Top PDF Electron transfer pathways in cytochrome c oxidase

Electron transfer pathways in cytochrome c oxidase

Electron transfer pathways in cytochrome c oxidase

heme a 3 cannot be ruled out [14] . In spite of all the research done in this area, the exact path for electron transfer remains unknown. The most common method to study this kind of process consists of mutational experiments. However, even though this technique, in principle, points out the importance of some residues within a protein its results can also be misleading since mutations can produce changes in the original structure that may mask the real cause for the loss/gain of activity or, in this particular case, electron transfer. Alternatively, computational tools can provide a quick and cheap alternative in mapping electron transfer pathways with the key advantage of not introducing signi ficant perturbations on the system [25 –27] . Much effort has also been dedicated to the assessment of the time evolution of an electron transfer process by diverse approaches as described in the following work and references within [28,29] . In the present work, however, we will explore the electron transfer pathway, in cytochrome c oxidase, by means of a new computation technique [30] that has already shown to give good results [31] . This method, QM/MM e-pathway, consists basically in activating –deactivating different regions in the quantum region, by means of a QM/MM scheme, which allows us to follow the progress of an electron and so describe its path between a donor and acceptor. The theoretical analysis indicates a mechanism for the electron transfer that follows the sequence Cu A →heme a→heme a 3 , which is signi ficantly affected by electrostatic
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Deuterium isotope effect of proton pumping in cytochrome c oxidase

Deuterium isotope effect of proton pumping in cytochrome c oxidase

The X-ray crystal structure of the ED286 mutant CytcO indicates that the shorter side chain of D286 compared to that of E286 leaves a pocket with enough space for an additional water molecule in the mutant enzyme (see Fig. 3 ). Due to the limited resolution of the mutant CytcO structure (3.0/3.5 Å), water molecules were not included in the refinement. How- ever, in the difference Fourier electron density map (shown as a green mesh in Fig. 3 ) there is a peak between the D286 side chain and the water molecule hydrogen-bonded to E286 in the wild-type structure. This electron density could corres- pond to a new water molecule, or alternatively, the reallo- cation of one already present in wild-type CytcO. We also see that the hydrogen bond between residue 286 and the carbonyl oxygen of M107, observed in the wild-type CytcO, is not present in the ED286 mutant CytcO structure as the distance is too large.
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Variable proton-pumping stoichiometry in structural variants of cytochrome c oxidase

Variable proton-pumping stoichiometry in structural variants of cytochrome c oxidase

In general, the proton-pumping stoichiometry is presumably determined by the relative proton-transfer rates from Glu286 to the catalytic site and the pump site, respectively. These rates are presumably dependent on the local environment of the Glu286, not only because changes in the environment would alter the proton connectivity, but more importantly because the Glu286 presumably has to isomerize during the transfer. Such an isomerization would be sensitive to any changes in the hydrogen-bonding pattern of the Glu. A structural isomerization of the Glu286 side chain, linked to proton transfer was originally proposed by Iwata et al. [10] based on an inspection of the P. denitri ficans CytcO crystal structure (because of the lack of protonic connectivity between the Glu and the catalytic site or the pump site). This proposal is supported by results from theoretical calculations [50,151 –155] , which suggested that Glu286 conforma- tional isomerization may be involved in proton gating. Results from more recent computational studies suggested that Glu286 may function as a proton valve, which can adopt distinct conformations thereby controlling the proton connectivity to solution, the catalytic site and the pump site [156,157] . However, as pointed out by Warshel and colleagues [54] , an isomerization of a side chain by itself cannot establish a gate (because of microscopic reversibility). Nevertheless, changes in the equilibrium constant between the Glu286 conformers are expected to result in proton leaks that would decrease the proton- pumping stoichiometry [157] .
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Decoding Cytochrome C Oxidase Biogenesis: New Insights Into Copper Trafficking

