Nucleotide Metabolism Pyrimidine - PowerPoint Presentation

Slide1

Nucleotide Metabolism

Pyrimidine

Met.

Purine

Met.

Slide2

Learning Objectives

1. How

Are Purines Synthesized?

2.

How Are Purines Catabolized?

3.

How Are

Pyrimidines

Synthesized and

Catabolized

?

 

4.

How Are

Ribonucleotides

Converted to

Deoxyribonucleotides

?

 

5. How

Is

dUTP

Converted to

dTTP

?

6

.

Abnormal metabolism of uric acid.

7. Anticancer drugs targets

Slide3

Two types of pathways lead to nucleotides: the

de novo pathways and the salvage pathways.

De novo

synthesis of nucleotides begins with their metabolic precursors:

amino acids

,

ribose 5-phosphate

,

CO

2

, and

NH

3

.

Salvage pathways

recycle the free bases and nucleosides released from nucleic acid breakdown.

Slide4

Salvage and de Novo Pathways

Slide5

The purine ring structure is built up one or a few atoms at a time, attached to

ribose

throughout the process.

The

pyrimidine

ring is synthesized as

orotate

, attached to

ribose phosphate

, and then converted to the common

pyrimidine

nucleotides required in nucleic acid synthesis.

Although the free bases are not intermediates in the de novo pathways, they are intermediates in some of the salvage pathways.

Slide6

de Novo Pathway for Purine Nucleotide Synthesis

Slide7

The synthesis of the purine ring is more

complex. The only major component is

glycine

,

which

donates C-4 and C-5, as well as N-7

. All of the other atoms in the ring are incorporated individually. C-6 comes from HCO3–. Amide groups from

glutamine provide

the atoms N-3 and N-9. The amino group donor for the inclusion of N-1 is

aspartate

,

which is converted into

fumarate

in the process, in the same way as in the urea cycle . Finally, the carbon atoms C-2 and C-8 are derived from

formyl

groups in N10-

formyl-tetrahydrofolate

Slide8

The Purine Ring System Is Assembled on Ribose Phosphate

Glutamine

phosphoribosyl

amidotransferase

catalyzes this reaction.

Slide9

De novo

purine

biosynthesis, like

pyrimidine

biosynthesis, requires PRPP, but for

purines

, PRPP provides the foundation on which the bases are constructed step by step. The initial committed step is the

displacement of pyrophosphate by ammonia

, rather than by a preassembled base, to produce

5-phosphoribosyl-1-amine, with the amine in the

β

configuration.

Slide10

de Novo Purine Biosynthesis

Slide11

Inosinate Formation

Slide12

Generating AMP and GMP

Slide13

Slide14

Slide15

Slide16

Slide17

Slide18

Salvage Pathways Economize Intracellular Energy Expenditure

Two salvage enzymes with different specificities recover

purine

bases.

Adenine

phosphoribosyltransferase

catalyzes the formation of

adenylate

Slide19

whereas

hypoxanthine-guanine

phosphoribosyltransferase

(HGPRT)

catalyzes the formation of

guanylate

as well as

inosinate

(

inosine

monophosphate

, IMP), a precursor of

guanylate

and

adenylate

Slide20

Pyrimidine Nucleotides Are Made from Aspartate

, PRPP, and

Carbamoyl

Phosphate

The common

pyrimidine

ribonucleotides

are

cytidine

5-monophosphate (CMP;

cytidylate

) and

uridine

5-monophosphate (UMP;

uridylate

), which contain the

pyrimidines

cytosine and

uracil

. De novo

pyrimidine

nucleotide Biosynthesis proceeds in a somewhat different manner from

purine

nucleotide synthesis;

the six-

membered

pyrimidine

ring is made first and then attached to ribose 5-phosphate.

Slide21

In the first step of the

carbamoyl

phosphate synthesis pathway, bicarbonate is

phosphorylated

by ATP to form

carboxyphosphate

and ADP. Ammonia then reacts with

carboxyphosphate

to form

carbamic

acid and inorganic phosphate.

Slide22

Carbamoyl

phosphate reacts with

aspartate

to form

carbamoylaspartate

in a reaction catalyzed by

aspartate

Transcarbamoylase

.

Carbamoylaspartate

then

cyclizes

to form

dihydroorotate

which is then oxidized by

NAD+ to form

orotate

.

Slide23

Orotate

reacts with PRPP to form

orotidylate

, a

pyrimidine

nucleotide. This reaction is driven by the hydrolysis of pyrophosphate. The enzyme that catalyzes this addition,

pyrimidine

phosphoribosyltransferase

,

Slide24

Pyrimidine synthesis

Slide25

Carbamoyl phosphate reacts with

aspartate

to yield

N-

carbamoylaspartate

in the first committed step of

pyrimidine

biosynthesis . This reaction is catalyzed by

aspartate

transcarbamoylase

Slide26

By removal of water from

N-

carbamoylaspartate

, a reaction catalyzed by

dihydroorotase

, the

pyrimidine

ring is closed to form

L-

dihydroorotate

.

This compound is oxidized to the

pyrimidine

derivative

orotate

, a reaction in which NAD is the ultimate electron acceptor.

Slide27

Once

orotate

is formed, the ribose 5-phosphate side chain, provided once again by PRPP, is attached to yield

orotidylate

.

Orotidylate

is then

decarboxylated

to

uridylate

, which is

phosphorylated

to UTP. CTP is formed from UTP by the action of

cytidylate

synthetase

.

