Upload
roshan-mahat
View
1.643
Download
4
Tags:
Embed Size (px)
Citation preview
Metabolism of Nucleotides
Guided by
Dr. D V Bhale
( prof. & HOD)
By
Roshan Kumar Mahat
(PG Student)
Department Of Biochemistry
Objectives to know
1. Nitrogen bases i.e. purines & pyrimidines
2. Nucleosides
3. Nucleotides
4. Synthesis of purine nucleotides
5. Regulation of purine nucleotides synthesis
6. Inhibitors of purine nucleotide synthesis
7. Disorders of purine metabolism
8. Synthesis of pyrimidine nucleotides
9. Inhibitors of pyrimidine nucleotides synthesis
10. Disorders of pyrimidine nucleotide synthesis
Pyrimidines and PurinesPyrimidine and purine are the names of the
parent compounds of two types of nitrogen-containing heterocyclic aromatic compounds.
N
N
N
N
N
NH
Pyrimidine Purine
Important Pyrimidines• Pyrimidines that occur in DNA are cytosine
and thymine. Cytosine and uracil are the pyrimidines in RNA. HN
NH
O
O
Uracil
HN
NH
O
O
CH3
Thymine
HN
NH
NH2
O
Cytosine
Important Purines• Adenine and guanine are the principal
purines of both DNA and RNA.
Adenine
N
N
NH2
N
NH
Guanine
O
HN
NH
N
N
H2N
Caffeine and Theobromine• Caffeine (coffee) and theobromine (coffee
and tea) are naturally occurring purines. Caffeine
N
N
O
N
N
H3C
O
CH3
CH3
Theobromine
O
HN
NN
N
CH3
CH3
O
Nucleosides
• Is a structure formed by the combination of nitrogen base and sugar.
N2 base Sugars Nucleoside
Adenine Deoxyribose/Ribose Adenosine
Guanine Deoxiribose/Ribose Guanosine
Thymine Deoxyribose Thymidine
Cytosine Deoxyribose/Ribose Cytidine
Uracil Ribose Uridine
Nucleotides
• Nucleotides are phosphoric acid esters of nucleosides.
Nucleoside Phosphoric acid Nucleotides
Adenosine Phosphoric acid Adenylate (AMP)
Guanosine Phosphoric acid Guanylate ( GMP)
Thymidine Phosphoric acid Thymidylate ( TMP)
Cytidine Phosphoric acid Cytidylate (CMP)
Uridine Phosphoric acid Uridylate (UMP)
Synthesis of purine nucleotides
Synthesis of purine nucleotidesDenovo synthesis
Synthesis of purine base step by step on the ribose 5-phosphate
Salvage pathwayAddition of ribose 5-phosphate to the preformed purine bases or
addition of phosphate to the purine nucleosides
Sources of different atoms of purine ring
Denovo synthesis of purine nucleotides
Tissue and site of synthesis
Tissues- major tissue is liver
Site- cytosol
Inhibitors
1. Sunfonamide Are structural analogues of PABA Act as competitive inhibitors of synthesis of folic
acid from PABA in bacteria. They inhibit the reactions of purine nucleotide
synthesis requiring folic acid ( GAR transformylase and AICAR transformylase)
Used as bacteriostatic drugs to control bacterial infection.
2. Methotrexate and AminopterinAre structural analogue of folic acid.They act as a competitive inhibitors of
dihydrofolate reductase thus blocking the biosynthesis of tetrahydrofolic acid.
They inhibit the reaction requiring folic acid for purine nucleotide synthesis.
Used in t/t of cancers like leukemia choriocarcinoma.
3. Trimethoprim structural analogue of folic acid.Acts as a competitive inhibitors of
dihydrofolate reductase in bacteria thus blocking the biosynthesis of tetrahydrofolic acid.
Inhibit the reaction requiring folic acid to purine nucleotide synthesis.
Used in the t/t of bacterial infections and UTI.
4. 6-mercaptopurine is a structural analogue of purine bases. is converted to 6- thioionosine
monophosphate by the enzyme HGPRT, called lethal synthesis.
6-thio IMP inhibits the conversion of IMP to AMP and GMP.
6-thio IMP also feed back inhibits glutamine PRPP amidotransferase.
Used as an anticancer drug.
5. Thioguanine Is a guanine analogue. It is converted to 6-thio GMP by the
enzyme HGPRT. 6-thio GMP inhibits the conversion of IMP
to GMP. Also inhibits glutamine PRPP
amidotransferase. Used as an anticancer drug.
6. Azaserine is a structural analogue of glutamine. is a glutamine antagonist. inhibits the enzyme reactions in purine and
pyrimidine nucleotide synthesis that utilize glutamine as a substrste.
it is highly toxic to the cells so it is not used clinically as a drug.
Regulation
1. Intracellular conc. Of PRPP-
depends upon 2 factors i.e. its synthesis & utilization.
Synthesis depends on- Availability of R-5-P. Action of enzyme PRPP synthetase.
Utilization depends on- Denovo synthesis. Salvage pathway.
