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[Biochemistry](https://durgical.com/en/biochemistry/) · Metabolic pathway

# Pyrimidine synthesis

Pyrimidines are built the other way round from purines: ring first (orotate), sugar second. UMP, CTP and dTMP come from there.

Updated 2026-11-01

## Interactive map

## Key facts

- Ring first, sugar second: orotate joins PRPP at step 5.
- CPS-II (cytosol, uses glutamine) is the rate-limiting step in humans; UTP inhibits it and PRPP and ATP activate it. Not to be confused with CPS-I (mitochondria, ammonia, urea cycle).
- Three proteins do almost everything: CAD (steps 1–3), DHODH (step 4, in mitochondria) and UMP synthase (steps 5–6).
- Deoxyribonucleotides come from ribonucleotide reductase (diphosphates); dTMP forms by methylation of dUMP by thymidylate synthase.
- Ring atoms: aspartate (N1, C4, C5, C6), CO₂ (C2) and glutamine (N3).

## In detail

- CTP is made from UTP by CTP synthetase, which takes nitrogen from glutamine.
- Steps 9–10: NDP kinase converts dUDP to dUTP and dUTPase cuts it to dUMP, which keeps uracil out of DNA.
- Another source of dUMP is deamination of dCMP (dCMP deaminase).
- DHODH hands its electrons to ubiquinone, linking pyrimidine synthesis with the respiratory chain.
- Thymidylate synthase oxidises folate to DHF; DHFR returns it to THF. That is why methotrexate also stops thymidine synthesis.

**Also called**
Orotate pathway

## Regulation

- CPS-II: UTP (−), PRPP and ATP (+); phosphorylation by MAP kinases activates it when the cell grows.
- Ribonucleotide reductase is regulated by dATP (off) and ATP (on) and chooses its substrate according to the bound dNTP.
- In bacteria control lies at aspartate transcarbamoylase (inhibited by CTP); in humans, at CPS-II.

## The steps, one by one

- **Step 1 · [Carbamoyl phosphate synthetase II (CAD)](https://durgical.com/en/biochemistry/enzymes/cad/)** irreversible [Glutamine](https://durgical.com/en/biochemistry/molecules/glutamine/) + CO₂ + [ATP](https://durgical.com/en/biochemistry/molecules/atp/) → Carbamoyl phosphate + [Glutamate](https://durgical.com/en/biochemistry/molecules/glutamate/) + ADP + phosphate (Pi) Rate-limiting step in the cytosol: bicarbonate and glutamine nitrogen form carbamoyl phosphate using 2 ATP. **If it fails:** CAD deficiency causes an epileptic encephalopathy that improves with uridine.
- **Step 2 · Aspartate transcarbamoylase (CAD domain)** irreversible Carbamoyl phosphate + Aspartate → Carbamoyl aspartate + phosphate (Pi) Joins aspartate to carbamoyl phosphate; most of the ring atoms enter here. **If it fails:** It is part of CAD; its defects appear as CAD deficiency.
- **Step 3 · Dihydroorotase (CAD domain)** Carbamoyl aspartate ⇌ Dihydroorotate + H₂O Closes the six-membered ring by removing a water molecule. **If it fails:** It is part of CAD; its defects appear as CAD deficiency.
- **Step 4 · [Dihydroorotate dehydrogenase (DHODH)](https://durgical.com/en/biochemistry/enzymes/dihydroorotate-dehydrogenase/)** irreversible Dihydroorotate + [Ubiquinone (CoQ)](https://durgical.com/en/biochemistry/molecules/ubiquinone/) → [Orotic acid](https://durgical.com/en/biochemistry/molecules/orotic-acid/) + Ubiquinol (CoQH₂) The only mitochondrial step: oxidises the ring to orotate and passes electrons to ubiquinone. **If it fails:** DHODH variants cause Miller syndrome (postaxial acrofacial dysostosis).
- **Step 5 · [UMP synthase: orotate phosphoribosyltransferase](https://durgical.com/en/biochemistry/enzymes/ump-synthase/)** [Orotic acid](https://durgical.com/en/biochemistry/molecules/orotic-acid/) + [PRPP](https://durgical.com/en/biochemistry/molecules/prpp/) ⇌ Orotidine monophosphate (OMP) + pyrophosphate The finished ring receives ribose phosphate from PRPP and becomes a nucleotide (OMP). **If it fails:** Its deficiency causes hereditary orotic aciduria. [Hereditary orotic aciduria →](https://durgical.com/en/conditions/hereditary-orotic-aciduria/)
- **Step 6 · [UMP synthase: OMP decarboxylase](https://durgical.com/en/biochemistry/enzymes/ump-synthase/)** irreversible Orotidine monophosphate (OMP) → Uridine monophosphate (UMP) + CO₂ Removes the orotate carboxyl and forms UMP, the first complete pyrimidine. **If it fails:** Its deficiency causes hereditary orotic aciduria. [Hereditary orotic aciduria →](https://durgical.com/en/conditions/hereditary-orotic-aciduria/)
- **Step 7 · UMP/CMP kinase** Uridine monophosphate (UMP) + [ATP](https://durgical.com/en/biochemistry/molecules/atp/) ⇌ Uridine diphosphate (UDP) + ADP Adds a phosphate, giving UDP, the substrate of ribonucleotide reductase. **If it fails:** It also phosphorylates analogues such as gemcitabine monophosphate.
- **Step 8 · [Ribonucleotide reductase](https://durgical.com/en/biochemistry/enzymes/ribonucleotide-reductase/)** irreversible Uridine diphosphate (UDP) → Deoxyuridine diphosphate (dUDP) + H₂O Removes the oxygen at C2' of the ribose: the ribonucleotide becomes a deoxyribonucleotide. **If it fails:** dATP that builds up in ADA deficiency switches it off and kills lymphocytes. [Adenosine deaminase deficiency (ADA-SCID) →](https://durgical.com/en/conditions/adenosine-deaminase-deficiency/)
- **Step 9 · Nucleoside diphosphate kinase** Deoxyuridine diphosphate (dUDP) + [ATP](https://durgical.com/en/biochemistry/molecules/atp/) ⇌ Deoxyuridine triphosphate (dUTP) + ADP Moves a phosphate from ATP to dUDP, forming dUTP. **If it fails:** An abundant, non-specific enzyme; no known disease from its loss.
- **Step 10 · dUTPase** irreversible Deoxyuridine triphosphate (dUTP) + H₂O → Deoxyuridine monophosphate (dUMP) + pyrophosphate Cuts dUTP to dUMP, keeping dUTP low so DNA polymerase does not put uracil into DNA. **If it fails:** When thymidylate synthase is blocked (5-FU), uracil enters DNA and breaks during repair.
- **Step 11 · [Thymidylate synthase](https://durgical.com/en/biochemistry/enzymes/thymidylate-synthase/)** irreversible Deoxyuridine monophosphate (dUMP) + N⁵,N¹⁰-Methylene-tetrahydrofolate → [dTMP](https://durgical.com/en/biochemistry/molecules/dtmp/) + Dihydrofolate (DHF) Puts a methyl on C5 of uracil: dUMP becomes dTMP and the folate is left oxidised (DHF). **If it fails:** Lack of folate or B12 slows this step: megaloblastic anaemia.

