ビタミンB群関連酵素異常症の分子生物学的研究

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Holocarboxylase synthetase (HCS) plays an essential role in biotin utilization in cells and its deficiency causes biotin-responsive multiple carboxylase deficiency in humans. We have clones the human HCS cDNA, which maps to chromosome 21 q 22. 1. Two mutations in the HCS genes of Japanese patients with HCS deficiency have been identified : a transition from T to C which causes an amino acid substitution of proline for leucine at position 237 (L237P) and a single guanine base deletion (ΔG 1067) followed by premature termination. Transient expression in cultured fibroblasts from a patient after site-directed mutagenesis showed that the L237P mutation was responsible for decreased HCS activity. Hybridization analysis using allele specific oligonucleotide probes demonstrated that the prevalence of the mutations-L237P and ΔG 1067-was 50% and 30%, respectively, among Japanese patients with HCS deficiency. Methylmalonyl CoA mutase (MCM) is an adenosylcobalamin dependent enzyme. Deficiency of the MCM apoenzyme causes an autosomal recessive disorder known as methylmalonic acidemia (MMA). Five of eight Japanese patients with MMA expressed remarkably decreased amount of MCMmRNA which could not be detected by Northern blot analysis. We were able to amplify and quantify the trace amount of mRNA (0.1-1.0%) by RT-PCR coupled with fluorescent analyzer. The trace amount of MCMmRNA was amplified by nested PCR to obtain sufficient amount of cDNA for sequencing analysis. Sequencing of the amplified cDNA revealed two novel mutations : a nonsense mutation (G to T at nucleotide position 425) and 2bp deletion (ΔCA 769) which resulted in the premature termination by frame shift. The nonsense mutation (G425T) accounts for about 20% of all MMA alleles in the Japanese population.
日本ビタミン学会の論文
1996-01-25

ビタミンB群関連酵素異常症の分子生物学的研究

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