Decoding Cytochrome C Oxidase Biogenesis: New Insights Into Copper Trafficking

supplement to produce an active cbb 3–Cox. Complementation of this mutant using wild-type genomic libraries unveiled a novel gene (ccoA) required for cbb 3 –Cox biogenesis in R. capsulatus. In the absence of CcoA, cellular content of Cu decreases, and cbb 3 –Cox assembly and activity becomes defective. CcoA shows pronounced homology to Major Facilitator Superfamily (MFS) type transporter proteins. Members of this family are known to transport small solutes or drugs, but so far, no MFS protein was implicated in cbb 3 –Cox biogenesis. In order to dissect the mechanism of Cu acquisition in the absence of CcoA, we isolated ΔccoA mutants that were cbb 3 –Cox defective after addition of Cu. Characterization of these mutants by genetic complementations revealed mutations in cytochrome c maturation (CCM) genes. These mutants were able to grow photosynthetically on the contrary to the usual phenotype of CCM genes deletion mutants. Here we show that these mutations are not directly involved in the Cu trafficking to CcoN but involved in the production of membrane bound cytochrome c subunits of cbb 3 –Cox. Although this study provides additional information about CCM system in R. capsulatus, the additional pathways of Cu acquisition to cbb 3 –Cox in the presence of exogenous Cu still remains to be identified. In the future, determination of ccoA bypass mutations will provide novel insights on the maturation and assembly of membrane-integral metalloproteins, and on hitherto unknown function(s) of MFS type transporters in bacterial Cu acquisition.
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Coordination of cytochrome c oxidase gene expression in the remodelling of skeletal muscle

Coordination of cytochrome c oxidase gene expression in the remodelling of skeletal muscle

In any multistep process, some steps will be more important in determining the overall rate. In the case of metabolic pathways, quantitative control analyses can assign a step a control coefficient, or, in more qualitative terms, some enzymes are considered to be ‘rate limiting’ or ‘rate determining’. These steps are often the slowest, and are typically subject to other forms of metabolic regulation. It is reasonable to speculate that some COX subunits may be more limiting than others and, if so, may have a greater role in determining changes in COX activity. By analogy with metabolic pathways, rate-determining subunits might be those where changes in mRNA level best paralleled COX activity. Many studies have used molecular genetic interventions to impose a reduction in the synthesis of individual subunits to assess the effects on COX biosynthesis. For example, dramatic reductions in COX arise when COX5A is disrupted using morpholinos in zebrafish (Baden et al., 2007) and RNAi in Caenorhabditis elegans (Suthammarak et al., 2009). Likewise, loss of COX activity is seen when COX6A synthesis is disrupted in a mouse knockout (Radford et al., 2002), Drosophila mutants (Liu et al., 2007) and human cell line knockouts (Fornuskova et al., 2010). Reductions in COX4 lead to a decrease in COX content in mammalian lines (Li et al., 2006; Fornuskova et al., 2010) and C. elegans (Suthammarak et al., 2009). In some cases, this intervention leads to the accumulation of assembly intermediates and enzymatic abnormalities (Fornuskova et al., 2010). Such studies are used to assess the importance of subunits in assembly or function but they are not intended to explore constraints on the rate of synthesis during adaptive remodelling. The question of whether any of these genes exert disproportionate control over COX synthesis has not been addressed experimentally. This study gives an indication of the sensitivity of COX synthesis to variation in subunit expression. Thus, we consider these data to
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A study of cytochrome C oxidase subunit 1 polymorphism in Malay population

A study of cytochrome C oxidase subunit 1 polymorphism in Malay population

Cytochrome c oxidase subunit 1 DNA size is around 1548 bp which has 70.2% total T content (Shaikevich and Zakharov., 1993). In addition, Cytochrome c oxidase protein (EC 1.9.3.1) is the last enzyme complex of respiratory electron transport chain which is located in the mitochondrial membrane (Valnotet al., 2000).This enzyme is acting as a dioxygen activator in aerobic life by transferring the electron from the reduced cytochrome c to the oxygen (Valnotet al., 2000). There are three different subunits of cytochrome c oxidase, COI, COII and COIII. Between the three mitochondrial genes which are coding the cytochrome c oxidase subunits, COI is the largest one and the most conserved between them (Beard et al., 1993). Based on a survey about COI gene on some of the search engines such as GeneBank BLAST and BOLD, it can be concluded that the COI gene reliably identifies species where the reference sequence data is present (Dawnayet al., 2006).
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Electron Transfer of Co-immobilized Cytochrome c and Horseradish Peroxidase in Chitosan-Graphene Oxide Modified Electrode

