Three Molecules of Ubiquinone Bind Specifically to Mitochondrial Cytochrome bc 1Complex

Bartoschek, Stefan; Johansson, Malin; Geierstanger, Bernhard H.; Okun, Jürgen G.; Lancaster, C. Roy D.; Humpfer, Eberhard; Yu, Linda; Yu, Chao; Griesinger, Christian; Brandt, Ulrich

doi:10.1074/jbc.c100365200

Cited by 61 publications

(53 citation statements)

References 30 publications

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“…Bifurcated electron transport is thought to originate at these two quinones through a quinhydrone charge-transfer intermediate. However, the quinhydrone has never been directly observed and its possible involvement remains controversial (28)(29)(30)(31). Here we show evidence that a quinhydrone is found in DsbB.…”

Section: The Putative Quinhydrone In Dsbb[cccc] Responds To Ph In Thementioning

confidence: 55%

Disulfide bond formation involves a quinhydrone-type charge–transfer complex

Regeimbal

Gleiter

Trumpower

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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The chemistry of disulfide exchange in biological systems is well studied. However, the detailed mechanism of how oxidizing equivalents are derived to form disulfide bonds in proteins is not clear. In prokaryotic organisms, it is known that DsbB delivers oxidizing equivalents through DsbA to secreted proteins. DsbB becomes reoxidized by reducing quinones that are part of the membranebound electron-transfer chains. It is this quinone reductase activity that links disulfide bond formation to the electron transport system. We show here that purified DsbB contains the spectral signal of a quinhydrone, a charge-transfer complex consisting of a hydroquinone and a quinone in a stacked configuration. We conclude that disulfide bond formation involves a stacked hydroquinone-benzoquinone pair that can be trapped on DsbB as a quinhydrone charge-transfer complex. Quinhydrones are known to be redox-active and are commonly used as redox standards, but, to our knowledge, have never before been directly observed in biological systems. We also show kinetically that this quinhydronetype charge-transfer complex undergoes redox reactions consistent with its being an intermediate in the reaction mechanism of DsbB. We propose a simple model for the action of DsbB where a quinhydrone-like complex plays a crucial role as a reaction intermediate.M any extracellular proteins require the formation of one or more disulfide bonds to achieve their correct tertiary structure. Because of the importance of disulfides in the folding of many proteins, the process of disulfide formation in biological systems has been studied extensively. The DsbA-DsbB oxidation-reduction pathway is responsible for de novo disulfide formation in Escherichia coli, and has been heavily investigated by several groups for almost 15 years. As a result, much is known about this system. For recent reviews, see refs. 1 and 2.DsbA is a 21-kDa periplasmic protein that is a member of the thioredoxin superfamily. DsbA has a very reactive active-site high-redox-potential disulfide that can readily transfer oxidizing equivalents to a wide variety of substrate proteins (1, 2). When DsbA donates its disulfide bond to a secreted target protein, DsbA is released in a reduced state. To function as a catalyst, DsbA must be reoxidized. This reoxidation is accomplished by its partner protein, DsbB (3). DsbB is a 20-kDa innermembrane protein that is predicted to have four transmembrane helices and two periplasmic loops (4). DsbB has four conserved cysteines that are required for activity; the first pair is located in the N-terminal periplasmic loop and the second pair is located in the C-terminal periplasmic loop (5). The overall reaction that DsbB catalyzes is the formation of a disulfide bond within DsbA and the reduction of a membrane-bound quinone. This reaction provides a pathway for the oxidative power of the electron transport system to be used to form a disulfide within DsbA and ultimately within secreted proteins (3, 6, 7). The mechanism of DsbB begins with the reduction of a qu...

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Section: The Putative Quinhydrone In Dsbb[cccc] Responds To Ph In Thementioning

confidence: 55%

Disulfide bond formation involves a quinhydrone-type charge–transfer complex

Regeimbal

Gleiter

Trumpower

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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“…Generation of R. sphaeroides Cytochrome bc 1 Mutants-Mutations were constructed using the QuikChange site-directed mutagenesis kit (Stratagene) with supercoiled double-stranded pGEM7Zf(ϩ)-fbcB as a template. Forward and reverse primers were used for PCR amplification ( Table 1).…”

Section: Methodsmentioning

confidence: 99%

“…Mutant bc 1 Complexes-Because the cytochrome bc 1 complex is absolutely necessary for photosynthetic growth of R. sphaeroides, a mutant with a substitution at a critical position will not grow photosynthetically, whereas mutants with substitutions at noncritical positions will grow. Thus, observing the photosynthetic growth behavior, one can determine whether substitutions are at critical positions.…”

