Properties of the two terminal oxidases of Escherichia coli

Abstract

Proton translocation coupled to oxidation of ubiquinol by O2 was studied in spheroplasts of two mutant strains of Escherichia coli, one of which expresses cytochrome d, but not cytochrome bo, and the other expressing only the latter. O2 pulse experiments revealed that cytochrome d catalyzes separation of the protons and electrons of ubiquinol oxidation but is not a proton pump. In contrast, cytochrome bo functions as a proton pump in addition to separating the charges of quinol oxidation. E. coli membranes and isolated cytochrome bo lack the CuA center typical of cytochrome c oxidase, and the isolated enzyme contains only 1Cu/2Fe. Optical spectra indicate that high-spin heme o contributes less than 10% to the reduced minus oxidized 560-nm band of the enzyme. Pyridine hemochrome spectra suggest that the hemes of cytochrome bo are not protohemes. Proteoliposomes with cytochrome bo exhibited good respiratory control, but H+/e- during quinol oxidation was only 0.3-0.7. This was attributed to an "inside out" orientation of a significant fraction of the enzyme. Possible metabolic benefits of expressing both cytochromes bo and d in E. coli are discussed.

Role of Cytochrome bd Oxidase from Corynebacterium glutamicum in Growth and Lysine Production

“…Although inhibitory effects due to the overproduction of four membrane proteins cannot be excluded, the more likely reason for the reduced growth is the difference in bioenergetic efficiency between the cytochrome bc 1 -aa 3 supercomplex and cytochrome bd oxidase (6). As outlined previously (3), the former pathway should couple transport of two electrons from menaquinol to oxygen to the transfer of six protons from the cytoplasm to the outside, whereas the latter pathway can transfer only two protons to the outside per two electrons transported to oxygen (20,28) (Fig. 1).…”

Mutation of Arg-54 Strongly Influences Heme Composition and Rate and Directionality of Electron Transfer in Paracoccus denitrificans Cytochrome c Oxidase

“…The concentrations of different heme species were determined using the following extinction coefficients for the reduced-minus-oxidized difference spectra (31): ⑀ 587-620 ϭ 25 mM Ϫ1 cm Ϫ1 for heme A and ⑀ 552-536 ϭ 24 mM Ϫ1 cm Ϫ1 for heme O. The latter value was based on the assumption that the peak-to-trough absorbance difference is similar for hemes B and O (32). HPLC 1 Heme Analysis-Hemes were extracted from the isolated enzyme complexes essentially as described (33,34).…”

Time-resolved electrometric and optical studies on cytochrome bd suggest a mechanism of electron-proton coupling in the di-heme active site

“…Unlike the heme-copper oxidases (4,5), cytochrome bd does not pump protons (6). However, the oxidation of ubiquinol by O 2 catalyzed by cytochrome bd is linked to the generation of transmembrane electric potential difference (⌬⌿) (7)(8)(9)(10)(11) by virtue of the fact that the protons resulting from the oxidation of ubiquinol are released into the bacterial periplasm, whereas the protons used to convert O 2 to H 2 O are taken from the bacterial cytoplasm.…”