Prokaryotic expression analysis of I269L and R270K mutations of the phenylalanine hydroxylase gene

The I269L (c.805A→C) and R270K (c.809G→A) mutations in exon 7 of the human phenylalanine hydroxylase (PAH) gene were identified in the Portuguese phenylketonuric (PKU) population with a frequency of 0.4 % and 6.2 %, respectively. To confirm that these changes at the DNA level are responsible for the PKU phenotype presented by those patients, and to establish a correlation between the genotype and the presented phenotype, these two mutations were produced in a prokaryotic expression system and characterized. In the present study we show that, using the pTrcHis system, the R270K mutation results in a severe loss of PAH enzyme activity and that mutation I269L only causes a moderate reduction in the specific activity of the enzyme. The obtained results are compatible with the clinical/metabolic phenotype of the affected patients.