Purification and Properties of Two 2‐Oxoacid:Ferredoxin Oxidoreductases from Halobacterium halobium

Abstract

Pyruvate: ferredoxin oxidoreductase and 2-oxoglutarate: ferredoxin oxidoreductase were obtained from cell-free extracts of Halobacterium halobium as homogeneous proteins after ammonium sulfate precipitation, salting-out chromatography with ammonium sulfate on unsubstituted agarose, gel filtration and chromatography on hydroxyapatite. The respective molecular weights are 256000 and 248000. Both enzymes consist of two sets of nonidentical subunits of M_r 86000 and 42000 in the case of the pyruvate-degrading enzyme and of 88000 and 36000 in the case of the 2-oxoglutarate-degrading enzyme. Analyses indicate that an intact enzyme molecule contains two [4 Fe-4S]^2+(2+.1+) clusters and two molecules of thiamin diphosphate. Flavin nucleotides, lipoic acid and pantetheine are absent. Thus the enzymes are very similar to the 2-oxoacid: ferredoxin oxidoreductases from fermentative and photosynthetic anaerobes described previously, but are clearly different from the 2-oxoacid dehydrogenase multienzyme complexes which commonly occur in aerobic organisms.