Complex Mutation Analysis of the PAH Gene in Slovak Patients Affected by Phenylketonuria

Czech version

Authors: E. Polák ¹; A. Ficek ¹; M. Baldovič ^1,2; E. Feráková ¹; A. Šoltýsová ¹; J. Strnová ³; O. Ürge ³; L. Kovács ⁴; Ľ. Kádaši ^1,2
Authors‘ workplace: Katedra molekulárnej biológie PrírF UK, Bratislava vedúci doc. RNDr. Ľ. Kádasi, DrSc. ¹; Ústav molekulárnej fyziológie a genetiky SAV, Bratislava vedúci doc. RNDr. Ľ. Kádasi, DrSc. ²; Klinika deti a dorastu Andreja Getlíka. SZU, Bratislava prednostka doc. MUDr. K. Furková, CSc., mim. prof. ³; 2. detská klinika LFUK a DFNsP, Bratislava prednosta prof. MUDr. L. Kovács, DrSc, MPH ⁴
Published in: Čes-slov Pediat 2008; 63 (10): 528-534.
Category: Original Papers

Overview

This work continues previous studies to detect molecular basis of phenylketonuria in Slovak patients. DHPLC screening for all 13 exons of the PAH gene was introduced and optimized. DNA samples from 48 subjects with biochemically proven phenylketonuria were analyzed in whom direct DNA diagnostics (detection of 11 most frequent mutations) allowed to identify mutations on only one or none PKU chromosome. In this study, 16 new mutations were detected, (Q20X, F39L, R68S, F233I, R176X, P281L, R270I, T278N, R243X, A300S, I306V, W326X, L348V, S349P, E390G, Y387X), which were not known in this population as yet. Individual mutations were allocated to the appropriate phenotypic categories of phenylketonuria.

The present results will be applied for routine diagnostic practice of PKU in this country. In addition to causative mutations, several genetic polymorphisms were detected in introns, but also in the exons of the PAH gene. Up to date, 84,3% of Slovak PKU alleles are already identified, detection of the remaining 15,7 % needs additional studies and investigational approaches.

Key words:
phenylketonuria, mutations, mutational analysis, DNA diagnostics

Sources

1. Eisensmith RC, Woo SLC. Molecular basis of phenylketonuria and related hyperphenylalaninemias: Mutations and polymorphisms in the human phenylalanine hydroxylase gene. Hum. Mutat. 1992;1: 13–23.

2. Scriver C, et al. The Metabolic and Molecular Bases of Inherited Disease. New York: McGraw-Hill, 1995: 1015–1075.

3. Sršeň Š, Sršňová K. Základy klinickej genetiky a jej molekulárna podstata. Martin: Osveta, 2005: 445. ISBN 80-8063-185-9.

4. Kaufman S. Phenylketonuria: biochemical mechanisms. Adv. Neurochem. 1976;2: 1–132.

5. Tyfield LA, Meredith AL, Osborn MJ, Harper PS. Identification of the haplotype patern associated with the mutant PKU allele in the Gypsy population in Wales. J. Med. Genet. 1989;26: 450–499.

6. Bickel H, Bachmann C, Beckers R. Neonatal mass screening for metabolic disorders. Eur. J. Pediatr. 1981;137: 133–139.

7. Kozák L, Kuhrová V, Blažková M, Fajkusová L, Dvořáková D, Pijáčková A. Pokroky v diagnostice fenylketonurie zavedením přímé detekce mutací v PAH genu. Čas. Lék. čes. 1995;134: 385–387.

8. Kádasi Ľ, Poláková H, Feráková E, Krivušová T, Hudecová S, Szomolayová I, Strnová J, Hruškovič I, Ferák V. PKU in Slovakia: mutation screening and haplotype analysis. Hum. Genet. 1995;95: 112–114.

9. Gűttler, F. Hyperphenylalaninemia: Diagnosis and classification of the various types of phenylalanine hydroxylase deficiency. Acta Paediat. 1980;280: 1–80.

10. Gűttler F, Guldberg P. The influence of mutations on enzyme activity and phenylalanine tolerance in phenylalanine hydroxylase deficiency. Eur. J. Pediatr. 1996;155: S6–S10.

11. Guldberg P, Rey F, Zschocke J, Romano V, Baudouin F, et al. A European Multicenter Study of Phenylalanine Hydroxylase Deficiency: Classification of 105 mutations and a general system for genotype-based prediction of metabolic phenotype. Am. J. Hum. Genet. 1998;63: 71–79.

12. Woo SLC, Lidsky AS, Güttler F, Chandra T, Robson K. Cloned human phenylalanine hydroxylase gene allows prenatal diagnosis and carrier detection of classical phenylketonuria. Nature 1983;306: 151–155.

13. Lidsky AS, Law ML, Morse HG, Kao FT, Rabin M, et al. Regional mapping of the phenylalanine hydroxylase gene and the phenylketonuria locus in the human genome. Proc. Natl. Acad. Sci. USA 1985;82: 6221–6225.

