamate
• Formation of Orotic Acid
• Carbamoyl phosphate condenses with aspartate in
the presence of aspartate transcarbamylase to
yield N-carbamylaspartate which is then converted
to dihydroorotate.
• In man, CPSII, asp-transcarbamylase, and
dihydroorotase activities are part of a
multifunctional protein.
• Oxidation of the ring by a complex, poorly
understood enzyme produces the free pyrimidine,
orotic acid. This enzyme is located on the outer face
of the inner mitochondrial membrane, in contrast to
the other enzymes which are cytosolic. Note the
contrast with purine synthesis in which a nucleotide
is formed first while pyrimidines are first
synthesized as the free base.
• Salvaging Purines
As a salvage process though, we are dealing with
purines. There are two enzymes, A-PRT and HGPRT. A-PRT is not very important because we
generate very little adenine. (Remember that the
catabolism of adenine nucleotides and nucleosides
is through inosine). HG-PRT, though, is
exceptionally important and it is inhibited by both
IMP and GMP. This enzyme salvages guanine
directly and adenine indirectly. Remember that
AMP is generated primarily from IMP, not from
free adenine
• Lesch-Nyhan Syndrome
• HG-PRT is deficient in the disease called
Lesch-Nyhan Syndrome, a severe
neurological disorder whose most blatant
clinical manifestation is an uncontrollable
self-mutilation. Lesch-Nyhan patients have
very high blood uric acid levels because of
an essentially uncontrolled de novo
synthesis. (It can be as much as 20 times the
normal rate). There is a significant increase
in PRPP levels in various cells and an inability
to maintain levels of IMP and GMP via
salvage pathways. Both of these factors
could lead to an increase in the activity of
the amidotransferase.
• Purine Catabolism
•
The end product of purine catabolism in
man is uric acid. Other mammals have the
enzyme urate oxidase and excrete the
more soluble allantoin as the end product.
Man does not have this enzyme so urate is
the end product for us. Uric acid is formed
primarily in the liver and excreted by the
kidney into the urine.
• Bases to Uric Acid
• Both adenine and guanine nucleotides converge at the
common intermediate xanthine. Hypoxanthine,
representing the original adenine, is oxidized to xanthine
by the enzyme xanthine oxidase. Guanine is deaminated,
with the amino group released as ammonia, to xanthine.
If this process is occurring in tissues other than liver, most
of the ammonia will be transported to the liver as
glutamine for ultimate excretion as urea.
• Xanthine, like hypoxanthine, is oxidized by oxygen and
xanthine oxidase with the production of hydrogen
peroxide. In man, the urate is excreted and the hydrogen
peroxide is degraded by catalase. Xanthine oxidase is
present in significant concentration only in liver and
intestine. The pathway to the nucleosides, possibly to the
free bases, is present in many tissues.
• Gouts and Hyperuricemia
• Both undissociated uric acid and the monosodium salt
(primary form in blood) are only sparingly soluble. The
limited solubility is not ordinarily a problem in urine unless
the urine is very acid or has high [Ca2+]. [Urate salts
coprecipitate with calcium salts and can form stones in
kidney or bladder.] A very high concentration of urate in the
blood leads to a fairly common group of diseases referred to
as gout. The incidence of gout in this country is about
3/1000.
• Gout is a group of pathological conditions associated with
markedly elevated levels of urate in the blood (3-7 mg/dl
normal). Hyperuricemia is not always symptomatic, but, in
certain individuals, something triggers the deposition of
sodium urate crystals in joints and tissues. In addition to the
extreme pain accompanying acute attacks, repeated attacks
lead to destruction of tissues and severe arthritic-like
malformations. The term gout should be restricted to
hyperuricemia with the presence of these tophaceous
deposits.
• Urate in the blood could accumulate either through an
overproduction and/or an underexcretion of uric acid. In
gouts caused by an overproduction of uric acid, the defects
are in the control mechanisms governing the production of
- not uric acid itself - but of the nucleotide precursors. The
only major control of urate production that we know so
far is the availability of substrates (nucleotides,
nucleosides or free bases).
• One approach to the treatment of gout is the drug
allopurinol, an isomer of hypoxanthine.
• Allopurinol is a substrate for xanthine oxidase, but the
product binds so tightly that the enzyme is now unable to
oxidized its normal substrate. Uric acid production is
diminished and xanthine and hypoxanthine levels in the
blood rise. These are more soluble than urate and are less
likely to deposit as crystals in the joints. Another approach
is to stimulate the secretion of urate in the urine.
• Pyrimidine Catabolism
• In contrast to purines, pyrimidines undergo ring cleavage
and the usual end products of catabolism are beta-amino
acids plus ammonia and carbon dioxide. Pyrimidines
from nucleic acids or the energy pool are acted upon by
nucleotidases and pyrimidine nucleoside phosphorylase
to yield the free bases. The 4-amino group of both
cytosine and 5-methyl cytosine is released as ammonia.
• Ring Cleavage
• In order for the rings to be cleaved, they must first be
reduced by NADPH. Atoms 2 and 3 of both rings are
released as ammonia and carbon dioxide. The rest of the
ring is left as a beta-amino acid. Beta-amino isobutyrate
from thymine or 5-methyl cytosine is largely excreted.
Beta-alanine from cytosine or uracil may either be
excreted or incorporated into the brain and muscle
dipeptides, carnosine (his-beta-ala) or anserine (methyl
his-beta-ala).