Genetic diagnosis and molecular characterization of three novel variations in the phenylalanine hydroxylase gene from Chinese patients with phenylketonuria.

Loss-of-function variants in the human phenylalanine hydroxylase (PAH) gene are the most common genetic causal factors for Phenylketonuria (PKU). Currently, a broad spectrum of variations is recognized in the human PAH gene. However, the molecular function and clinical significance of some novel PAH variants remain unclear. Here, we report on five PKU-affected families carrying three novel PAH variants, including one missense variant (PAH: c.271C>A (p.Leu91Met)) and two deletions (PAH: c.206_208delCTT (p.Ser70del) and PAH: c.541_544delGAGG (p.Glu181Lysfs*13)). These variations constitute different compound heterozygous genotypes with other known pathogenic variants such as PAH: c.721C>T (p.Arg241Cys), PAH: c.168+5G>C, and PAH: c.1238G>C (p.Arg413Pro), which probably led to the patients' PKU etiopathology. qRT-PCR and immunoblotting showed that the protein levels of PAH (S70del) and PAH (E181Kfs*13) were significantly reduced compared with the wild-type control, although their transcript levels were not. Also, the enzyme activity of PAH (S70del) and PAH (E181Kfs*13) mutants was significantly decreased relative to the wild type (P < 0.001). PAH: c.271C>A (p.Leu91Met) had no significant effect on PAH mRNA and protein levels or enzyme activity. Collectively, our data demonstrate that the two deletions PAH: c.206_208delCTT and PAH: c.541_544delGAGG are clinically significant for pathogenicity. Our findings are anticipated to contribute to the advancement of prenatal diagnosis, population-based carrier screening, and genetic counseling for individuals affected by PKU, and is expected to help reduce the incidence of PKU and ameliorate the associated disease burden. See also the graphical abstract(Fig. 1).