Over-expression of cbaAB genes of Bacillus stearothermophilus produces a two-subunit SoxB-type cytochrome c oxidase with proton pumping activity

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Abstract

We constructed expression plasmids containing cbaAB, the structural genes for the two-subunit cytochrome bo3-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 bo3-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 aa3-type oxidases belonging to the SoxM-type.

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Nikaido, K., Sakamoto, J., Noguchi, S., & Sone, N. (2000). Over-expression of cbaAB genes of Bacillus stearothermophilus produces a two-subunit SoxB-type cytochrome c oxidase with proton pumping activity. Biochimica et Biophysica Acta - Bioenergetics, 1456(1), 35–44. https://doi.org/10.1016/S0005-2728(99)00102-4

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