ISOLATION, CHARACTERIZATION AND STRUCTURAL ORGANIZATION OF THE GENE AND PSEUDOGENE FOR THE DIHYDROLIPOAMIDE SUCCINYLTRANSFERASE COMPONENT OF THE HUMAN 2-OXOGLUTARATE DEHYDROGENASE COMPLEX

In the present study, the dihydrolipoamide succinyltransferase gene of the 2-oxoglutarate dehydrogenase complex was isolated from a human ge nomic DNA library and its entire nucleotide sequence was determined. T his 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 initi ating translation codon was the transcription initiation site of the h uman dihydrolipoamide succinyltransferase mRNA. Sequence analysis of t he promoter-regulatory region showed the presence of a CAAT-box-like s equence 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 t he gene during evolution. Moreover, a processed pseudogene of dihydrol ipoamide succinyltransferase was isolated and sequenced. The nucleotid e sequence of the pseudogene is 93% similar to the sequence of the hum an dihydrolipoamide succinyltransferase cDNA, but the pseudogene is no t functional for base changes, deletions and insertions of the pseudog ene. Southern-blot analysis showed the presence of a single copy of th is gene and a single copy of a pseudogene in the human genome. In addi tion, a possible relationship between dihydrolipoamide succinyltransfe rase and familial Alzheimer's disease is discussed.