## Clinical notes

- Hereditary orotic aciduria: UMP synthase fails, orotate builds up and a megaloblastic anaemia appears that does not respond to B12 or folate.
- In OTC deficiency (urea cycle) mitochondrial carbamoyl phosphate spills into the cytosol and orotic acid also rises, but with hyperammonaemia.
- Drugs: leflunomide (DHODH), 5-fluorouracil (thymidylate synthase), methotrexate (DHFR) and hydroxyurea (ribonucleotide reductase).

## Related conditions

- [Hereditary orotic aciduria](https://durgical.com/en/conditions/hereditary-orotic-aciduria/)

## Related

- [CAD (carbamoyl phosphate synthetase II)](https://durgical.com/en/biochemistry/enzymes/cad/)
- [Dihydroorotate dehydrogenase (DHODH)](https://durgical.com/en/biochemistry/enzymes/dihydroorotate-dehydrogenase/)
- [Orotic acid](https://durgical.com/en/biochemistry/molecules/orotic-acid/)
- [UMP synthase](https://durgical.com/en/biochemistry/enzymes/ump-synthase/)
- [PRPP (phosphoribosyl pyrophosphate)](https://durgical.com/en/biochemistry/molecules/prpp/)
- [Ribonucleotide reductase](https://durgical.com/en/biochemistry/enzymes/ribonucleotide-reductase/)
- [Thymidylate synthase](https://durgical.com/en/biochemistry/enzymes/thymidylate-synthase/)
- [Dihydrofolate reductase (DHFR)](https://durgical.com/en/biochemistry/enzymes/dihydrofolate-reductase/)
- [dTMP (thymidine monophosphate)](https://durgical.com/en/biochemistry/molecules/dtmp/)
- [Drugs acting on nucleotide metabolism](https://durgical.com/en/biochemistry/topics/drugs-acting-on-nucleotide-metabolism/)
- [Nucleotide structure](https://durgical.com/en/biochemistry/topics/nucleotide-structure/)

## Test yourself

Think of the answer, then tap the question to see it.

## How do CPS-I and CPS-II differ?

CPS-I is mitochondrial, uses ammonia and is activated by N-acetylglutamate (urea); CPS-II is cytosolic, uses glutamine and is inhibited by UTP (pyrimidines).

## Which lab finding separates OTC deficiency from hereditary orotic aciduria?

Both raise urinary orotic acid, but only OTC deficiency comes with hyperammonaemia.

## Which enzyme of the pathway sits in mitochondria, and which drug blocks it?

Dihydroorotate dehydrogenase; leflunomide (through its metabolite teriflunomide) blocks it.

[Add to my cards (3)](https://durgical.app/estudiar/?add=/en/biochemistry/pathways/pyrimidine-synthesis/)

Educational information, not medical advice. For any symptom, see a health professional.

Sources: [Pyrimidine biosynthesis](https://reactome.org/PathwayBrowser/#/R-HSA-500753) (Reactome, 2026); [Interconversion of nucleotide di- and triphosphates](https://reactome.org/PathwayBrowser/#/R-HSA-499943) (Reactome, 2026); [CAD: Multifunctional protein CAD (human)](https://www.uniprot.org/uniprotkb/P27708/entry) (UniProt, 2026); Textbook of Biochemistry for Medical Students, 6th ed. (Jaypee Brothers Medical Publishers (Vasudevan DM, Sreekumari S, Vaidyanathan K), 2011).

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Representative model, for educational purposes only.

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