The Study of Carbamoyl Phosphate Synthetase 1 Deficiency Sheds Light on the Mechanism for Switching On/Off the Urea Cycle

J Genet Genomics. 2015 May 20;42(5):249-60. doi: 10.1016/j.jgg.2015.03.009. Epub 2015 Apr 1.


Carbamoyl phosphate synthetase 1 (CPS1) deficiency (CPS1D) is an inborn error of the urea cycle having autosomal (2q34) recessive inheritance that can cause hyperammonemia and neonatal death or mental retardation. We analyzed the effects on CPS1 activity, kinetic parameters and enzyme stability of missense mutations reported in patients with CPS1 deficiency that map in the 20-kDa C-terminal domain of the enzyme. This domain turns on or off the enzyme depending on whether the essential allosteric activator of CPS1, N-acetyl-L-glutamate (NAG), is bound or is not bound to it. To carry out the present studies, we exploited a novel system that allows the expression in vitro and the purification of human CPS1, thus permitting site-directed mutagenesis. These studies have clarified disease causation by individual mutations, identifying functionally important residues, and revealing that a number of mutations decrease the affinity of the enzyme for NAG. Patients with NAG affinity-decreasing mutations might benefit from NAG site saturation therapy with N-carbamyl-L-glutamate (a registered drug, the analog of NAG). Our results, together with additional present and prior site-directed mutagenesis data for other residues mapping in this domain, suggest an NAG-triggered conformational change in the β4-α4 loop of the C-terminal domain of this enzyme. This change might be an early event in the NAG activation process. Molecular dynamics simulations that were restrained according to the observed effects of the mutations are consistent with this hypothesis, providing further backing for this structurally plausible signaling mechanism by which NAG could trigger urea cycle activation via CPS1.

Keywords: Allosteric regulation; Carbamoyl phosphate synthetase 1; Enzyme; Hyperammonemia; Inborn errors; Restrained molecular dynamics; Site-directed mutagenesis; Urea cycle diseases.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Ammonia / metabolism*
  • Carbamoyl-Phosphate Synthase (Ammonia) / chemistry*
  • Carbamoyl-Phosphate Synthase (Ammonia) / genetics
  • Carbamoyl-Phosphate Synthase (Ammonia) / metabolism*
  • Carbamoyl-Phosphate Synthase I Deficiency Disease / enzymology
  • Carbamoyl-Phosphate Synthase I Deficiency Disease / genetics
  • Carbamoyl-Phosphate Synthase I Deficiency Disease / metabolism*
  • Carbamoyl-Phosphate Synthase I Deficiency Disease / pathology
  • Enzyme Stability
  • Glutamates / metabolism
  • Humans
  • Kinetics
  • Molecular Dynamics Simulation
  • Mutation
  • Protein Structure, Tertiary
  • Signal Transduction
  • Urea / metabolism*


  • Glutamates
  • Ammonia
  • Urea
  • Carbamoyl-Phosphate Synthase (Ammonia)
  • N-acetylglutamic acid