Molecular cloning of the Corynebacterium glutamicum ('Brevibacterium lactofermentum' AJ12036) odhA gene encoding a novel type of 2-oxoglutarate dehydrogenase.
The Corynebacterium glutamicum ('Brevibacterium lactofermentum' AJ12036) odhA gene, encoding 2-oxoglutarate dehydrogenase (E1o subunit of the 2-oxoglutarate dehydrogenase complex), has been isolated and identified as an homologous counterpart of the Escherichia coll sucA and Bacillus subtilis odhA genes. The nucleotide sequence of a 4394 bp chromosomal fragment containing the C. glutamicum odhA gene was determined. The odhA gene comprised 3771 bp (1257 codons, including the initiation codon) and a molecular mass of 138656 Da was predicted for the OdhA polypeptide. Northern blot analysis revealed a 3.9 kb transcript. The size of the transcript, together with the presence of a rho-independent terminator-like structure, suggests that C. glutamicum odhA is monocistronic. Cells harbouring plasmids carrying C. glutamicum odhA showed a threefold increase in specific 2-oxoglutarate dehydrogenase complex activity and expression of a protein with an apparent molecular mass of 136 kDa, in good agreement with the predicted size of the OdhA polypeptide. The C-terminal region of the C. glutamicum OdhA protein shows strong sequence similarity to E1os from other organisms. C. glutamicum OdhA has an N-terminal extension not found in previously reported E1os. The amino acid sequence of this extension shows similarity to that of the C-terminal region of dihydrolipoamide S-succinyltransferase (E2o) subunits of 2-oxoglutarate dehydrogenase complexes and dihydrolipoamide S-acetyltransferase (E2p) subunits of pyruvate dehydrogenase complexes. It suggests that the C. glutamicum odhA gene might encode a novel bifunctional protein with E1o and E2o activities.
Usuda
Y
Y
Central Research Laboratories, AJINOMOTO Co., Inc., Kawasaki-shi, Japan.
Tujimoto
N
N
Abe
C
C
Asakura
Y
Y
Kimura
E
E
Kawahara
Y
Y
Kurahashi
O
O
Matsui
H
H
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology; D001483:Base Sequence; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D003062:Codon; D003352:Corynebacterium / Q000201:enzymology* / Q000235:genetics*; D005798:Genes, Bacterial; D007655:Ketoglutarate Dehydrogenase Complex / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011768:Pyruvate Dehydrogenase Complex / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid
D003062:Codon; D046911:Macromolecular Substances; D011768:Pyruvate Dehydrogenase Complex; D011994:Recombinant Proteins; D007655:Ketoglutarate Dehydrogenase Complex