The first reaction catalysed by CPS I is the rate-limiting enzyme reaction or committed step in urea synthesis. CPS I is allosterically activated by NAG. It is synthesised from glutamate and acetyl CoA by synthase and degraded by a hydrolase.

The rate of urea synthesis in liver is correlated with the concentration of NAG. The consumption of protein-rich meal increases the level of NAG in liver, leading to enhanced urea synthesis.

CPS I and glutamate dehydrogenase are localised in the mitochondria. They coordinate with each other in the formation of NH₃ and its utilisation for the synthesis of carbamoyl phosphate. The remaining four enzymes of urea cycle are mostly controlled by the concentration of their respective substrates.