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Transcript
Metabolism of Nucleotides
核 苷 酸 代 谢
Jiao Li
Department of biochemistry and molecular biology
65986142
[email protected]
Is nucleic acid dietarily essential?
Nucleic acid health product
What makes up of our DNA and RNA?
Learning Objectives
After completion of this section, you should be able to :
1. Indicate the source of each atom in purine and pyrimidine ring.
2. Describe anabolism (de novo and salvage) and
catabolism of purine and pyrimidine.
3. Outline the sequence of reactions in de novo and salvage
and the conversion of IMP to AMP or GMP, and UMP to
CTP or TMP.
4. Understand the mechanism of anticancer drugs and
allopurinol in treating gout.
Outline of the section
1. Nomenclature and structure of nucleotide.
2. Biosynthesis of purine and pyrimidine
3. Feedback inhibition and cross regulation in purine and
pyrimidine synthesis.
4. Catabolism of purine and pyrimidine.
5. Clinical significance.
Section 1
Nomenclature and structure of nucleotide
1. (Deoxy)Ribonucleotide is basic subunit of
DNA or RNA
(1) Nomenclature and structure of bases
purines
Adenine
Guanine
pyrimidines
Cytosine Uracil
Thymine
U is only present in RNA
T is present in DNA
These are purine and pyrimidine bases.
(2) Types of sugar
Ribose is only present in RNA.
Deoxyribose is only present in DNA.
(3) Deoxyribonucleotide in DNA
Base
Adenine
Guanine
Cytosine
Thymine
Deoxynucleoside
deoxyadenosine
deoxyguanosine
deoxythymidine
deoxycytidine
Deoxyribonucleotide
adenosine monophosphate
(AMP)
deoxyadenosine triphosphate
(dATP)
(4) Ribonucleotide in RNA
Base
Adenine
Guanine
Cytosine
Uracil
Nucleotide
adenosine
guanosine
Nucleoside
uridine
cytidine
Nucleotide
adenosine monophosphate
(AMP)
deoxyadenosine triphosphate
(dATP)
Section 2
Biosynthesis of purine and pyrimidine
Overview of dietary nucleotide
metabolism
Dietary nucleoprotein
protein
Gastric acid
Nuclei acid(RNA or DNA)
(deoxy)ribonuclease from pancreas
nucleotide
Nucleotidase from intestinal
endothelium
nucleoside
phosphate
nucleoside phosphorylase from
intestinal endothelium
Uric acid
base
inside intestinal endothelium
(deoxy)ribose
We do not need to eat
purines and
pyrimidines in our diet because we have ways
to make them.
Our body has two ways of making purine
and pyrimidine nucleotides:
De novo
synthesis and the salvage pathway.
New or recycled. We recycle more than we
make new.
1. De novo synthesis of purine nucleotide
•Source of nitrogen and carbon atoms of
the purine ring
CO2
glycine
Aspartate
N5,N10methenyl-
tetrahydrofolate
N10 formyl tetrahydrofolate
glutamine
(amide nitrogen)
• De novo synthesis of purine nucleotide
includes two major processes:
(1) Synthesis from PRPP to IMP
(2) Conversion of IMP to GMP and AMP.
• Major site: liver
• De novo synthesis of purine is weak in brain and
erythrocyte.
Overview of de novo pathway for synthesis of purine
Key step1
Key step2
IMP
(1) Synthesis from PRPP to IMP
1st step
Key enzyme
The 1st step in the pathway is the synthesis of
PRPP by PRPP synthetase. This step is regulated by
feedback inhibition.
Function of PRPP
PRPP is the source of the ribose sugar
and 5’ phosphate found in purine
nucleotides.
2nd step—— Base synthesis starts
PRPP amidotransferase catalyzes this step. This is
the first committed step in de novo synthesis of purines.
This step adds an amine from glutamine to PRPP.
3rd step
GAR
synthetase
The ribose sugar and
the phosphate are
indicated by P-Ribose
because this part does
not change in any of
the steps.
Glycine is the source of
2 carbons and a
nitrogen of the purine
ring.
Glycinamide ribosyl-5-phosphate
4th step
N5,N10- methenyl
tetrahydrofolate
Formyltransferase
Formylglycinamide
ribosyl-5-phosphate
In this step one carbon is donated by
N5,N10- methenyl tetrahydrofolate.
5th step
Synthetase
Formylglycinamidine
ribosyl-5-phosphate
In this step glutamine donates an amine.
6th step
ATP
The ring closes to make
the 5 membered ring.
Synthetase
ADP+Pi
Aminoimidazole ribosyl-5-phosphate
7th step
Carboxylase
Aminoimidazole carboxylate
ribosyl-5-phosphate
A carbon atom is donated by carbon dioxide.
8th step
synthetase
Aminoimidazole succinyl
carboxamide ribosyl-5-phosphate
Aspartate is added. Only the nitrogen of
aspartate will remain in the final purine ring.
9th step
Most of the aspartate
atoms are removed.
Only the nitrogen of
aspartate remains.
