Purine & Pyrimidine Metabolism

Synthesis and degradation of nucleotides.

Biochemical Importance of Nucleotides

• Nucleotides are precursors to DNA and RNA.

• Nucleotides are precursors to metabolites like UDP-glucose (glycogen) and CDP-diacylglicerol (phosphoglicerides).

• Nucleotides serve as metabolic regulators (through phosphorylation, etc.)

• Intracellular messengers. (cAMP and cGMP)

• Important diseases like gout, Lesh-Nyhan, and ADA deficiency.

• Drugs like AZT, and anticancers drugs.

Gout

“goute” = drop

“drop” = uric acid crystal deposition

What it is

• A metabolic disease caused by elevated uric acid (hyperuricemia)

• Leads to crystal deposition in joints

Cause

• Breakdown of purines → uric acid

• When uric acid is too high → forms monosodium urate crystals

These crystals deposit in joints → trigger inflammation

Classic presentation

• Sudden, severe joint pain (acute gout attack)

• Most commonly:

  • Big toe (1st MTP joint) → podagra

• Other signs:

  • Redness

  • Swelling

  • Warmth

Pathophysiology

• Uric acid crystals are recognized as “foreign”

• Activate immune response → neutrophils → inflammation

Key connection to your slide

• Gout is a disorder of nucleotide (purine) metabolism
  👉 Excess breakdown → excess uric acid

Lesh-Nyhan Syndrome

• Named after two physicians:

  • Michael Lesch

  • William Nyhan

They first described the disorder in 1964

Lesch–Nyhan syndrome

What it is

• A genetic disorder of purine metabolism

• Causes overproduction of uric acid

• X-linked recessive (mostly affects males)

Cause

• Deficiency of the enzyme:
  Hypoxanthine-guanine phosphoribosyltransferase (HGPRT)

HGPRT = an enzyme that transfers a phosphoribosyl group to hypoxanthine and guanine

It recycles purines instead of making new ones

This enzyme is part of the purine salvage pathway

Pathophysiology (high-yield)

• Normally: HGPRT recycles purines

• Without it:

  • ❌ Purines are not salvaged

  • ↑ De novo purine synthesis

  • ↑ Breakdown → ↑ uric acid

Leads to gout-like symptoms

Classic triad (VERY important)

1. Hyperuricemia

   • Gout

   • Kidney stones

   • “Orange sand” (urate crystals in diapers)

2. Neurologic dysfunction

   • Developmental delay

   • Choreoathetosis (abnormal movements)

3. Self-mutilation

   • Biting lips, fingers

This is the buzzword feature

ADA Deficiency

What is ADA?

• ADA = adenosine deaminase

• Enzyme in purine metabolism

What happens in deficiency

• ADA normally converts:

  • adenosine → inosine

• Without ADA:

  • ↑ adenosine

  • ↑ deoxyadenosine

  • ↑ dATP

dATP buildup is toxic to lymphocytes

Key effect

• Inhibits ribonucleotide reductase

• ↓ DNA synthesis

Lymphocytes (T & B cells) cannot proliferate

Disease result

• Severe Combined Immunodeficiency (SCID)

Both:

• ❌ T cells

• ❌ B cells

What is AZT?

• AZT = Azidothymidine

• Also called Zidovudine

• A nucleoside reverse transcriptase inhibitor (NRTI)

Etymology

• Azi-do → refers to an azide (–N₃) group

• Thymidine → a DNA nucleoside (thymine + deoxyribose)

So AZT = a modified thymidine molecule

Mechanism (high-yield)

1. AZT mimics thymidine

2. Gets incorporated into viral DNA by reverse transcriptase

3. BUT:

   • Lacks a 3′-OH group

❌ DNA chain cannot elongate → chain termination

Use

• Treatment of HIV

• Prevents viral replication

Why it works

• HIV relies on reverse transcriptase

• AZT specifically targets this step

AZT = fake thymidine → stops HIV DNA synthesis (chain termination)

-thymine is DNA, uracil is RNA and replaces thymine.

anticancer drugs are nucleotide analogs, why?

Cancer cells:

• Divide rapidly

• Require constant DNA synthesis

Target DNA building → target cancer

How nucleotide analogs work

They are “fake nucleotides” that interfere with DNA/RNA synthesis:

1. Chain termination

• Incorporated into DNA

• Missing a 3′-OH
  DNA cannot elongate

We will start start talking now on the synthesis of nucleotides.

The parent compound for the purine nucleotides is IMP (inositol-monophosphate).

Both purine nucleotides are derived from IMP. (Because IMP (inosine monophosphate) is the first fully formed purine nucleotide made in the de novo synthesis pathway.)