2. Activity of enzyme PRPP amidotransferase.Increased activity of PRPP
amidotransferase leads to Increased synthesis of AMP amd GMP, which feedbackly inhibit the enzyme PRPP amidotransferase.
3. Both AMP and GMP inhibit their own formation by feedback inhibition of adenylosuccinate synthetase and IMP dehydrogenase.
Salvage pathway
It refers to the formation of purine nucleotides by the
1. Addition of ribose phosphate ( from PRPP) to the preformed purine bases.
2. Addition of phosphate to the preformed purine nucleosides.
Significance
Salvage pathway provide a pathway for the utilization of purine bases derived from diet (exogenous) and normal turnover of the nucleic acids.
In erythrocytes, denovo syntheis of purine nucleotides does not occur because of absence of PRPP amidotransferase. The requirement of purine nucleotides is met by the salvage pathway.
Synthesis of purine nucleotides from purine bases
Catalyzed by HGPRT and APRT.
Adenine + PRPP AMP + PPi
Hypoxanthine + PRPP IMP + PPi
Guanine + PRPP GMP+ PPi
APRT
HGPRT
HGPRT
Synthesis of purine nucleotides from purine nucleosides
Adenosine + ATP AMP+ ADP Adenosine kinase
Degradation of purine nucleotides
Disorders of purine metabolism
1. Gout
2. Lesch nyhan syndrome
3. Immunodeficiency associated with purine metabolism
4. Infantile autism
Gout
Metabolic disorders associated with overproduction of uric acid.
At physiological form, uric acid is found in more soluble form as sodium urate.
In severe hyperuricemia, crystal of sodium urate get deposited in the soft tissues, particularly in joints. Such deposits are commonly known as tophi.
This causes inflammation of joints resulting in gouty arthritis.
The prevalence of gout is about 3 per 1000 persons, mostly affecting males.
Post menopausal women, however are as susceptable as men for this disease.
Historically, gout was found to be associated with high living, over eating and alcohol consumption.
Lead poisoning also causes gout by decreasing uric acid excretion.
Types of gout
Gout
Primary gout
Metabolic gout
Renal gout
Secondary gout
Metabolic gout
Renal gout
Primary metabolic goutIt is an inborn error of purine metabolism
due to overproduction of uric acid.
Causes:
1. Increased activity of PRPP synthetase
2. Overactivity of PRPP amidotransferase
3. HGPRT deficiency
4. Glucose 6-phosphatase deficiency
5. Elevation of glutathione reductase
Primary renal gout
It is due to failure of uric acid excretion from the body so that uric acid level in the body gets increased.
Secondary metabolic gout
Secondary gout is due to secondary to certain diseases like leukemia, polycythemia, lymphoma, psoriasis and increased tissue breakdown like in trauma, starvation etc.
Secondary renal gout
It is due to secondary to defective glomerular filtration of urate due to generalized renal failure.
Tratment of gout
Is by
1. Use of colchicine & uricosuric drugs. To remove urates from the joint, colchine
is the drug of choice. To remove the urates from the body,
urocosuric drugs such as probenecid, sulfinpyrazole, salicylates etc are used.
2. Use of allopurinol-inhibits the activity of enzyme xanthine oxidase as a result of which uric acid is not produced.
Lesch Nyhan syndromeFist described tn 1964 by Michael Lesch( a
medical student) and William L. Nyhan (his teacher).
It is X linked metabolic disorder since the structural genes for HGPRT is located on the X chromosome.
It affects only males and is characterized by excessive uric acid production and neurological abnormalities such as mental retardation, aggressive behaviour, learning disability etc.
The patients of this disorder have an irresistible urge to bite their fingers and lips,ofen causing self-mutilation.
Biochemical basisHGPRT deficiency spares the utilization of
PRPP through salvage and the accumulated PRPP takes part in the purine biosynthesis by the denovo pathway finally leading to hyperuricemia.
The biochemical basis for neurological abnormalities are big enegma till date.
Indeed, it is surprising that the deficiency of a single enzyme can cause such an abnormal behavioural changes.
few explanations are putforth in this regard.
Neurological symptoms may be due to decreased availability of purines to the developing brain which has a low capacity for denovo purine synthesis and hence depends on purine salvage pathway for the supply of purine nucleotides it requires.
Treatment
allopurinol is used to treat hyperuricemia but it has no effect on the neurological menefestation in theses patients.
Treatment for the neuro-behavioural features are limited to behavioural therapy and providing protective physical device to prevet self-mutilation.
Immunodeficiency diseases associated with purine metabolism
Two different immunodeficiency disorders associated with degredation of purine nucleotides are known.
The enzyme defects are adenosine deaminase and purine nucleoside phosphorylase, involved in uric acid synthesis.
The deficiency of ADA causes SCID involving T- cell and usually B- cell dysfunction.
It is explained that ADA deficiency results in the accumulation of Datp which is an inhibitor of ribonucleotide reductase and thus DNA synthesis, replication are adversely affected.
Different modes of t/t such as blood transfusion, bone marrow transplantation are tried to cure the diseases but with limited effects.
But, like in any other inborn error, the real hope for the future is only gene therapy.
