Large-scale purification and characterization of a highly active four-subunit cytochrome bc1 complex from Rhodobacter sphaeroides

Andrews, Katherine M.; Crofts, Antony R.; Gennis, Robert B.

doi:10.1021/bi00463a004

Cited by 86 publications

(83 citation statements)

References 53 publications

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“…The reduced minus oxidized difference spectrum of cytochrome bL displayed an asymmetric a band with an absorption maximum at 564 and a shoulder at 559 nm, as was also reported for cytochrome bL from R. rubrum (17). Cytochrome bH displayed the characteristic symmetric a band with an absorption maximum at 559.5 nm (1,15,17). The titration of c-type cytochromes (Fig.…”

supporting

confidence: 74%

Abundance, subunit composition, redox properties, and catalytic activity of the cytochrome bc1 complex from alkaliphilic and halophilic, photosynthetic members of the family Ectothiorhodospiraceae

et al. 1993

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Ubiquinol-cytochrome c oxidoreductase (cytochrome bej) complexes were demonstrated to be present in the membranes of the alkaliphilic and halophilic purple sulfur bacteria Ectothiorhodospira halophila, Ectothiorhodospira mobilis, and Ectothiorhodospira shaposhnikovii by protoheme extraction, immunoblotting, and electron paramagnetic resonance spectroscopy. The gy values of the Rieske [2Fe-2S] clusters observed in membranes of E. mobifis and E. halophila were 1.895 and 1.910, respectively. In E. mobilis membranes, the cytochrome be1 complex was present in a stoichiometry of approximately 0.2 per reaction center. This complex was isolated and characterized. It contained four prosthetic groups: low-potential cytochrome b (cytochrome bL; Em = -142 mV), high-potential cytochrome b (cytochrome bH; E. = 116 mV), cytochrome cl (Em = 341 mV), and a Rieske iron-sulfur cluster. The absorbance spectrum of cytochrome bL displayed an asymmetric a-band with a maximum at 564 nm and a shoulder at 559 nm. The a bands of cytochrome bH and cytochrome cl peaked at 559.5 and 553 nm, respectively. These prosthetic groups were associated with three different polypeptides: cytochrome b, cytochrome cl, and the Rieske iron-sulfur protein, with apparent molecular masses of 43, 30, and 21 kDa, respectively. No evidence for the presence of a fourth subunit was obtained.Maximal ubiquinol-cytochrome c oxidoreductase activity of the purified complex was observed at pH 8; the turnover rate was 57 mol of cytochrome c reduced (mol of cytochrome cj)-1 s-1. The complex showed a strikingly low sensitivity towards typical inhibitors of cytochrome bc complexes.

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supporting

confidence: 74%

Abundance, subunit composition, redox properties, and catalytic activity of the cytochrome bc1 complex from alkaliphilic and halophilic, photosynthetic members of the family Ectothiorhodospiraceae

et al. 1993

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“…Nevertheless, there are many precedents for it, such as Cytf (46), the c 556 High Potential heme of the photosynthetic RC from Blastochloris viridis (57), and the b L -heme of the bc 1 complex of Rb. sphaeroides (58).…”

Section: Discussionmentioning

confidence: 99%

On the Spatial Organization of Hemes and Chlorophyll in Cytochrome b 6 f

Schoepp¹,

Chabaud²,

Breyton³

et al. 2000

Journal of Biological Chemistry

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The organization of chromophores in the cytochrome b 6 f from Chlamydomonas reinhardtii has been studied spectroscopically. Linear dichroism (LD) measurements, performed on the complex co-reconstituted into vesicles with photosynthetic reaction centers as an internal standard, allow the determination of the orientations of the chromophore with respect to the membrane plane. The orientations of the b H -and b L -hemes are comparable to those determined crystallographically on the cytochrome bc 1 . The excitonic CD signal, resulting from the interaction between b-hemes, is similar to that reported for the cytochrome bc 1 . LD and CD data are consistent with the differences between the b 6 f and bc 1 leaving the orientation of the b-hemes unaffected.

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“…Some bacterial cytochromes bc 1 (for example in Rhodobacter capsulatus and Paracoccus denitrificans) can just be composed of these three subunits (21,142,216,272). Other cytochromes bc 1 may have accessory subunits, which number varies from one in bacterial cytochrome bc 1 (for example, in Rhodobacter sphaeroides) (6,273) to eight in mitochondrial cytochrome bc 1 (19,268). These subunits do not contain redox cofactors and are not involved in energy conversion.…”

Section: Cofactor Chains Of Cytochrome Bc 1 As a Frame For The Q mentioning

confidence: 99%

Electronic Connection Between the Quinone and CytochromecRedox Pools and Its Role in Regulation of Mitochondrial Electron Transport and Redox Signaling

Sarewicz

Osyczka

2015

Physiological Reviews

140

141

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Physiol Rev 95: 219 -243, 2015; doi:10.1152/physrev.00006.2014.-Mitochondrial respiration, an important bioenergetic process, relies on operation of four membranous enzymatic complexes linked functionally by mobile, freely diffusible elements: quinone molecules in the membrane and water-soluble cytochromes c in the intermembrane space. One of the mitochondrial complexes, complex III (cytochrome bc 1 or ubiquinol: cytochrome c oxidoreductase), provides an electronic connection between these two diffusible redox pools linking in a fully reversible manner two-electron quinone oxidation/reduction with one-electron cytochrome c reduction/oxidation. Several features of this homodimeric enzyme implicate that in addition to its well-defined function of contributing to generation of proton-motive force, cytochrome bc 1 may be a physiologically important point of regulation of electron flow acting as a sensor of the redox state of mitochondria that actively responds to changes in bioenergetic conditions. These features include the following: the opposing redox reactions at quinone catalytic sites located on the opposite sides of the membrane, the inter-monomer electronic connection that functionally links four quinone binding sites of a dimer into an H-shaped electron transfer system, as well as the potential to generate superoxide and release it to the intermembrane space where it can be engaged in redox signaling pathways. Here we highlight recent advances in understanding how cytochrome bc 1 may accomplish this regulatory physiological function, what is known and remains unknown about catalytic and side reactions within the quinone binding sites and electron transfers through the cofactor chains connecting those sites with the substrate redox pools. We also discuss the developed molecular mechanisms in the context of physiology of mitochondria.

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Large-scale purification and characterization of a highly active four-subunit cytochrome bc1 complex from Rhodobacter sphaeroides

Cited by 86 publications

References 53 publications

Abundance, subunit composition, redox properties, and catalytic activity of the cytochrome bc1 complex from alkaliphilic and halophilic, photosynthetic members of the family Ectothiorhodospiraceae

Abundance, subunit composition, redox properties, and catalytic activity of the cytochrome bc1 complex from alkaliphilic and halophilic, photosynthetic members of the family Ectothiorhodospiraceae

On the Spatial Organization of Hemes and Chlorophyll in Cytochrome b 6 f

Electronic Connection Between the Quinone and CytochromecRedox Pools and Its Role in Regulation of Mitochondrial Electron Transport and Redox Signaling

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