Adenylosuccinase
Aminoimidazole
carboxamide
ribosyl-5-phosphte
10th step
Formyltransferase
Formaminoimidazole carboxamide
ribosyl-5-phosphate
A carbon atom is donated by N10-formyltetrahydrofolate.
11th (last) step
Cyclohydrolase
In the last step the ring closes to make IMP.
You are
not
expected to memorize
pathways!
De novo synthesis of purine is on the
basis of PRPP by addition of simple
molecules step by step.
IMP is an important intermediate in de
novo synthesis of purine.
IMP
AMP
GMP
(2) Conversion of IMP to GMP and AMP
fumarate
Asp
Adenylosuccinate
① Adenylosuccinate synthetase
③ IMP dehydrogenase
② Adenylosuccinase
④ Transamidinase
2. Salvage pathway of purine synthesis
What are salvage pathways?
Salvage pathways recover bases and nucleosides by
converting them to nucleotides.
(1) Save energy
(2) Provide purines to erythrocyte and brain(no de novo)
How do salvage pathways proceed ?
(1) Base is converted to nucleotide
A to adenosine
monophosphate
G to guanosine
monophosphate
How do salvage pathways proceed ?
(2) Nucleoside is converted to nucleotide
Adenosine
(dAdenosine,
dGuanosine)
Adenosine kinase
ATP
ADP
AMP
(dAMP,
dGMP)
Relationship of nucleotides,
nucleosides, nucleobases
All compounds in this
box are nucleotides
Nucleobases in this box
Nucleosides
in this box
Case 1: Lesch Nyhan syndrome
HGPRT is the "salvage
enzyme" for the purines,
failure of this enzyme has two
results:
*Cell breakdown products
cannot be reused, and are
therefore degraded. This gives
rise to increased uric acid.
*The de novo pathway is
stimulated due to an excess of
PRPP.
X-linked
genetic disease
with:
Neurologic
disfunction;
behavial
disturbance;
high uric acid
Abnormal metabolism leads to disease
The CNS involvement is not caused
by uric acid. The brain has an
extremely low capacity for de novo
purine biosynthesis, so a complete
absence of the salvage enzyme
damages the brain by depriving it of
purine nucleotides.
3. De novo synthesis of pyrimidine
•Source of nitrogen and carbon atoms of
the pyrimidine ring
CO2
• De novo synthesis of pyrimidine nucleotide
includes two major processes:
(1) Synthesis from PRPP to UMP
(2) Conversion of UMP to CTP and TMP.
(1) Synthesis from PRPP to UMP
Key step
UMP
(2) Conversion of UMP to CTP and TMP
UMP
4. Salvage pathway of pyrimidine synthesis
Salvage pathway of pyrimidine
transferase
Pyrimidine + PRPP
nucleoside+ PPi
Uridine kinase
Uridine + ATP
UMP +ADP
Thymidine kinase
Thymidine + ATP
dTMP +ADP
Overview of synthesis for pyrimidine
Uridine
kinase
5. Formation of deoxyribonucleotides
by ribonucleotide reductase
Ribonucleotide reductase makes DNA
precursors
Ribonucleotides, the precursors
for RNA, are made first in the de
novo pathway.
Deoxyribonucleotides, the
precursors for DNA, are made later
and on the basis of NDP.
Ribonucleotide reductase reduces all 4
ribonucleotides to deoxyribonucleotides
NDP(A,C,G,U)
dNDP
kinase
dNDP + ATP
dNTP + ADP
Section 3
Feedback inhibition and cross regulation in purine and
pyrimidine synthesis
(1) Regulation of purine synthesis
When we speak of regulation of purine
synthesis what we mean is


(a) regulation of IMP synthesis, and
(b) regulation of synthesis of GMP and AMP
from IMP
α-D-Ribose 5-phosphate
PRPP
activation
Phosphoribosylamine
Feedback
inhibition
Feedback
inhibition
Cross-regulation
(2) Regulation of pyrimidine synthesis
PRPP(+)
In bacteria synthesis is activated
by ATP and inhibited by CTP.
In mammals synthesis is activated
by PRPP and inhibited by UTP.
UTP(-)
Setion 4
Catabolism of purine and pyrimidine
(1) Catabolism of purines
The end product of purine catabolism
in man and birds is uric acid.
Uric acid is formed primarily in the
liver and is excreted by the kidneys
into the urine.
(1) Catabolism of purines
Xanthine
oxidase
Guanine
deaminase
Adenosine
deaminase
Xanthine
oxidase
Nucleoside
phosporylase
Uric acid
(not very soluble)
Case 2:
Gout
Uric acid and urate salts are not
very soluble.
Crystals of sodium urate
precipitate in joints, especially the
big toe, causing severe pain.
Sodium urate crystals can damage
the kidneys, causing death.
3 out of 1000 people have gout.
Causes of gout
Overproduction of purine
nucleotides, leading to excessive
uric acid synthesis.
Inadequate salvage of purines.
Impaired uric acid excretion
through the kidneys.
