The genes for anabolic 2-oxoglutarate: Ferredoxin oxidoreductase from Hydrogenobacter thermophilus TK-6

2-Oxoglutarate: ferredoxin oxidoreductase (OGOR) of a thermophilic, chemoli thoautotrophic hydrogen-oxidizing bacterium, Hydrogenobacter thermophilus T K-6, is involved in carbon dioxide fixation via a reductive TCA cycle. The enzyme is a heterodimer comprising subunits of 70 and 35 kDa. The structura l genes for the subunits (korAB) were cloned with primers designed from N-t erminal sequences of the purified enzyme. The korAB genes were followed by two open reading frames of unknown function (orf3 and orf4). KorA carried a binding motif for coenzyme A, and KorB carried binding motifs for Fe-S clu ster and thiamine pyrophosphate. Active recombinant enzyme from korAB was p roduced in E. coli under the control of the lac promoter. orf3 and orf4 wer e not necessary for the expression of active enzyme in E. coli. The recombi nant enzymes had high substrate specificity toward a-oxoglutarate as in the case of the native enzyme purified from strain TK-6. (C) 2001 Academic Pre ss.