期刊名称：Journal of Nutritional Science and Vitaminology
印刷版ISSN：0301-4800
电子版ISSN：1881-7742
出版年度：1992
卷号：38
期号：Special
页码：397-400
DOI：10.3177/jnsv.38.Special_397
出版社：Center for Academic Publications Japan
摘要：Genomic clones encompassing the entire genes for the human pyruvate dehydrogenase α and β subunits (PDHα or β) have been isolated by screening the leukocyte genomic libraries with a nick-translated human foreskin fibroblast PDHα or β cDNA probe. These genomic clones were characterized by restriction enzyme analysis, extensive DNA sequencing and primer extension analysis. The PDHα gene spans 17.08 kilobases and is composed of 11 exons and 10 introns within its coding region. The 18-kilobase clone of PDHβ gene is composed of 10 exons and 9 introns. All intron-exon splice junctions of two genes follow the GT/ AG rule. A total of seven Alu repeats in the PDHα gene were found in five introns and two Alu family in the PDHβ gene were found in intron 2 and 8. The 5'-flanking region of the PDHα gene contains typical CCAAT and TATA-like consensus promoter sequence and two Sp1 binding sequences. That of the PDHβ gene contains a TCAAT sequence but no TATA sequence. Primer extension analyses indicated that the PDHα and β genes transcription start sites are thymine and adenine residues located 124 and 132 bases upstream from initiation codon in exon 1, respectively. Genomic DNA of patient, died 93 hours after birth with acidemia and defect of PDH activity, was isolated and all of the exons of PDHα and β genes were amplified by PCR. Two single mutations in PDHα gene, TGT to TTT at codon 116 of Exon 5 and CTG to CCG at codon 162 of Exon 6, were identified by sequence analysis and resulted in a Cys to Phe and Leu to Prosubstitution in PDHα protein. The mutation causes the loss of PDH activity and missing two subunit protein bands revealed by immunoblot analysis.
