Mutation analysis of the 6‐pyruvoyl‐tetrahydropterin synthase gene in Chinese hyperphenylalaninemia caused by tetrahydrobiopterin synthesis deficiency

Abstract

Hyperphenylalaninemia (HPA) may be caused by deficiency of phenylalanine hydroxylase or tetrahydrobiopterin (BH4), the essential cofactor for the aromatic amino acid hydroxylases. 6-Pyruvoyl-tetrahydropterin synthase (PTPS) deficiency is a major cause of BH4 deficient HPA. In this study, seven single base mutations at nucleotides 73 (C>G), 155 (A>G), 166 (G>A), 209 (T>A), 259 (C>T), 286 (G>A), and 317 (C>T) on PTPS cDNA were detected in Chinese PTPS-deficient HPA by polymerase chain reaction and solid phase DNA sequencing. These nucleotide alterations result in R25G, N52S, V56M, V70D, P87S, D96N, and T106M amino acid substitutions, respectively. The R25G, V56M, V70D, and T106M were novel mutations found in PTPS gene. By analysis of 38 PTPS mutant alleles from 19 unrelated Chinese PTPS-deficient HPA families, the allele frequency of these mutations in Chinese PTPS-deficient HPA were determined to be approximately 5.3% (R25G), 34.2% (N52S), 7.9% (V56M), 2.6% (V70D), 36.8% (P87S), 7.9% (D96N), and 2.6% (T106M), respectively. Two common mutations, N52S and P87S, were found to account for 71% of the Chinese PTPS mutant alleles. The N52S mutation accounts for 48% of the southern Chinese PTPS mutation, but only one (9%) of the northern Chinese PTPS mutant allele was found to be N52S, which suggested that the N52S mutation might be southern Chinese. Clinically, the V56M mutation was found to associate with the mild form of PTPS deficiency. However, the R25G, N52S, P87S, and D96N were found mainly in the patients with severe clinical symptom. Using polymerase chain reaction-based mutation analysis, a fetus at risk of PTPS deficiency was diagnosed prenatally to be a carrier of N52S mutation.

Mutation spectrum of and founder effects affecting the PTS gene in East Asian populations

“…14 In some cases, genomic DNA was extracted from blood samples, cultured fibroblasts or lymphoblasts by standard protocols. The DNA fragments for all patients as well as the normal controls were amplified by PCR followed by bidirectionally fluorescent sequencing on an ABI 377 or ABI 3730xl DNA analyzer (Life Technologies, Carlsbad, CA, USA) with BigDye chemistry (Life Technologies).…”

Mutation Profile of the MUT Gene in Chinese Methylmalonic Aciduria Patients

“…The method for genomic DNA isolated from dried blood spot sample has been described previously (Hong et al 1999;Liu et al 1998). All 13 exons of the MUT gene, including coding and non-coding exons, and exon-intron boundary sequences were PCR-amplified and directly sequenced by thermocycle sequencing using the BigDye ® terminator 3.1 sequencing kit (Applied Biosystems, Foster City, CA, USA) following analysis by an ABI 3730XL DNA Analyzer (Applied Biosystems).…”

Mutation spectrum of MMACHC in Chinese patients with combined methylmalonic aciduria and homocystinuria

“…The method for genomic DNA isolated from a dried blood spot collected on filter paper has been described elsewhere. [20][21] The whole coding region sequence and exon-intron boundary sequence of the MMACHC gene were PCR amplified using primer sets for exons 1, 2, 3 and 4a as described by Lerner-Ellis et al 7 For patients with only one mutation identified in the coding exons and flanking sequence, sequence analysis was extended into the 3¢-untranslated region (exons 4b, 5a and 5b in Lerner-Ellis et al 7 ). The four coding exons and exon-intron boundary sequence of the MMACHC gene were also sequenced in 70 anonymous Chinese controls.…”