14. DiLella AG, Marvit J, Lidsky AS, Güttler F, Woo SLC. Tight linkage between a splicing mutation and a specific DNA haplotype in phenylketonuria. Nature 1986;332: 799–803.

15. Daiger SP, Reed L, Huang SS, Zeng YT, Wang T, et al. Polymorphic DNA haplotypes at the phenylalanine hydroxylase (PAH) locus in Asian families with phenylketonuria (PKU). Am. J. Hum. Genet. 1989;45: 319–324.

16. Kádasi Ľ. Molekulárna genetika vybraných monogénne dedičných ochorení. Bratislava: Veda, 2005: 224. ISBN 80-224-0869-7.

17. Kayaalp E, Treacy E, Waters JP, Byck S, Nowacki P, Scriver CR. Human phenylalanine hydroxylase mutations and hyperphenylalaninemia phenotypes: A metanalysis of genotype-phenotype correlations. Am. J. Hum. Genet. 1997;61: 1309–1317.

18. http://www.pahdb.mcgill.ca

19. Erlandsen H, Patch MG, Gamez A, Straub M, Stevens RC. Structural studies on phenylalanine hydroxylase and implications toward understanding and treating phenylketonuria. Pediat. 2003;112: 1557–1565.

20. Richard I, Beckmann JS. How neutral are synonymous codon mutations? Nat. Genet. 1995;35: 301–304.

21. Chao HK, Hsiao KJ, Su TS. A silent mutation induces exon skipping in the phenylalanine hydroxylase gene in phenylketonuria. Hum. Genet. 2001;108: 14–19.

22. Okano Y, Eisensmith RC, Güttler F, Lichter Konecki U, et al. Molecular basis of phenotypic heterogenity in phenylketonuria. N. Engl. J. Med. 1991;324: 1232–1238.

23. Desviat LR, Perez B, Ugarte M. Molecular basis of non-PKU hyperphenylalaninaemia in Spain: prevalence of A403V, a mutation with high residual activity. J. Inherit. Metab. Dis. 1996;19: 227–230.

24. Gűttler F, Guldberg P. Mutation analysis anticipates dietery requirements in phenylketonuria. Eur. J. Pediatr. 2000;159: 150–153.

25. Feráková E, Ferák V, Kadáši Ľ, Poláková H, Hejcmanová L, Pijačková A. A unique RFLP haplotype at the phenylalanine hydroxylase locus in Czechoslovak Gypsies with phenylketonuria. Funct. Develop. Morph. 1992;2: 139–140.

26. Jeruzelska J, Matuszak R, Lyonnet S, Rey F, Rey J, et al. Genetic background of clinical homogenity of phenylketonuria in Poland. J. Med. Genet. 1993;30: 232–234.

27. http://frodo.wi.mit.edu/

28. Búcová, D. Polymorfizmus typu STR v géne pre fenylalanín hydroxylázu a jeho využitie pri diagnostike fenylketonúrie v slovenskej populácii. Diplomová práca. Bratislava: PrírF UK, 1996.

29. Feráková E, Kádaši Ľ. Multiple origin for PKU in Slovakia. Symposium “The Origin of the Hungarian Populations and Mitochondrial Diseases”. Budapest, 1996.

30. Kojšová S. Detekcia mutácií L48S, I65T, R243X a Y414C v géne pre fenylalanín hydroxylázu v súbore slovenských pacientov s fenylketonúriou. Rigorózna práca. Bratislava: PrírF UK, 2004.

31. Bräutigam S, Kujat A, Kirst A, Seidel J, Lüleyap ÜH, Froster UG. DHPLC mutation analysis of phenylketonuria. Mol. Gen. Metab. 2003;78: 205–210.

32. Dobrowolski SF, Ellingson C, Coyne T, Grey J, Martin R, et al. Mutations in the phenylalanine hydroxylase gene identified in 95 patients with phenylketonuria using novel systems of mutation scanning and specific genotyping based upon thermal melt profiles. Mol. Genet. Metab. 2007;10: 12–22.

33. Kozák L, Blažková M, Kuhrová V, Pijáčková A, Ružičková Š, Šťastná S. Mutation and haplotype analysis of the phenylalanine hydroxylase alleles in classical PKU patients from Czech Republic: identification of four novel mutations. J. Med. Genet. 1997;34: 893–898.

34. Barić I, Mardešić D, Sarnavska V, Lichter-Konecki U, Konecki DS, Trefz FK. Geographical distribution of the P281L mutation at the phenylalanine hydroxylase locus: possible origin in southeastern Europe. J. Inherit. Metab. Dis. 1994;17: 376–377.

35. Ürge O, Strnová J, Mosendzová B. Fenylketonúria u adolescentov a dospelých na Slovensku. Čes.-slov. Pediat. 2003;58: 423–425.