Electron Transfer of Co-immobilized Cytochrome c and Horseradish Peroxidase in Chitosan-Graphene Oxide Modified Electrode

A novel bi-protein (Cyt c and HRP) electrode was fabricated based on a nanocomposite of GO and CHIT through self-assembly method. An electron transfer protein of Cyt c was used as a test system for direct electron transfer (DET) of redox proteins and for communication in the HRP stacks by co-immobilizing it with HRP. The experimental results indicated that the biocompatible matrix supplied a necessary pathway for the co-immobilized proteins to achieve DET and promoted the electron transfer between the coexist proteins and underlying electrode. Furthermore, the co- immobilized electrode exhibited a larger electron transfer rate constant compared to the electrode modified with single protein. The bi-protein electrode exhibited fast electron transfer and good electro- catalytic activity toward the reduction of H 2 O 2 and O 2 . The resulted biosensor exhibited a sensitive and
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Biogenesis of cbb3-type cytochrome c oxidase in Rhodobacter capsulatus

Biogenesis of cbb3-type cytochrome c oxidase in Rhodobacter capsulatus

domain of CcmI recognizes and binds tightly the most C-terminal helix of apocytochrome c 2 in the absence of heme b. The folding pro- cess of cytochrome c (and cytochrome c 2 ) [124] indicates that their most C-terminal helix interacts with their most N-terminal heme binding helix to form a stable folding intermediate that can trap heme b non-covalently [125] . Altogether, these observations led to the proposal that at least some c-type apocytochromes are first recog- nized via their C-terminal helices by the periplasmic CcmI-2 domain of the CcmFHI core complex ( Fig. 4 , middle), and then released from this domain upon transfer of heme b from CcmF to apocytochrome, and subsequently, the thioether bonds are formed. Once the thioether bonds are formed, cytochrome c folds into its final structure.
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Structure/function of oxygen regulated isoforms in cytochrome c oxidase

Structure/function of oxygen regulated isoforms in cytochrome c oxidase

The means by which electron transfer is coupled to proton translocation in cytochrome c oxidase is unclear, but it has been suggested by many laboratories that the enzyme has distinct redox-sensitive conformational states at its heme–copper binuclear center and that these states couple electron and proton movement (Chan and Li, 1990; Woodruff, 1993; Wikstrom et al. 1994; Iwata et al. 1995; Wittung and Malmstrom, 1996). There is ample evidence that both the oxidized and reduced forms of the enzyme may exist in multiple conformational states and that these are sensitive to pH. For the oxidized enzyme, these are called ‘fast’ and ‘slow’ states (Moody, 1996). They can be distinguished by their spectral signatures. The ‘fast’ form is considered to be a fully oxidized active enzyme, while the ‘slow’ form is considered to be an artifact that arises during the purification and/or storage of the enzyme. The ‘fast’ form converts to the ‘slow’ form at low pH, and the ‘slow’ form may be converted back to the ‘fast’ form by a cycle of reduction and reoxidation. Neither the nature of these pH-induced changes nor their relevance to catalysis is known. For the reduced enzyme, conformers are observable by using Fourier transform infrared Table 1. Correspondence between yeast and mammalian
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Intracytoplasmic Copper Homeostasis Controls Cytochrome c Oxidase Production