(warning): there is a long sequence of reactions coming up.

We will go reaction by reaction. This is the overall series of reactions that are involved in the synthesis of IMP.

IMP is the last molecule you see in this sequence. You need 11 reactions to get to IMP.

We will be looking at the reactions in some detail and notice that all the reactions have a number and will be referred to by that number.

Purine Synthesis cont.

1. Formation of PRPP

2. Formation of N-glycosidic bond (Commitment step)

The first reaction is the synthesis of PRPP (phospho-ribo-pyro-phosphate).

1. a-D-ribose-5-phosphate (PRPP synthase) → PRPP

This first reaction requires ATP. The ATP will donate the pyro-phosphate (two phosphates)

2. PRPP (PRPP Glutamyl Amido-transferase) → 5-phospho-ribosylamine

PRPP Glutamyl Amido-transferase: Enzyme that transfers an amide nitrogen from glutamine to PRPP

Reaction #2 is the introduction of nitrogen. This nitrogen group is donated by glutamine. Glutamine will use the energy of the pyro-phosphates to move the ammonia into PRPP.

Reaction #2 is a very important reaction for two reasons. The first reason is it is the commitment step. It is the commitment step because there is no other use of 5-phospho-ribosylamine than to create Purine nucleotides.

The second reason is that, in reaction #2, once you put the nitrogen there, you are creating the glycosidic bond. The glycosidic bond is the bond that holds the nitrogen base to the sugar.

Purine Synthesis cont.

3. 5-phospho-ribosylamine (Glycinamide ribonucleotide synthetase (GAR synthetase) → glycinamide ribosyl-5-phosphate

Incorporation of glycine

Brings in carbon from the amino acid glycine. Glycine will donate its skeleton, it’s attached to the nitrogen of the glycosidic bond and is the start of the nitrogen base. This reaction requires ATP, this reaction consumes one phosphate group. One phosphate group bond, the energy, is used to put glycine in place.

4. glycinamide ribosyl-5-phosphate (Formyl Transferase) → Formylglycinamide ribosyl-5-phosphate

Formyltransferase = enzyme that transfers a formyl (–CHO) group,

Uses: N¹⁰-formyl-tetrahydrofolate (THF) as the donor

Transfer of a formyl group, inhibited by folic acid antagonists (aminopterin)

• Folate antagonists (e.g., aminopterin, methotrexate):

  • ↓ THF

  • ❌ Block formyl transfer

  • ❌ Stop purine synthesis → ↓ DNA synthesis

That’s why they’re used as anticancer drugs

Reaction #4 is the introduction of a formyl group, derived from formic acid. It’s a one carbon residue. The molecule that usually donates that one carbon units is methyl-tetra-hydryl-folate. This one carbon residue is introduced, this is a place where folic acid antagonists, like aminopterin, will inhibit.

Remember, we will be encountering a couple of reactions that need tetrahydrofolate, things that will inhibit tetrahydrolfolic, for example, lack of folic acid, or aminopterin will inhibit those specific reactions.

This is the first time we encounter the donation of the one carbon mioety of the tetrahydrolfolate.

5. Glutamine donates another nitrogen.

5. Formylglycinamide ribosyl-5-phosphate (FGAR) → Formylglycinamidine ribosyl-5-phosphate (FGAM)
   (with glutamine donating nitrogen, becomes glutamate)

FGAM synthetase

The next step, #5, is the introduction of another nitrogen. Nitrogen is brought in by glutamine and attaches to the oxygen. This is a mechanism that requires the presence of ATP, but ATP is hydrolyzed to provide the energy.

6. Formylglycinamidine ribosyl-5-phosphate (VI) → Aminoimidazole ribosyl-5-phosphate (VII)

is catalyzed by: AIR synthetase

Removal of water closes the ring. (Usually, in reactions where the circle is closed, you will see the removal of water).

7. Aminoimidazole ribosyl-5-phosphate (VII) → Aminoimidazole carboxylate ribosyl-5-phosphate (VIII)
   CO₂ addition. Here, we are starting to build the larger ring of the purine nitrogen base.

8. Aminoimidazole carboxylate ribosyl-5-phosphate (VIII) → Aminoimidazole succinyl carboxamide ribosyl-5-phosphate (IX)

Aspartate is added. The donor of the nitrogen is Aspartate, it’s not glutamine. Aspartate will be bound to the molecule in reaction 8, and water will come out.

9. Aminoimidazole succinyl carboxamide ribosyl-5-phosphate (IX) → Aminoimidazole carboxamide ribosyl-5-phosphate (X)
   ADENYLOSUCCINASE

Fumarate is liberated.

