Molecular assessment and bioinformatic analysis of two common mutations of phenylalanine hydroxylase (PAH) gene by HRM

Phenylketonuria (PKU) is the most prevalent inborn error of amino acid metabolism in the world and its incidence rate is increasing in Iran. The aim of the present study was to assess the efficiency of HRM technique as a fast and suitable method in identifying common mutations of phenylalanine hydroxylase gene including IVS10-11G>A and P281L in order to improve the early detection of the disease to prevent the occurrence of it.
In this case-control study, 20 DNA samples including one sample with IVS10-11G>A mutation , one sample with P281L mutation and 18 control samples were extracted from peripheral blood collected in Medical Genetic Center of Isfahan and were genotyped with HRM technique. To validate the mutations, the mutant samples were genotyped using sequencing. Bioinformatic analyses were used for determining structural and functional effects of P281L mutation on the of PAH protein.
FINDINGS: HRM analysis identified IVS10-11G>A and P281L mutations with a sensitivity and specificity of 100% and the mutant and normal samples differentiated well in normalized and difference plots. Bioinformatic analyses demonstrated instability and pathological effects of PAH protein containing P281L mutation.
HRM is a simple and fast technique detecting the two IVS10-11G>A and P281L mutations with 100% sensitivity and specificity.