Three enzymes defect
lead to gout
Catabolism of purines
Xanthine
oxidase
Guanine
deaminase
Adenosine
deaminase
Xanthine
oxidase
Nucleoside
phosporylase
Uric acid
(not very soluble)
Mechanism of gout
The unneeded excess purine
nucleotides are degraded to uric
acid. Uric acid is not very soluble.
The excess uric acid cannot stay
in solution. It precipitates,
causing gout.
Gout or death can be caused by
impaired uric acid secretion
Chemotherapy for cancer can overload
the kidneys with degraded purines after
the death of cells.
Kidney stones may form and lead to
death due to kidney failure.
Verify the dose of chemotherapy drugs
Treatment 1 of gout
Allopurinol inhibits
xanthine oxidase,
thus reducing the
amount of uric
acid made.
Limit intake of foods that are rich in nucleic
acids, for example meat. Limit alcohol intake
because alcohol affects kidney function.
Catabolism of purines
Xanthine
oxidase
(d)
Guanine
deaminase
Adenosine
deaminase
Xanthine
oxidase
Nucleoside
phosporylase
Uric acid
(not very soluble)
Gouty toe
After allopurinol
Treatment 2 of gout
Another way to treat
gout is with
probenecid, a drug that
increases renal
secretion of uric acid.
Probenecid
Probenecid
Uric acid
×
Colchicine
CH 3O
H
N
C
CH 3O
OCH 3
CH 3
O
O
OCH 3
COLCHICINE
Toxic natural product from Colchicum autumnale (Meadow Saffron).
Cochicine relieves the pain of gout. It is anti-inflammatory.
Catabolism of purines
Xanthine
oxidase
(d)
Guanine
deaminase
Adenosine
deaminase
Xanthine
oxidase
Nucleoside
phosporylase
Uric acid
(not very soluble)
Case 3: Combined immunodeficiency
Excess dATP caused by adenosine
deaminase deficiency inhibits
ribonucleotide reductase and
leads to immunodeficiency
disease.
Catabolism of purines
dATP
Xanthine
oxidase
(d)
Guanine
deaminase
Adenosine
deaminase
Xanthine
oxidase
Nucleoside
phosporylase
Adenosine deaminase deficiency
• Clinical characteristics: inactive
immune system and therefore inability
to fight infections. Death occurs in
early childhood.
When ribonucleotide reductase is inhibited
by dATP there are not enough nucleotides for
DNA synthesis. This makes it impossible for
white cells to proliferate rapidly. White cells
must proliferate for an immune response to
occur.
First
gene
therapy
disease
(2) Catabolism of pyrimidines
deaminase
Very water soluble
Section 5
Clinical significance
We have described several
diseases associated with purine
metabolism
Gout
Lesch-Nyhan syndrome
Combined immunodeficiency
disease
Some purine or pyrimidine analogs
are used as anticancer drugs
5-fluorouracil(5-FU)
6-mercaptopurine(6-MP)
Methotrexate
(1) 5-FdUMP inhibit thymidylate synthase
Breast cancer and colon cancer are
treated with 5-fluorouracil.
F
5-Fluorouracil
5-fluorouracil is an analog of thymine
F
5-fluorouracil
uracil
thymine
5-fluorouracil is an inactive precursor drug.
It must be converted to a nucleotide by the
salvage pathway before it is an active drug.
Transferase;
kinase
Active drug form
FdUMP inhibits thymidylate synthase.
As a result, DNA synthesis slows down.
FdUMP
Proliferation of cancer cells slows down.
(2) 6-MP inhibit salvage of purine
TIMP inhibit de novo synthesis of purine
(—)
hypoxanthine
(I)
HGPRT
6-mercaptopurine
(6-MP)
6-thioinosine-5-monophosphate
(TIMP) (—)
PRPP-glutamyl
amidotransferase
(3)Methotrexate inhibt dihydrofolate
reductase
Both compounds are similar and bind to dihydrofolate
reductase.
Dihydrofolate reductase activity is
essential for production of dTMP
Dihydrofolate
reductase
dTMP
Dihydrofolate reductase regenerates methylene FH4, the
donor of the CH3 group in dTMP.
Methotrexate
methotrexate
Methotrexate inhibits dihydrofolate reductase.
Cells cannot produce adequate amounts of dTMP
for DNA synthesis. In this way methotrexate
inhibits proliferation of cancer cells.
Summary
1. Source of each atom in purine and pyrimidine is from
simple molecule.
2. Tow major way of biosynthesis of purine and pyrimidine-
De novo and salvage pathway.
3. Feedback inhibition and cross regulation in purine and
pyrimidine synthesis.
4. End product of purine catabolism is uric acid.
5. Gout, Lesh-Nyhan syndrome and Combined
immunodeficiency syndrome are associated with
abnormal metabolism of purine.
Study question:
1. What is the mechanism of action of allopurinol for
treatment of gout?
a. Allopurinol solubilizes uric acid crystals by
binding to them.
b. Allopurinol increases excretion of uric acid by
the kidneys.
c. Allopurinol inhibits the enzyme that makes uric
acid.
d. Allopurinol decreases the inflammatory response.