Identification of five novel inactivating mutations in the human thyroid peroxidase gene by denaturing gradient gel...

Thyroid peroxidase (TPO) is the key enzyme in the synthesis of thyroid hormones. Defects in the TPO gene are reported to be the cause of congenital hypothyroidism due to a Total Iodide Organification Defect (TIOD). This type of defect, where iodide taken up by the thyroid gland cannot be oxidized and bound to protein, is the most common hereditary inborn error causing congenital hypothyroidism in the Netherlands. Denaturing Gradient Gel Electrophoresis (DGGE) of PCR amplified genomic DNA was used to screen for mutations in the TPO gene of TIOD patients from nine apparently unrelated families, and seven different mutations were detected. Three frameshift mutations were found: a 20 bp duplication in exon 2, a 4 bp duplication in exon 8, and an insertion of a single nucleotide (C) at pos. 2505 in exon 14. In addition, four single nucleotide substitutions were identified: one single-base mutation resulted in a premature termination codon (C-->T at pos. 1708 in exon 10), two single-base substitutions changed an amino acid in highly conserved regions of the gene (Tyr-->Asp in exon 9 and Glu-->Lys in exon 14). The fourth single-base mutation located at the exon 10/intron 10 border altered a conserved Gly into Ser and could also affect splicing. Nine TIOD patients from five families were compound heterozygotes and six patients from four families were homozygous for one of the mentioned mutations in the TPO gene.