Electron-proton interactions in terminal oxidases

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One key role of many cellular membranes is to hold a transmembrane electrochemical ion gradient that stores free energy, which is used, for example, to generate ATP or to drive transmembrane transport processes. In mitochondria and many bacteria, the gradient is maintained by proton-transport proteins that are part of the respiratory (electron-transport) chain. Even though our understanding of the structure and function of these proteins has increased significantly, very little is known about the specific role of functional protein-membrane and membrane-mediated proteinprotein interactions. Here, we have investigated the effect of membrane incorporation on proton-transfer reactions within the membrane-bound proton pump cytochrome c oxidase. The results show that the membrane acts to accelerate proton transfer into the enzyme's catalytic site and indicate that the intramolecular proton pathway is wired via specific amino acid residues to the twodimensional space defined by the membrane surface. We conclude that the membrane not only acts as a passive barrier insulating the interior of the cell from the exterior solution, but also as a component of the energy-conversion machinery.cytochrome aa3 | electron transfer | energy transduction | membrane protein | respiration

Section: Discussionmentioning

confidence: 91%

Functional interactions between membrane-bound transporters and membranes

Öjemyr

Lee

Gennis

et al. 2010

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“…However, it should be noted that we propose this role for the pathway only during formation of the P r intermediate (where the charge movement also shuts off the proton transfer capability of the pathway). The pathway is used for proton transfer during reduction of the binuclear center and possibly also immediately after oxidation of the fully reduced enzyme (36,37).…”

Section: Discussionmentioning

confidence: 99%

On the role of the K-proton transfer pathway in cytochrome c oxidase

Brändén

Sigurdson

Namslauer

et al. 2001

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141

123

Cytochrome c oxidase is a membrane-bound enzyme that catalyzes the four-electron reduction of oxygen to water. This highly exergonic reaction drives proton pumping across the membrane. One of the key questions associated with the function of cytochrome c oxidase is how the transfer of electrons and protons is coupled and how proton transfer is controlled by the enzyme. In this study we focus on the function of one of the proton transfer pathways of the R. sphaeroides enzyme, the so-called K-proton transfer pathway (containing a highly conserved Lys(I-362) residue), leading from the protein surface to the catalytic site. We have investigated the kinetics of the reaction of the reduced enzyme with oxygen in mutants of the enzyme in which a residue [Ser(I-299)] near the entry point of the pathway was modified with the use of sitedirected mutagenesis. The results show that during the initial steps of oxygen reduction, electron transfer to the catalytic site (to form the ''peroxy'' state, P r) requires charge compensation through the proton pathway, but no proton uptake from the bulk solution. The charge compensation is proposed to involve a movement of the K(I-362) side chain toward the binuclear center. Thus, in contrast to what has been assumed previously, the results indicate that the K-pathway is used during oxygen reduction and that K(I-362) is charged at pH Ϸ 7.5. The movement of the Lys is proposed to regulate proton transfer by ''shutting off'' the protonic connectivity through the K-pathway after initiation of the O 2 reduction chemistry. This ''shutoff'' prevents a short-circuit of the protonpumping machinery of the enzyme during the subsequent reaction steps.flow-flash ͉ proton pumping ͉ cytochrome aa3 ͉ flash photolysis ͉ gating ͉ R. sphaeroides

“…Furthermore, internal electron transfer between the hemes and proton transfer, linked to redox changes at heme a 3 (29), were unaffected by the structural substitution (SI Text). The multiple turnover activity of the Gly171Asp CytcO was measured at pH 6.5 and found to be 34 Ϯ 11% of that of the wild-type CytcO.…”

mentioning

confidence: 99%

A mitochondrial DNA mutation linked to colon cancer results in proton leaks in cytochrome c oxidase

Namslauer

Brzezinski

2009

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An increasing number of cancer types have been found to be linked to specific mutations in the mitochondrial DNA, which result in specific structural changes of the respiratory enzyme complexes. In this study, we have investigated the effect of 2 such mutations identified in colon cancer patients, leading to the amino acid substitutions Ser458Pro and Gly125Asp in subunit I of cytochrome c oxidase (complex IV) [Greaves et al. (2006) Proc Natl Acad Sci USA 103:714 -719]. We introduced these mutations in Rhodobacter sphaeroides, which carries an oxidase that serves as a model of the mitochondrial counterpart. The lack of expression of the former variant indicates that the amino acid substitution results in severely altered overall structure of the enzyme. The latter mutation (Gly171Asp in the bacterial oxidase) resulted in a structurally intact enzyme, but with reduced activity (approximately 30%), mainly due to slowed reduction of the redox site heme a. Furthermore, even though the Gly171Asp CytcO pumps protons, an intrinsic proton leak was identified, which would lead to a decreased overall energy-conversion efficiency of the respiratory chain, and would also perturb transport processes such as protein, ion, and metabolite trafficking. Furthermore, the specific leak may act to alter the balance between the electrical and chemical components of the proton electrochemical gradient. cytochrome aa3 ͉ electrochemical ͉ electron transfer ͉ pump ͉ respiratory oxidase