CPS1 variants cause CPS1 deficiency

The CPS1 enzyme, located in the mitochondrial matrix, catalyzes the first and rate-limiting step of the urea cycle: the synthesis of carbamoyl phosphate from ammonia, bicarbonate, and ATP. This reaction is essential for the detoxification of ammonia in the liver (Pierson and Brien, 1980; Rubio and Grisolia, 1981; Pekkala et al, 2009; Diez-Fernandez et al, 2013; reviewed in Martinez et al, 2010; Nitzahn and Lipshutz, 2020).

Synthesis of carbamoyl phosphate by human CPS1 occurs in three linked biochemical steps. In the first step, ATP-dependent phosphorylation of bicarbonate yields the unstable intermediate carboxyphosphate. In the second step, carboxyphosphate reacts with ammonia to produce the next unstable intermediate, carbamate. In the final step, ATP-dependent phosphorylation of carbamate yields carbamoyl phosphate (reviewed in Martinez et al, 2010). Enzyme activity depends on the binding of the allosteric activator N-acetyl L-glutamate (NAG) to its binding domain in the C-terminal (Grisolia and Cohen 1953; Rubio et al, 1983. The ancestral CPS enzyme had an additional glutaminase activity that hydrolyzed glutamine to produce ammonia for use in the second step. The activity of this domain is conserved in the E.coli enzyme but has been lost in mammalian and other higher order enzymes (reviewed in Nitzahn and Lipshutz, 2020; Zhang et al, 2023).

Various functional domains contribute to the enzymatic activity and stability of the protein. The N-terminal region consists of a N-terminal extension of unknown function and the inactive glutaminase domain; these regions may contribute to overall structure and stabilization of the active conformation. The bicarbonate and carbamate phosphorylation domains are separated by an "integrating domain" (ID) that contributes to the structural integration of CPS1, influencing enzyme folding and stability and providing a tunnel for the carbamate to transfer between the two catalytic sites. Mutations in this domain can lead to misfolding and reduced enzyme activity. The C-terminal NAG binding domain binds the essential activator of CPS1; in consequence, any CPS1 nonsense mutation that precedes this domain is effectively non-functional.

Over 270 mutations have been identified in the CPS1 gene, with a predominance of missense mutations affecting various domains. Notably, mutations in the bicarbonate phosphorylation domain, the integrating domain and the NAG-binding domain are more likely to result in severe clinical manifestations due to their direct impact on enzyme function (de Cima et al, 2015; Yan et al, 2019; reviewed in Martinez et al, 2010)