The molecule fumarate will exit, leaving behind the nitrogen. The nitrogen is brought in by aspartate, and then it’s cleaved and the nitrogen stays behind and fumarate leaves.

IMP is Formed

10. Aminoimidazole carboxamide ribosyl-5-phosphate (X) → Formimidoimidazole carboxamide ribosyl-5-phosphate (XI)

FORMYLTRANSFERASE

(again it comes from tetrahydrylfolate)

Another formyl group added.

11. Formimidoimidazole carboxamide ribosyl-5-phosphate → Inosine monophosphate (IMP)

H₂O leaves
Ring closure

IMP, the first purine nucleotide because IMP has the sugar, the phosphate and the nitrogen base, it’s a nucleotide.

12. Inosine monophosphate (IMP) → adenylosuccinate (AMPS)

ADENYLOSUCCINATE SYNTHASE

GTP is needed , Mg²⁺

Reaction 12 is the donation of nitrogen by aspartate. Aspartate gets bound to the nitrogen base, leaving as fumarate, and leaves behind the nitrogen. In this way, you have AMP, Adenosine Monophosphate.

In Reaction 12, you need GTP to synthesize AMP. points to the circled GTP

13. adenylosuccinate (AMPS) → Adenosine monophosphate (AMP)

ADENYLOSUCCINASE

Reaction 12 and 13 is the formation of AMP. IMP is like a branch piont because it is the parent compound o both nitrogen bases, AMP and GMP.

You also need ATP to synthesize GMP.

The formation of GMP requires ATP.

4. glycinamide ribosyl-5-phosphate (Formyl Transferase) → Formylglycinamide ribosyl-5-phosphate

Formyltransferase = enzyme that transfers a formyl (–CHO) group,

Uses: N¹⁰-formyl-tetrahydrofolate (THF) as the donor

Transfer of a formyl group, inhibited by folic acid antagonists (aminopterin)

• Folate antagonists (e.g., aminopterin, methotrexate):

  • ↓ THF

  • ❌ Block formyl transfer

  • ❌ Stop purine synthesis → ↓ DNA synthesis

That’s why they’re used as anticancer drugs

10. Aminoimidazole carboxamide ribosyl-5-phosphate (X) → Formimidoimidazole carboxamide ribosyl-5-phosphate (XI)

FORMYLTRANSFERASE

(again it comes from tetrahydrylfolate)

Another formyl group added.

5. Formylglycinamide ribosyl-5-phosphate (FGAR) → Formylglycinamidine ribosyl-5-phosphate (FGAM)
   (with glutamine donating nitrogen, becomes glutamate)

FGAM synthetase

2. PRPP (PRPP Glutamyl Amido-transferase) → 5-phospho-ribosylamine

13. adenylosuccinate (AMPS) → Adenosine monophosphate (AMP)

ADENYLOSUCCINASE

Reaction 12 and 13 is the formation of AMP. IMP is like a branch piont because it is the parent compound o both nitrogen bases, AMP and GMP.

You also need ATP to synthesize GMP.

Folate antagonist — what it means

👉 A folate antagonist is a substance (often a drug) that blocks the function or metabolism of folate (vitamin B9).

Folate is essential for:

• DNA synthesis

• Making thymidine (dTMP)

• Supporting rapidly dividing cells

1. Blocking folate activation

• Inhibits Dihydrofolate reductase (DHFR)

• Prevents formation of tetrahydrofolate (THF) (active form)

2. Mimicking folate (competitive inhibition)

• Structurally resembles folate

• Competes for enzymes → blocks normal functio

Adenine phosphoribosyltransferase (APRT) 🔤 Etymology (decode the name)

• Adenine → the purine base

• Phosphoribosyl → ribose sugar + phosphate group

• Transferase → transfers a group

👉 “An enzyme that transfers a phosphoribosyl group onto adenine.”

defects in hypoxanthine-guanine phospho ribosyl transferase

unlike purine synthesis, the nitrogen base is assembled independently of PRPP.

Location: cytosol

end product: DHOA

location: mitochondria

end product: DHOA → OA (Orotic Acid)

UMP: first true pyrimidine nucleotide, made from Orotic Acid

Allopurinol is NOT directly involved in pyrimidine synthesis.
It primarily affects purine metabolism.

What allopurinol actually does

• Inhibits Xanthine oxidase

• Blocks:

  • Hypoxanthine → xanthine

  • Xanthine → uric acid

👉 Result:

• ↓ uric acid (used for gout)

Uric acid 🔤 Etymology

• Uric → from “urine” (it’s excreted there)

• Acid → weak organic acid

👉 “The end product of purine breakdown that is excreted in urine.”