Intracytoplasmic Copper Homeostasis Controls Cytochrome c Oxidase Production

Molecular genetic techniques. Standard molecular genetic tech- niques were performed as described previously (42). The gene transfer agent (GTA) (43) of R. capsulatus was used to construct chromosomal knockout alleles of desired genes as described earlier (44). Genomic DNA was extracted from 10-ml cultures of the wild type (MT1131), ⌬ ccoA mutant (SE8), and its Cox ⫹ revertants (SE8Ri with i ⫽ 1 to 6) that were grown in MPYE medium by using the DNeasy Blood & Tissue kit (Qiagen Inc.). The quality and quantity of the isolated genomic DNA were ana- lyzed by using a Nanodrop spectrophotometer and by agarose gel electro- phoresis prior to library preparations for NGS.
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Biogenesis and assembly of eukaryotic cytochrome c oxidase catalytic core

Biogenesis and assembly of eukaryotic cytochrome c oxidase catalytic core

Despite continuous efforts to fully understand the biogenesis and assembly of mitochondrial COX catalytic core subunits in several laboratories and the recent advances made in our understanding of the mechanisms involved, sorting out its complexity and how COX biogenesis is regulated remains a remarkable challenge. Whereas the number of COX ancillary factors identi fied continues increasing; the speci fic functions of most of them are only partially character- ized. Speci fic functions such as the protein-assisted assembly of Cox3, the dehydrogenase activity required for the last step of heme a biosyn- thesis, or the Cox1 heme insertion chaperone, among many others, wait to have a protein performer identi fied and/or assigned. Regulatory pathways coordinating COX core subunit synthesis, cofactor availability, maturation and assembly also remain to be fully characterized. A final challenge will involve the identi fication of COX assembly factors and regulatory pathways conserved from yeast to human and those that evolved to adapt to the tissue-speci fic requirements of multicellular organisms. This has a great biomedical relevance since lesions affect- ing the expression and assembly of COX catalytic core subunits result in severe human mitochondrial encephalomyopathies.
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Direct Electron Transfer of Cytochrome C on Cadmium Oxide Nanoparticles Modified Carbon Paste Electrode

Direct Electron Transfer of Cytochrome C on Cadmium Oxide Nanoparticles Modified Carbon Paste Electrode

Direct electron transfer reaction of cytochrome c has been obtained at a cadmium oxide nanoparticles modified electrode. These nanoparticles helped cytochrome c to have a favored orientation and reduce the effective electron transfer distance. Direct electron transfer of the cytochrome c immobilized in modified carbon paste electrode was easily achieved. A pair of well- defined and quasi-reversible redox peaks appeared at the modified electrode with the cathodic and anodic peak potential of −0.305 V and -0.244 V (vs. SCE) respectively, indicating that direct electrochemistry of cytochrome c had occurred. As a result, this novel biosensor showed high sensitivity, a wide linear range, low detection limit and good stability for electrochemical detection of H 2 O 2 . This work may represent a facile and promising approach for the fabrication of various
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Alternative electron transfer pathways in iron-metabolising bacteria

Alternative electron transfer pathways in iron-metabolising bacteria

Unlike in aerobic respiration, the ETC does not include complexes III and IV, as oxygen is not the terminal electron acceptor. Contrarily, electrons are transferred via the quinone pool to an inner membrane tetraheme c-type cytochrome (CymA), the first protein of Shewanella’s Mtr pathway. From CymA, electrons are transferred to the periplasm, where a decaheme c-type cytochrome (MtrA) is facing the periplasm embedded in an outer membrane porin (MtrB). MtrA transfers the electrons to MtrC, an outer membrane decaheme c-type cytochrome located at the other side of MtrB and exposed to the external media. Using the MtrB porin, MtrA and MtrC are connected so electrons can cross the outer membrane (Marritt et al., 2012; McMillan et al., 2012; Shi, Rosso, Clarke, et al., 2012; Beckwith et al., 2015). From the location of the hemes in the resolved structure of MtrC (Figure 1-4) it is thought that the hemes in MtrA and MtrC form an electric wire that facilitate electron transfer across the outer membrane (Edwards et al., 2015).
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Expanding the view of proton pumping in cytochrome c oxidase through computer simulation

Expanding the view of proton pumping in cytochrome c oxidase through computer simulation

