Understanding carbamoyl-phosphate synthetase I (CPS1) deficiency by using expression studies and structure-based analysis

Abstract

Carbamoyl‐phosphate synthetase I (CPS1) deficiency (CPS1D), a recessively inherited urea cycle error due to CPS1 gene mutations, causes life‐threatening hyperammonemia. The disease‐causing potential of missense mutations in CPS1 deficiency can be ascertained with the recombinant CPS1 expression and purification system reported here, which uses baculovirus and insect cells. We study with this system the effects of nine clinical mutations and one polymorphism on CPS1 solubility, stability, activity, and kinetic parameters for NAG. Five of the mutations (p.T471N, p.Q678P, p.P774L, p.R1453Q, and p.R1453W) are first reported here, in three severe CPS1D patients. p.P774L, p.R1453Q, and p.R1453W inactivate CPS1, p.T471N and p.Y1491H greatly decrease the apparent affinity for NAG, p.Q678P hampers correct enzyme folding, and p.S123F, p.H337R, and p.P1411L modestly decrease activity. p.G1376S is confirmed a trivial polymorphism. The effects of the C‐terminal domain mutations are rationalized in the light of this domain crystal structure, including the NAG site structure [Pekkala et al. Biochem J 424:211–220]. The agreement of clinical observations and in vitro findings, and the possibility to identify CPS1D patients who might benefit from specific treatment with NAG analogues because they exhibit reduced affinity for NAG highlight the value of this novel CPS1 expression/purification system. Hum Mutat 31:1–8, 2010. © 2010 Wiley‐Liss, Inc.

Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: Effects of CPS1 mutations that concentrate in a central domain of unknown function

“…1A) accounted for disease-causation because of impaired activity, stability or NAG activation. These results, which followed earlier pilot studies using baculovirus/insect cell-expressed rat CPS1 (a surrogate of human CPS1) [43], are extended now with those for 18 mutations affecting the UFSD.…”

“…We previously used the baculovirus/insect cell system utilized here for producing twenty CPS1 forms carrying different missense changes identified in CPS1D patients and spread all over the CPS1 protein except the UFSD [26,43]. We also reported [25] the effects of another five non-CPS1D-associated but functionally important mutations affecting the C-terminal domain (ASD).…”

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

“…); e When the contact was used as a restraint for model generation it is specified; f No clinical information available for the patient carrying this mutation; g Díez-Fernández et al, 2013; h Unknown when originally reported. New data gathered on the patient; i The late onset and coexistence with a null second allele (Q478*) indicate residual activity; j Pekkala et al, 2010; Not present in previous docking model (Pekkala et al, 2009); l The patient is alive, but the second CPS1 missense allele (Y959C) might not be inactivating; m The patient had a late onset presentation, but the second allele was not identified. …”

Carglumic acid for the treatment ofN-acetylglutamate synthase deficiency and acute hyperammonemia

“…This has been elaborated in studies identifying binding sites for NAG on the CPS1 protein [69,70]. The difficulty here will be to ascertain patients with CPS1 deficiency who will benefit from carglumic acid.…”