carbamoyl phosphate synthetase

**Reaction:** CO2 (as HCO3⁻) + NH4⁺ + 2 ATP → carbamoyl phosphate + 2 ADP + Pi. Two ATP are used. One ATP activates bicarbonate and the other phosphorylates carbamate. The reaction is irreversible and is the rate-limiting step of the urea cycle.

**Location and isoforms:** - **CPS I:** mitochondrial matrix, mainly in the liver. It uses free ammonia as the nitrogen source. It is the urea cycle enzyme, and it is required for disposal of nitrogen. - **CPS II:** cytosol. It is part of the multifunctional CAD protein (CPS II, Aspartate transcarbamoylase, Dihydroorotase). It uses glutamine as the nitrogen source. It is the rate-limiting step of de novo pyrimidine synthesis. It is inhibited by UTP and activated by PRPP and ATP. NAG does not regulate it.

**Regulation of CPS I:** It is allosterically activated by N-acetylglutamate (NAG), which is made by NAG synthase from acetyl-CoA + glutamate. Arginine activates NAG synthase. High amino acid catabolism raises glutamate and arginine, which raises NAG and so drives the urea cycle.

**Next step:** Carbamoyl phosphate + ornithine → citrulline, catalyzed by ornithine transcarbamylase (OTC) in the mitochondria. Citrulline is then exported to the cytosol.

**Clinical significance:** - **CPS I deficiency:** autosomal recessive. It presents in the neonatal period, after protein feeding starts, with poor feeding, vomiting, lethargy, hypotonia, seizures, and cerebral edema. Labs show hyperammonemia, respiratory alkalosis (ammonia stimulates the respiratory center), low BUN, low citrulline and arginine, and normal or low orotic acid. - **NAG synthase deficiency:** has the same biochemical picture as CPS I deficiency, but it responds to carglumic acid, a NAG analog. - **OTC deficiency:** X-linked recessive and the most common urea cycle disorder. Carbamoyl phosphate accumulates and spills into the cytosol, where it enters pyrimidine synthesis. This causes **high orotic acid** in the urine, with low citrulline and low BUN. This is the key discriminator from CPS I deficiency, which has normal or low orotic acid. - **Pathophysiology of toxicity:** Excess ammonia crosses the blood–brain barrier. Astrocytes convert it to glutamine with glutamine synthetase, which depletes glutamate and α-ketoglutarate. The glutamine causes osmotic swelling (cerebral edema), and α-ketoglutarate depletion impairs the TCA cycle and ATP production. - **Treatment:** Restrict dietary protein. Give nitrogen scavengers (sodium benzoate, sodium phenylbutyrate/phenylacetate), arginine or citrulline supplementation, and lactulose in some cases. Use hemodialysis for severe hyperammonemia. Carglumic acid is used for NAG synthase deficiency. - **Acquired hyperammonemia:** Liver disease (cirrhosis) impairs the urea cycle. Valproate can cause hyperammonemia through inhibition of CPS I and NAG synthase (carnitine depletion is also implicated).

**High-yield mnemonics:** - CPS I = urea cycle, mitochondria, ammonia, NAG-activated. CPS II = pyrimidines, cytosol, glutamine. - Hyperammonemia with high orotic acid → OTC deficiency. Hyperammonemia without high orotic acid → CPS I or NAG synthase deficiency.