One of the keys to conducting a meaningful MS-EVB simulation is the algorithm that dynamically selects diabatic states. Initially, the ex- cess proton is considered as a classical hydronium (H 3 O + ), which is the most favorable diabatic state (it possesses the maximum weight in the ground-state eigenvector). Then, a breadth- first search (BFS) is adopted on solvation shells, one at a time (for details please refer to Ref. [ 84 ], sec. 2.2). More importantly, states in an outer shell are searched by their hydrogen bond networks, which are the essential pathways for proton transport [80] . At each MD step, the state- search algorithm is conducted, which tracks the movement of the excess proton. Therefore, the critical area containing those atoms in- volved in the bond breaking/forming process, referred to as the “EVB-complex”, is dynamically created for the system during the MD simulation. Thus, the candidate diabatic states are intelligently selected based on the hydrogen bond topology, thus reducing the computational cost while still ensuring the key proton transfer path- ways are collected.
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A Heme-Sensing Mechanism in the Translational Regulation of Mitochondrial Cytochrome c Oxidase Biogenesis

A Heme-Sensing Mechanism in the Translational Regulation of Mitochondrial Cytochrome c Oxidase Biogenesis

*Correspondence: abarrientos@med.miami.edu http://dx.doi.org/10.1016/j.cmet.2012.10.018 SUMMARY Heme plays fundamental roles as cofactor and signaling molecule in multiple pathways devoted to oxygen sensing and utilization in aerobic organisms. For cellular respiration, heme serves as a prosthetic group in electron transfer proteins and redox enzymes. Here we report that in the yeast Saccharo- myces cerevisiae, a heme-sensing mechanism trans- lationally controls the biogenesis of cytochrome c oxidase (COX), the terminal mitochondrial respira- tory chain enzyme. We show that Mss51, a COX1 mRNA-specific translational activator and Cox1 chaperone, which coordinates Cox1 synthesis in mitoribosomes with its assembly in COX, is a heme-binding protein. Mss51 contains two heme regulatory motifs or Cys-Pro-X domains located in its N terminus. Using a combination of in vitro and in vivo approaches, we have demonstrated that these motifs are important for heme binding and effi- cient performance of Mss51 functions. We conclude that heme sensing by Mss51 regulates COX biogen- esis and aerobic energy production.
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Mutational Analysis of the Saccharomyces cerevisiae Cytochrome c Oxidase Assembly Protein Cox11p

Mutational Analysis of the Saccharomyces cerevisiae Cytochrome c Oxidase Assembly Protein Cox11p

⌬ cox11 mutant strains are hydrogen peroxide hypersensitive. We previously reported that a yeast sco1 null mutant has a hydrogen peroxide-hypersensitive phenotype (30). This obser- vation, in the context of the crystal structure of hSCO1, has led to the proposal that Sco1p may have a second, previously unanticipated, function. Both Cox11p and Sco1p are copper- binding proteins of the inner mitochondrial membrane, and both are proposed to participate in similar pathways, namely, copper acquisition from Cox17p and copper donation to Cox1p and Cox2p, respectively. In light of this, we wanted to deter- mine whether cox11 mutants might display a similar hypersen- sitivity to hydrogen peroxide. We subjected the cox11 null allele to acute hydrogen peroxide exposure and analyzed the effects of this exposure on the viability of the strain. As shown in Fig. 5, hydrogen peroxide had virtually no effect on the survival of wild-type aW303 cells. In contrast, the same treatment resulted in almost complete loss of viability of aW303 ⌬ COX11 cells. This hypersensitivity is clearly the result of loss of Cox11p, as a CEN plasmid-borne copy of COX11 rescued aW303 ⌬ COX11 from the peroxide-sensitive pheno- type (Fig. 5). This result was similar to that obtained with the aW303 ⌬ SCO1 strain that we reported earlier (30). Loss of COX assembly alone, such as that resulting from loss of Cox4p (aW303 ⌬ COX4), Cox6p (aW303 ⌬ COX6), or Cox9p (aW303- ⌬ COX9), does not result in peroxide sensitivity (Fig. 5), sug- gesting that peroxide sensitivity is not merely a consequence of the loss of assembled COX. In addition, a ␳ 0 derivative of
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Myocardial insufficiency is related to reduced subunit 4 content of cytochrome c oxidase