Slide28

de Novo Pathway for Pyrimidine Nucleotide Synthesis.

The C-2 and N-3 atoms in the

pyrimidine

ring

come from

carbamoyl

phosphate, where as the other atoms of the ring come from

aspartate

.

Slide29

The

pyrimidine

ring is made up of three components:

the nitrogen atom N-1 and carbons C-4 to C-6 are derived from

aspartate

, carbon

C-2 comes from

HCO3-

, and the second nitrogen (N-3) is taken from the amide group of

glutamine

.

Slide30

Slide31

Degradation of Purines and Pyrimidines

Produces

Uric Acid and Urea, Respectively

Purine

nucleotides are degraded by a pathway in which they lose their phosphate through the action of

5-Nucleotidase .

Adenylate

yields adenosine,

which is

deaminated

to

inosine

by

adenosine

deaminase

,

and

inosine

is hydrolyzed to hypoxanthine (its

purine

base) and D-ribose. Hypoxanthine is oxidized successively to

xanthine

and then uric acid by

xanthine

oxidase

, a

flavoenzyme

with an atom of molybdenum

and four iron-sulfur centers in its prosthetic group. Molecular oxygen is the electron acceptor in this complex reaction.

Slide32

Purine Catabolism

Purine

bases are converted first into

xanthine

and then into

urate

for excretion.

Xanthine

oxidase

catalyzes two steps in this process.

Slide33

Urate Crystals.

Micrograph of sodium

urate

crystals. Joints and kidneys are damaged by these crystals in

gout.

Slide34

Uric acid is the excreted end product of

purine

catabolism in

primates, birds, and some other animals

. A healthy adult human excretes uric acid at a rate of about 0.6 g/24 h; the excreted product arises in part from ingested

purines

and in part from turnover of the

purine

nucleotides of nucleic acids. In most mammals and many other vertebrates, uric acid is further degraded to

allantoin

by the action of

urate

oxidase

.

Slide35

Slide36

Slide37

Lesch-Nyhan syndrome

A genetic lack of

hypoxanthine-guanine

phosphoribosyltransferase

activity

, seen almost exclusively in male children, results in a bizarre set of symptoms

.

Children

with this genetic disorder, which becomes manifest by the age of 2 years, are sometimes poorly coordinated and mentally retarded. In addition, they are extremely hostile and show compulsive self-destructive tendencies:

they mutilate themselves by biting off their fingers,

toes, and lips.

Slide38

Excess Uric Acid Causes Gout

Long thought, erroneously, to be due to “high living,” gout is a disease of the joints caused by an elevated concentration of uric acid in the blood and tissues.

The joints become inflamed, painful, and arthritic, owing to the abnormal deposition of sodium

urate

crystals.

The kidneys are also affected, as excess uric acid is deposited in the kidney tubules.

Gout occurs predominantly in males. Its precise cause is not known, but it often involves an

underexcretion

of

urate

. A genetic deficiency of one or another enzyme of

purine

metabolism may also be a factor in some cases.

Slide39

Gout is effectively treated by a combination of nutritional and drug therapies. Foods especially rich in nucleotides and nucleic acids, such as liver or glandular products, are withheld from the diet. Major alleviation of the symptoms is provided by the drug

allopurinol

,

which inhibits

xanthine

oxidase

, the enzyme that catalyzes the conversion of

purines

to uric acid.

Slide40

multiple tophi on the hands (Panel A), feet, knees, Some of the tophi exuded a white, chalky material.

Laboratory studies were notable for a serum uric acid level of 8.5 mg per deciliter (506 µmol per liter), Xray hand:soft tissue swelling and pararticular erosions

Slide41

The Gout-By James Gilray-1799

Johnson and Rideout NEJM, 350 (11): 1071, Figure 1     March 11, 2004

Slide42

Slide43

Control of Purine Biosynthesis.

Feedback inhibition controls both the overall rate of

purine

biosynthesis

and the balance between AMP and GMP production

Slide44

Many Chemotherapeutic Agents Target Enzymes

in the Nucleotide Biosynthetic Pathways

The first set of agents includes compounds that inhibit

glutamine

amidotransferases

.

Recall

that

glutamine

is a nitrogen donor in at least half a dozen separate reactions in nucleotide biosynthesis. The binding sites for glutamine and the mechanism by which NH4 is extracted are quite similar in many of these enzymes. Most are strongly inhibited by glutamine analogs such as

azaserine

and

acivicin

Slide45

Several Valuable Anticancer Drugs Block the Synthesis of

Thymidylate

One inhibitor that acts on

thymidylate

synthase

,

fluorouracil, is an important

chemotherapeutic agent. Fluorouracil itself is not the enzyme inhibitor. In the cell, salvage pathways convert it to the

deoxynucleoside

monophosphate

FdUMP

, which binds to and inactivates the enzyme. Inhibition by

FdUMP

is a classic example of mechanism-based enzyme inactivation. Another prominent chemotherapeutic agent,

methotrexate

, is an inhibitor

of

dihydrofolate

reductase

. This

folate

analog acts as a competitive inhibitor; the enzyme binds

methotrexate

with about 100 times higher affinity than

dihydrofolate

.

Slide46

Methotrexate

is a valuable drug in the treatment of many rapidly growing tumors, such as those in acute leukemia and

choriocarcinoma

, a cancer derived from placental cells. However,

methotrexate

kills rapidly replicating cells whether they are malignant or not.

Slide47

Anticancer Drug Targets

Slide48

END

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