Isolation, Characterization and Structural Organization of the Gene and Pseudogene for the Dihydrolipoamide Succinyltransferase Component of the Human 2‐Oxoglutarate Dehydrogenase Complex

Abstract

In the present study, the dihydrolipoamide succinyltransferase gene of the 2-oxoglutarate dehydrogenase complex was isolated from a human genomic DNA library and its entire nucleotide sequence was determined. This gene was approximately 23 kbp in size with 15 exons and 14 introns. All of the donor and acceptor splice sites of this gene conformed to the GT/AG rule. A quanine residue 43 bases upstreams of the ATG initiating translation codon was the transcription initiation site of the human dihydrolipoamide succinyltransferase mRNA. Sequence analysis of the promoter-regulatory region showed the presence of a CAAT-box-like sequence but the presence of a TATA-box-like sequence was not evidenced. Also located in this region were sequences resembling glucocorticoid-responsive and CAMP-responsive elements, and an Sp1 binding site. No nucleotide sequence corresponding to the E3-binding and/or E1-binding domain was found in any region of the gene. Therefore, the exon coding for the E3-binding and/or E1-binding domain may have been lost from the gene during evolution. Moreover, a processed pseudogene of dihydrolipoamide succinyltransferase was isolated and sequenced. The nucleotide sequence of the pseudogene is 93% similar to the sequence of the human dihydrolipoamide succinyltransferase cDNA, but the pseudogene is not functional for base changes, deletions and insertions of the pseudogene. Southern-blot analysis showed the presence of a single copy of this gene and a single copy of a pseudogene in the human genome. In addition, a possible relationship between dihydrolipoamide succinyltransferase and familial Alzheimer's disease is discussed.