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

Area of Science:

• Biochemistry
• Genetics
• Metabolic Disorders

Background:

• Carbamoyl-phosphate synthetase I (CPS1) deficiency (CPS1D) is a severe, recessively inherited urea cycle disorder.
• Mutations in the CPS1 gene lead to hyperammonemia, posing life-threatening risks.

Purpose of the Study:

• To establish a recombinant CPS1 expression and purification system for assessing the pathogenicity of CPS1 missense mutations.
• To investigate the functional impact of nine clinical mutations and one polymorphism on CPS1 enzyme properties.

Main Methods:

• Utilized a baculovirus and insect cell expression system for recombinant CPS1 production.
• Assessed CPS1 solubility, stability, activity, and kinetic parameters for N-acetylglutamate (NAG).
• Analyzed five novel mutations (p.T471N, p.Q678P, p.P774L, p.R1453Q, p.R1453W) in severe CPS1D patients.

Main Results:

• p.P774L, p.R1453Q, and p.R1453W mutations inactivated CPS1.
• p.T471N and p.Y1491H mutations significantly reduced NAG affinity.
• p.Q678P mutation impaired enzyme folding; other mutations moderately decreased activity. p.G1376S confirmed as a polymorphism.

Conclusions:

• The novel CPS1 system effectively evaluates missense mutation effects, correlating in vitro findings with clinical observations.
• This system identifies CPS1D patients potentially benefiting from NAG analogue therapy due to reduced NAG affinity.
• The study provides valuable insights into CPS1 structure-function relationships and disease mechanisms.