Over-expression of cbaAB genes of Bacillus stearothermophilus produces a two-subunit SoxB-type cytochrome c oxidase with proton pumping activity.
We constructed expression plasmids containing cbaAB, the structural genes for the two-subunit cytochrome bo(3)-type cytochrome c oxidase (SoxB type) recently isolated from a Gram-positive thermophile Bacillus stearothermophilus. B. stearothermophilus cells transformed with the plasmids over-expressed an enzymatically active bo(3)-type cytochrome c oxidase protein composed of the two subunits, while the transformed Escherichia coli cells produced an inactive protein composed of subunit I without subunit II. The oxidase over-expressed in B. stearothermophilus was solubilized and purified. The oxidase contained protoheme IX and heme O, as the main low-spin heme and the high-spin heme, respectively. Analysis of the substrate specificity indicated that the high-affinity site is very specific for cytochrome c-551, a cytochrome c that is a membrane-bound lipoprotein of thermophilic Bacillus. The purified enzyme reconstituted into liposomal vesicles with cytochrome c-551 showed H(+) pumping activity, although the efficiency was lower than those of cytochrome aa(3)-type oxidases belonging to the SoxM-type.
Nikaido
K
K
Department of Biochemical Engineering and Science, Kyushu Institute of Technology, 680-4 Kawazu, Iizuka, Fukuoka, Japan.
Sakamoto
J
J
Noguchi
S
S
Sone
N
N
D001426:Bacterial Proteins; D002384:Catalysis; D003573:Cytochrome b Group; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D003580:Cytochromes / Q000737:chemistry / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D001411:Geobacillus stearothermophilus / Q000235:genetics* / Q000378:metabolism; D010088:Oxidoreductases / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D017494:Proton Pumps / Q000235:genetics; D013379:Substrate Specificity
D001426:Bacterial Proteins; D003573:Cytochrome b Group; D003574:Cytochrome c Group; D003580:Cytochromes; D029968:Escherichia coli Proteins; D017494:Proton Pumps; C083850:cytochrome bo3, E coli; C017167:cytochrome C(551); D010088:Oxidoreductases; C064500:flavocytochrome c sulfide dehydrogenase; D003576:Electron Transport Complex IV