Download MBBS Pyrimidine Lecture PPT

Lesch-Nyhan Syndrome

? Inability of the body to salvage hypoxanthine and

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guanine due to the complete deficiency of HGPRTase
(Hypoxanthine-Guanine phosphoribosyl transferase)

? It is an X-linked inherited disorder of purine

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metabolism, the disease is limited to males only

? Different types of mutations in HGPRTase gene have

been identified in patients with Lesch Nyhan

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syndrome.

? Incidence is 1:10,000 males.

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? HGPRT deficiency results in the accumulation of

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PRPP and decrease in GMP and IMP.

? Increased level of Hypoxanthine and Guanine

in degradation to uric acid

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? Also PRPP accumulates

stimulates production of Purine nucleotides

increases their degradation to uric acid

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? Leads to hyperuricemia---Gout-like symptoms

Nephrolithiasis ( Renal stones)

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Neurological symptoms

? self mutilation
? spasticity,
? aggressiveness,

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? mental retardation
DIAGNOSIS

? Increase urinary urate / creatinine ratio
? Absent / reduced enzyme activity in

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lymphocytes or fibroblast

? Mutation analysis of Hypoxanthine-Guanine

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phosphoribosyl transferase (HGPRT) gene.

Severe combined immunodeficiency (SCID)

? The deficiency of adenosine deaminase (ADA) causes

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severe

combined

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immunodeficiency

(SCID)

involving T-cell and usually B-cell dysfunction.

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? ADA deficiency results in the accumulation of dATP.
? dATP is an inhibitor of ribonucleotide reductase

which causes reduced synthesis of other dNTPs and

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therefore DNA synthesis and cell replication is
inhibited.

? Thus proliferation and differentiation of immune cells

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is compromised.



SCID

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? Lymphocytes usually contain high levels of ADA.
? Therefore, ADA deficiency is mainly manifested as

reduced lymphocytes.

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? This leads to impaired cellular and humoral immunity.
? Hypouricemia is due to defective breakdown of

purine nucleotides.

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ADA estimation in CSF is used for the diagnosis of
tuberculous meningitis.
ADA levels can be estimated in various body fluids like
blood, CSF, pleural fluid, pericardial fluid, ascitic fluid, etc.

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SCID - Treatment

? Antibiotics and periodic injections of

immunoglobulin will be lifesaving.

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? Bone marrow stem cells will increase both T

and B cells in the patients.

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? Enzyme replacement therapy with ADA-

Polyethylene glycol ( the first successful

application of enzyme replacement therapy for

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an inherited disease.

? Gene therapy- recently, ADA gene has been

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successfully transfected into stem cells of

ADA deficient children.

Purine Nucleoside Phophorylase

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Deficiency

? Less severe than ADA deficiency
? Associated with severe deficiency of T- cells

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but apparently normal B- cell function.

? Immune dysfunction appear to result from

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accumulation of dGTP, and dATP, which
inhibit ribonucleotide reductase and thereby
deplete cells of DNA precursors.

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METABOLISM OF

PYRIMIDINE

DR. SUDHANSHU SHEKHAR

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ASSOCIATE PROFESSOR

DEPT. OF BIOCHEMISTRY
Pyrimidine is a heterocyclic ring.

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4

? 5 5

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3

5

2

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1

6

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Sources of different atoms of pyrimidine rings
Synthesis of pyrimidine nucleotides

A. Denovo synthesis

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B. Salvage pathway

Denovo synthesis of

Formation

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of

pyrimidine nucleotide

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pyrimidine nucleotides

refers to the formation of

from pyrimidine bases

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pyrimidine ring structure
followed by the addition
of ribose phosphate
Denovo synthesis

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? The synthesis of pyrimidines is a much simpler

process compared to that of purines.

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? Aspartate, Glutamine and bicarbonate contribute to

atoms in the formation of pyrimidine ring.

? Pyrimidine ring is first synthesized and then attached

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to ribose 5-phosphate.

? This is in contrast to purine nucleotide synthesis

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where in purine ring is built upon a pre-existing
ribose-5-phosphate.
Synthesis of pyrimidine nucleotides

Tissue and site of synthesis

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Mainly occurs in the liver.

The reaction occurs in cytosol and mitochondria.

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The formation of orotate from dihydroorotate

occurs ie mitochondria and all other reactions

occur in the cytosol.

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Step 1: Carbamoyl Phosphate Synthesis

? The reaction occurs in

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cytoplasm

(in

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urea

synthesis, the reaction is

in mitochondria).

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? The

nitrogen

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of

glutamine, ATP and

bicarbonate react to form

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carbamoyl

phosphate

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(step 1).

? The enzyme is carbamoyl

phosphate synthetase II

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(CPS II). N


Step 2: Rate Limiting Step :Condensation

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? Carbamoyl phosphate and

aspartate combine to form

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carbamoyl aspartate

? The enzyme is aspartyl trans

carbamoylase (ATC), which

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is allosterically regulated


? The atoms C2 and N3 are

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derived from carbamoyl

phosphate and the rest are

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from aspartate.


Step 3: Formation of Pyrimidine Ring

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? The 3rd nitrogen and 4th

carbon are joined by a
covalent

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bond

and

carbamoyl aspartate is

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cyclized.

Ring closure:

? Dihydo orotic acid is

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produced.

? The enzyme is dihydro

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orotase (DHOase)


Step 4: Oxidation

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O

? Hydrogen

atoms

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are

removed from C5 and C6
positions, so that orotic

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acid is produced

? Enzyme is dihydro orotate

dehydrogenase (DHODH).

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? It requires NAD as co-

enzyme.