Section: Photosynthetic Growth Behaviors Of the Wild-type Andmentioning

confidence: 99%

“…The bc 1 complex/PL mixtures were incubated at 0°C for 30 min before overnight dialysis at 4°C against 100 volumes of 50 mM sodium phosphate buffer (pH 7.4) containing 100 mM KCl with three changes of buffer. The mixture was then dialyzed against 100 volumes of 150 mM KCl for 3-4 h. 1 Complex-PL Vesicles-Proton translocation coupled to electron flow through the bc 1 complex-PL vesicles was measured at room temperature using an Accumet Model 10 pH meter and Model 13-620-96 combination pH electrode. 25 nmol of Q 0 C 10 BrH 2 was added to the 1.6-ml reaction mixture containing 150 mM KCl, 4 M ferricytochrome c, 1 M valinomycin, and an appropriate amount of bc 1 complex-PL vesicles (30 -50 l).…”

Section: Primermentioning

confidence: 99%

“…It is an essential segment of cellular energy-conserving electron transport chains in animals, plants, and bacteria (1,2). This complex catalyzes electron transfer from ubiquinol to cytochrome c (or cytochrome c 2 in bacteria) and concomitantly translocates protons across the membrane to generate a membrane potential and pH gradient for ATP synthesis.…”

mentioning

confidence: 99%

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Effect of Mutations of Arginine 94 on Proton Pumping, Electron Transfer, and Superoxide Anion Generation in Cytochrome b of the bc1 Complex from Rhodobacter sphaeroides

Zhou

2013

Journal of Biological Chemistry

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Background:The bc 1 complex catalyzes electron transfer from ubiquinol to cytochrome c and translocates protons across the membrane. Results: Substitution of Arg-94 decreased the electron transfer activity and proton pumping capability and increased the superoxide production. Conclusion: Arg-94 is an important amino acid for the proton pumping capability of the bc 1 complex. Significance: The result is significant for understanding the catalytic mechanism of the bc 1 complex.

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Iron: Heme Proteins & Electron Transport

Durham

Millett

2005

Encyclopedia of Inorganic Chemistry

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This chapter discusses the structure and function of cytochromes, which are a class of iron‐containing heme proteins primarily involved in biological electron‐transfer reactions. Subcellular organelles called mitochondria are responsible for carrying out oxidative phosphorylation, the major energy‐transduction process in eukaryotic cells. Enormous progress has been made in our understanding of the molecular mechanism of the mitochondrial electron‐transport chain over the past decade. X‐ray crystal structures have been determined for three of the four electron‐transfer complexes: succinate–ubiquinone reductase, cytochrome bc 1 , and cytochrome c oxidase (CcO). Cytochrome c (Cc) is a small heme protein with a molecular weight of 12 500 Da that transports electrons from cytochrome bc 1 to CcO. It is a very positively charged protein, and is known to bind to both cytochrome bc 1 and CcO by means of electrostatic interactions. Extensive chemical modification studies have demonstrated that the binding domain on Cc for both proteins involves lysines immediately surrounding the heme crevice, and Cc functions as a mobile shuttle during electron transport. The reactions of Cc with its redox partners are too fast to resolve by conventional techniques such as stopped‐flow spectroscopy. A new method to study biological electron transfer has been introduced that utilizes a photoactive tris(bipyridine)ruthenium complex, Ru(II), which is covalently attached to a protein such as Cc. Photoexcitation of Ru(II) to the metal‐to‐ligand charge‐transfer state, Ru(II*), a strong reductant, leads to rapid electron transfer to the ferric heme group in Cc. Subsequent electron transfer from photoreduced heme c to redox center(s) in another protein can be measured on a timescale as short as 50 ns. This technique has been used to measure intracomplex electron transfer between Cc and its physiological partners, CcO, cytochrome bc 1 , cytochrome c peroxidase (CcP), and cytochrome b 5 . The ruthenium technique was used to characterize sequential electron transfer in cytochrome bc 1 from ubiquinol to the Rieske iron–sulfur protein (2Fe2S), cytochrome c 1 , and finally to Cc. Cytochrome oxidase contains four redox centers, Cu A , heme a, heme a 3 , and Cu B . A ruthenium Cc derivative was used to demonstrate that the initial site of electron entry into CcO is Cu A , followed by electron transfer to heme a, and then heme a 3 .

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Three Molecules of Ubiquinone Bind Specifically to Mitochondrial Cytochrome bc 1Complex

Cited by 61 publications

References 30 publications

Disulfide bond formation involves a quinhydrone-type charge–transfer complex

Disulfide bond formation involves a quinhydrone-type charge–transfer complex

Effect of Mutations of Arginine 94 on Proton Pumping, Electron Transfer, and Superoxide Anion Generation in Cytochrome b of the bc1 Complex from Rhodobacter sphaeroides

Iron: Heme Proteins & Electron Transport

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