In 1990, a 5 year old girl suffering from SCID was successfully cured by transfecting the ADA gene into stem cells of the patients.
This is considered as landmark in the history of trating inborn errors of metabolism.
The deficiency of purine nucleoside phosphorylase is associated with impairement of T cell function but has no effect on B cell function.
It is believed that d GTP inhibits the development of normal T-cells.
Infantile Autism
Recently it was observed that children suffering from infantile autism exihibited increased excretion of uric acid but surprisingly the serum concentrations are within normal limits.
The biochemical basis for this is unknown.An oral dose of uridine is tried in the t/t.
Synthesis of pyrimidine nucleotides
Synthesis of pyrimidine nucleotides
Denovo synthesisSynthesis of pyrimidine nucleotide
refers to the formation of pyrimidine ring structure followed
by the addition of ribose phosphate.
Salvage pathwayFormation of pyrimidine
nucleotides from pyrimidine bases
Sources of different atoms of prrimidine rings
Denovo synthesis of pyrimidine nucleotides
Tissue and site of synthesisMainly occurs in the liver.The reaction occurs in cytosol and
mitochondria. The formation of orotate from dihydroorotate occurs ie mitochondria and all other reactions occur in the cytosol.
2 ATP + HCO3- + Glutamine + H2O
CO
O PO3-2
NH2
Carbamoyl Phosphate
NH2
CNH
CH
CH2
C
COOO
HO
O
Carbamoyl Aspartate
HN
CNH
CH
CH2
C
COOO
O
Dihydroorotate
HN
CNH
C
CHC
COOO
O
Orotate
HN
CN
C
CHC
COOO
O
HH
CH2
OH OH
H HO
O2-O3P
Orotidine-5'-monophosphate(OMP)
HN
CN
CH
CHC
O
O
HH
CH2
OH OH
H HO
O2-O3P
Uridine Monophosphate(UMP)
2 ADP +Glutamate + Pi
CarbamoylPhosphateSynthetase II
AspartateTranscarbamoylase(ATCase)
Aspartate
Pi
H2O
Dihydroorotase
Quinone
ReducedQuinone
DihydroorotateDehydrogenase
PRPP PPi
Orotate PhosphoribosylTransferase
CO2
OMP Decarboxylase
Pyrimidine Synthesis
UMP UTP and CTP
• Nucleoside monophosphate kinase catalyzes transfer of Pi to UMP to form UDP; nucleoside diphosphate kinase catalyzes transfer of Pi from ATP to UDP to form UTP
• CTP formed from UTP via CTP Synthetase driven by ATP hydrolysis – Glutamine provides amide nitrogen for C4 in
animals
UDP dUDP
dUDP dUMP
dUMP dTMP
N5,10 formyl THF formyl THF
Ribonucleotide reductase
Thymidylate synthetase
Regulation of pyrimidine synthesis
In bacteria, aspartate transcarbamoylase catalyses a committed step in pyrimidine biosynthesis.
Aspartate transcarbamoylase is a good example of an enzyme controlled by feedback mechanism by the end product CTP.
In certain bacteria, UTP also inhibits aspartate transcarbamoylase. ATP, however stimulates aspartate transcarbamoylase activity.
Carbamoyl phosphate synthase II is the regulatory enzyme of pyrimidine synthesis in animals.
It is activated by PRPP and ATP and inhibited by UDP and UTP.
OMP decarboxylase inhibited by UMP and CMP, also controls pyrimidine formation.
Inhibitors of pyrimidine synthesis
SulfonamidesMethotrexateTrimethoprim5-fluorouracilFluorocytosine
Salvage pathwaySalvage pathway of pyrimidine nucleotide
synthesis refers to the formation of pyrimidine nucleotides from pyrimidine bases.
Significance Salvage pathway provide a pathway for the
utilization of pyrimidine bases derived from diet(exogenous) and normal turnover of nucleic acids.
Enzymes and reactions
There are 2 enzymes that catalyze the reactions of salvage pathway. They are uracil phosphoribosyl transferase (UPRT) and thymidine kinase.
Uracil + PRPP UMP + PPi
Thymidine + ATP TMP+ ADP
UPRT
Thymidine kinase
Disorders of pyrimidine metabolism
Disorders of pyrimidine metabolism includes:Orotic aciduriaReye’s syndrome
Orotic aciduriaIs a rare metabolic disorder characterized by the
excretion of orotic acid in urine, severe anemia and retarded growth.
It is due to the deficiency of the enzymes orotate phosphoribosyl transferase and OMP decarboxylase of pyrimidine synthesis.
Both these enzymes activities are present on a single protein as domains (bifunctional enzyme).
Treatment Feeding diet rich in uridine or cytidine is an
effective t/t of orotic aciduria.These compounds provide pyrimidine
nucleotides required for DNA and RNA synthesis.
Reye’s syndrome
Is considered as a secondary orotic aciduria.It is believed that a defect in ornithine
transcarbamoylase (of urea cycle) causes the accumulation of carbamoyl phosphate.
This is then diverted for the increased synthesis and excretion of orotic acid.
THANK YOU