Phenylalanine hydroxylase deficiency in the Slovak population: Genotype–phenotype correlations and genotype-based predictions of BH4-responsiveness |
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Authors: | Emil Polak Andrej Ficek Jan Radvanszky Andrea Soltysova Otto Urge Eleonora Cmelova Dana Kantarska Ludevit Kadasi |
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Institution: | 1. Comenius University, Faculty of Natural Sciences, Department of Molecular Biology, Mlynska Dolina, 842 15 Bratislava, Slovak Republic;2. Institute of Molecular Physiology and Genetics, Slovak Academy of Science, Vlarska 5, 833 34 Bratislava, Slovak Republic;3. University Hospital Bratislava, A.Getlik clinic for children and adolescents, Antolska 11, 851 07 Bratislava, Slovak Republic;4. University Hospital Bratislava, Department of Clinical Genetics, Limbova 5, 833 05 Bratislava, Slovak Republic;5. Faculty Hospital of F.D.Roosvelt, Department of Medical Genetics, Nam. L. Svobodu 1, 975 17 Banska Bystrica, Slovak Republic |
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Abstract: | We investigated the mutation spectrum of the phenylalanine hydroxylase gene (PAH) in a cohort of patients from 135 Slovak PKU families. Mutational screening of the known coding region, including conventional intron splice sites, was performed using high-resolution melting analysis, with subsequent sequencing analysis of the samples showing deviated melting profiles compared to control samples. The PAH gene was also screened for deletions and duplications using MLPA analysis. Forty-eight different disease causing mutations were identified in our patient group, including 30 missense, 8 splicing, 7 nonsense, 2 large deletions and 1 small deletion with frameshift; giving a detection rate of 97.6%. The most prevalent mutation was the p.R408W, occurring in 47% of all alleles, which concurs with results from neighboring and other Slavic countries. Other frequent mutations were: p.R158Q (5.3%), IVS12 + 1G>A (5.3%), p.R252W (5.1%), p.R261Q (3.9%) and p.A403V (3.6%). We also identified three novel missense mutations: p.F233I, p.R270I, p.F331S and one novel variant: c.− 30A>T in the proximal part of the PAH gene promoter. A spectrum of 84 different genotypes was observed and a genotype based predictions of BH4-responsiveness were assessed. Among all genotypes, 36 were predicted to be BH4-responsive represented by 51 PKU families. In addition, genotype–phenotype correlations were performed. |
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Keywords: | BH4 tetrahydrobiopterin (6RL-erythro-5 6 7 8-tetrahydrobiopterin) Bp base pairs BIOPKUdb International Database of Patients and Genotypes Causing HPA/PKU including BH4-responsive Phenotype Conc concentration cPKU classic phenylketonuria dNTP deoxyribonucleoside triphosphate EDTA ethylenediaminetetraacetic acid HRM High Resolution Melting HumVar human variation MC melting curve MHP mild hyperphenylalaninemia MLPA Multiplex Ligation-dependent Probe Amplification mPKU mild phenylketonuria mRNA messenger RNA HPA hyperphenylalaninemia Kb kilobases PAH gene encoding phenylalanine hydroxylase PAH phenylalanine hydroxylase PAHdb PAH Locus Knowledgebase PAHvdb International database of variations in Phenylalanine Hydroxylase gene PCR polymerase chain reaction Phe phenylalanine PKU phenylketonuria PRA predicted residual activity RFLP restriction-fragment length polymorphism Ta annealing temperature Taq Thermus aquaticus Tyr tyrosine U unit |
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