title: The Missense p.S231F phenylalanine hydroxylase gene mutation causes complete loss of enzymatic activity in vitro.
abstract: Phenylketonuria (PKU), the most frequent disorder of amino acid metabolism, is caused by mutations in human phenylalanine hydroxylase gene (PAH), leading to deficient enzyme activity. Previously reported but uncharacterized PAH gene mutation, p.S231F (c.692C > T), was detected in Serbian patients with classical PKU. We analyzed p.S231F PAH protein in prokaryotic (Escherichia coli) and eukaryotic expression system (hepatoma cells). In both systems the mutant enzyme was unstable. Residual enzyme activity in vitro was approximately 1%. Mutation p.S231F PAH was not activated by pre-incubation with phenylalanine substrate. We found no GroEL/GroES chaperone effect and slightly positive effect of the (6R)-L-erythro-5,6,7,8-tetrahydrobiopterin (BH(4)) on the stabilization of the protein structure. Our findings were in accordance with severe patients' phenotypes. In conclusion, p.S231F should be classified as a functionally null PAH gene mutation as it drastically reduces stability and activity of the PAH enzyme in vitro.
authors.affiliation: Institute of Molecular Genetics and Genetic Engineering, Vojvode Stepe 444a, 11000, Belgrade, Serbia.
authors.forename: Maja
authors.initials: M
authors.lastname: Stojiljkovic
authors.forename: Belén
authors.initials: B
authors.lastname: Pérez
authors.forename: Lourdes R
authors.initials: LR
authors.lastname: Desviat
authors.forename: Cristina
authors.initials: C
authors.lastname: Aguado
authors.forename: Magdalena
authors.initials: M
authors.lastname: Ugarte
authors.forename: Sonja
authors.initials: S
authors.lastname: Pavlovic
mesh_terms: D045744:Cell Line, Tumor; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D020125:Mutation, Missense; D010651:Phenylalanine Hydroxylase / Q000235:genetics* / Q000378:metabolism*; D010661:Phenylketonurias / Q000201:enzymology*
chemical_list: D010651:Phenylalanine Hydroxylase