Molecular characterization of phenylketonuria in Japanese patients

We characterized phenylalanine hydroxylase (PAH) genotypes of Japanese pati ents with phenylketonuria (PKU) and hyperphenylalaninemia (HPA). PKU and HP A mutations in 41 Japanese patients were identified by denaturing gradient gel electrophoresis and direct sequencing, followed by restriction fragment length polymorphism analysis to find a large deletion involving exons 5 an d 6. Of 82 mutant alleles, 76 (92%) were genotyped showing 21 mutations. Th e major mutations were R413P (30.5%), R243Q (7.3%), R241C (7.3%), IVS4nt-1 (7.3%), T278I (7.3%), E6nt-96A-->g (6.1%), Y356X (4.9%), R111X (3.7%), and 442-706delE5/6 (2.4%). Eight new mutations (L52 S, delS70, S70P, Y77X, IVS3 nt-1, A132 V, W187 C, and C265Y) and a polymorphism of IVS10nt-14 were dete cted. In vitro PAH activities of mutant PAH cDNA constructs were determined by a COS cell expression system. Six mutations, viz., R408Q, L52 S, R241 C , S70P, V388 M, and R243Q, had 55%, 27%, 25%, 20%, 16% and 10% of the in vi tro PAH activity of normal constructs, respectively . The mean pretreatment phenylalanine concentration (0.83+/-0.21 mmol/l) of patients carrying the R408Q, R241 C, or L52 S mutation and a null mutation was significantly lowe r (P<0.0005) than that (1.99+/-0.65 mmol/l) of patients with both alleles c arrying mutations associated with a severe genotype. Simple linear regressi on analysis showed a correlation between pretreatment phenylalanine concent rations and predicted PAH activity in 29 Japanese PKU patients (y=31.9-1.03 x, r=0.59, P<0.0001). Genotype determination is useful in the prediction of biochemical and clinical phenotypes in PKU and can be of particular help i n managing patients with this disorder.