Molecular analysis of 16 Turkish families with DHPR deficiency using denaturing gradient gel electrophoresis (DGGE)

Dihydropteridine reductase (DHPR) catalyses the conversion of quinonoid dih ydrobiopterin (qBH(2)) to tetrahydrobiopterin (BH4), which serves as the ob ligatory cofactor for the aromatic amino acid hydroxylases, DHPR deficiency , caused by mutations in the QDPR gene, results in hyperphenylalaninemia an d deficiency of various neurotransmitters in the central nervous system, wi th severe neurological symptoms as a consequence. We have studied, at the c linical and molecular levels, 17 patients belonging to 16 Turkish families with DHPR deficiency. The patients were detected at neonatal screening for hyperphenylalaninemia or upon the development of neurological symptoms. To identify the disease causing molecular defects, we developed a sensitive sc reening method that rapidly scans the entire open reading frame and all spl ice sites of the QDPR gene. This method combines PCR amplification and "GC- clamping" of each of the seven exonic regions of QDPR, resolution of mutati ons by denaturing gradient gel electrophoresis (DGGE), and identification o f mutations by direct sequence analysis. A total of ten different mutations were identified, of which three are known (G23D, Y150C, R221X) and the rem aining are novel (G17R, G18D, W35fs, Q66R, W90X, S97fs and G149R). Six of t hese mutations are missense variants, two are nonsense mutations, and two a re frameshift mutations. All patients had homoallelic genotypes, which allo wed the establishment of genotype-phenotype associations. Our findings sugg est that DGGE is a fast and efficient method for detection of mutations in the QDPR gene, which may be useful for confirmatory DNA-based diagnosis, ge netic counselling and prenatal diagnosis in DHPR deficiency.