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Step 5:Transfer of ribose phosphate & Formation of OMP

? Ribose-5-phosphate

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is

added to orotic acid, so as to

?

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produce orotidylic acid or
orotidine

monophosphate

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(OMP).

? PRPP is the donor of ribose-

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5-P.

O

? The enzyme is orotate

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phosphoribosyl transferase
(OPRTase)

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Step 6: Decarboxylation

O

? The C7 of OMP is removed

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as carbon dioxide, so that

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uridine

monophosphate

(UMP) is produced

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? This is the first pyrimidine

that is synthesized.

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? The

enzyme

is

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OMP-

decarboxylase (OMPDC).

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? 6-aza-uridine inhibits this

step, and so used as an
anticancer drug

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Uridine monophosphate (UMP)


Step 7: Synthesis of

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Triphosphates

? UMP is phosphorylated to

form

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UDP

(uridine

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diphosphate) with the help of

ATP

? The enzyme is nucleoside

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monophosphatekinase (UMP

kinase).

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? The UDP is phosphorylated to

UTP (uridine triphosphate)

with the help of ATP

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? The enzyme is nucleoside

diphosphate kinase

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Step 8: Formation of CTP

? UTP is converted to

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CTP by adding an
amino group from
glutamine catalyzed

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by CTP synthetase.

? It needs ATP

Step 8

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Step 9 .Reduction of ribonucleoside diphosphates

to their corresponding dNDP's

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Step10.Formation of TMP

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from UDP

? dUMP is substrate for

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Step9

TMP synthesis.

? dUDP

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is

dephosphorylated

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to

dUMP

? Methylation of dUMP

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occurs at C5 by
N5,N10methyleneTHF,
forming TMP.

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? This reaction is catalysed

Step10

by Thymidylate synthase.

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SALVAGE PATHWAY OF PYRIMIDINE

SYNTHESIS

Pyrimidine base

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PRPP



Pyrimidine Phosphoribosyl

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Transferase


PPi
Pyrimidine nucleotide

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Regulation of pyrimidine synthesis

CPSII and Aspartate transcarbomylase are main

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regulatory enzymes.

There is feedback regulation to maintain optimal

pyrimidine nucleotide concentrations.

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CPS II ?
- inhibited by UTP .
- activated by PRPP
Aspartate transcarbomylase :

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- inhibited by CTP
- activated by ATP
Cross Regulation of Purine and pyrimidine

synthesis:

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? PRPP is required for the synthesis of both purines and

pyrimidines, so its regulation by both ensures
coordinated purine and pyrimidine synthesis

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? PRPP stimulates the purine and pyrimidine synthesis

through amidotransferase and carbamoyl phosphate
synthase respectively.

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? So both purine and pyrimidine feedback inhibit PRPP

synthase

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? Increase synthesis of pyrimidines (TDP) leads to

allosteric inhibition of PRPP synthase.

? Purine (ADP) also inhibit PRPP synthase.

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Degradation

of

pyrimidine

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nucleotides

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?The

pyrimidine

nucleotides

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undergo

similar

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reactions

(dephosphorylation, deamination
and cleavage of glycosidic bond) to
liberate the nitrogenous bases

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cytosine, uracil and thymine.

?The bases are then degraded to
highly soluble products -alanine
and -aminoisobutyrate.

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?These are the amino acid which
undergo transamination and other
reactions to finally produce acetyl
CoA and succinyl CoA

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DISORDERS OF PYRIMIDINE METABOLISM

1.OROTIC ACIDURIA

Features :

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? Orotic aciduria type I ?

? Due to lack of feedback

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deficiency of

inhibition orotic acid production

a) Orotate phosphoribosyl

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is excessive.(UMP inhibits

transferase and

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OMP decarboxylase)

b ) OMP ?decarboxylase.

Rapidly growing cells are

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? Orotic aciduria type II :

affected ?

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- Rare

a)

anemia

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- deficeincy of ONLY

b)

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Retarded growth

OMP decarboxylase.

c)

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Crystals excreted in urine

causing urinary obstruction.

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? Both types are inherited as

autosomal recessive disorders.

? Both types respond to uridine ,

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as it is converted to UTP . This

acts as feed back inhibitor
Other causes of orotic aciduria

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1.Deficeincy

of

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liver

mitochondrial

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ornthine

?

trancarbomylase (X-linked).

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under utilised substrate carbomyl phosphate enters cytosol

Stimulates pyrimidine nucleotide biosynthesis

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Leading to orotic aciduria
2. Drugs may precipitate orotic aciduria:

ALLOPURINOL , a purine analog is a substrate for

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Orotate phosphoribosyl transferase.


It competes for phosphoribosylation with natural substrate,

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orotic acid.

The resulting nucleotide product inhibits

OMP DECARBOXYLASE

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leading to Orotic aciduria and orotiduniria
Reye's syndrome

? This is considered as a secondary orotic

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aciduria.

? It is believed that a defect in ornithine

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trascarbamoylase (or urea cycle ) causes the
accumulation of carbamoyl phosphate.

? This is then diverted for the increased synthesis

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and excretion of orotic acid.
Anti-Folate Drugs

? Cancer cells consume dTMP quickly for DNA

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replication

? Interfere with thymidylate synthase reaction to

decrease dTMP production

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? Fluorodeoxyuridylate ? irreversible inhibitor ? also

affects rapidly growing normal cells (hair follicles,
bone marrow, immune system, intestinal mucosa)

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? Dihydrofolate reductase step can be stopped

competitively (DHF analogs)

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? Anti-Folates:

Aminopterin,

methotrexate,

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trimethoprim


Inhibitors

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5-Flurouracil and methotrexate