Myocardial insufficiency is related to reduced subunit 4 content of cytochrome c oxidase

Deterioration of myocardial contractility is an obvious indicator for reduced oxygen supply. In coronary heart disease, when arteriosclerotic plaque formation reduces blood flow, the reduced coronary blood supply results in ischemia and damage to the myocardium. Myocardial respiration depends on Cytochrome c- Oxidase (CytOx) activity. It represents the rate limiting step for the func- tion of the mitochondrial respiratory chain, also known as electron transmission chain (ETC). If cardiac failure is related to the compromised cellular respiration of the heart, remains unclear. But contractility requires abundant supply of adenosine triphosphate (ATP), and this kind of “energy currency” is produced in mitochondria (see Fig. 1) where oxygen consumption for water production at Cyto- chrome c- Oxidase (E.C. 1.9.3.1.) is a rate-limiting step. Decreased expression of COX 4 results in an impaired Cytochrome c oxidase activity [4, 5]. We hypothesize sub- sequent mitochondrial dysfunction associated with the
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The cyclope Gene of Drosophila Encodes a Cytochrome c Oxidase Subunit VIc Homolog

The cyclope Gene of Drosophila Encodes a Cytochrome c Oxidase Subunit VIc Homolog

Cytochrome c oxidase is the terminal enzyme of the mitochondrial electron transfer chain. In eukaryotes, the enzyme is composed of 3 mitochondrial DNA-encoded subunits and 7–10 (in mammals) nuclear DNA- encoded subunits. This enzyme has been extensively studied in mammals and yeast but, in Drosophila, very little is known and no mutant has been described so far. Here we report the genetic and molecular characterization of mutations in cyclope (cype) and the cloning of the gene encoding a cytochrome c oxidase subunit VIc homolog. cype is an essential gene whose mutations are lethal and show pleiotropic phenotypes. The 77-amino acid peptide encoded by cype is 46% identical and 59% similar to the human subunit (75 amino acids). The transcripts are expressed maternally and throughout development in localized regions. They are found predominantly in the central nervous system of the embryo; in the central region of imaginal discs; in the germarium, follicular, and nurse cells of the ovary; and in testis. A search in the Genome Annotation Database of Drosophila revealed the absence of subunit VIIb and the presence of 9 putative nuclear cytochrome c oxidase subunits with high identity scores when compared to the 10 human subunits.
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Computational studies of electron transfer in the bacterial deca-heme cytochrome MtrF

Computational studies of electron transfer in the bacterial deca-heme cytochrome MtrF

1.3 Thesis statement The aim of this work is to provide detailed insight into the properties of OM cytochromes of S. oneidensis, specifically MtrF and MtrC, on a molecular level using computational modelling methods. These allow us to address questions that are difficult to tackle by experiment. Specifically, thermody- namic and kinetic parameters for electron transfer between the ten cofactors in MtrF are calculated, and hence for transport through the entire protein, starting from the atomistic structure of MtrF. Using the framework of nona- diabatic Marcus theory allows us to understand electron transfer in terms of driving forces, free energies of (protein and solvent) reorganization and elec- tronic couplings between heme cofactors. The results not only allow for pre- dictions in regard to overall trans-protein transport rates and hypotheses re- garding the function of different parts of the protein but also enable to draw correlations between protein structure and function and thus to deduce design principles. Fig. 1.6 illustrates the multi-scale nature of this topic: To describe electron transport through MtrF as a whole (A), ET rates are needed for pairs of adjacent hemes (B) which requires calculation of the electronic structure of individual hemes and averaging of heme properties over the thermally fl- cutuating protein conformations (C).
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Studies on the structure, function and mechanisms of the cbb3 type cytochrome c oxidase from Vibrio cholerae and the cytochrome bo3 ubiquinol oxidase from Escherichia coli

Studies on the structure, function and mechanisms of the cbb3 type cytochrome c oxidase from Vibrio cholerae and the cytochrome bo3 ubiquinol oxidase from Escherichia coli

82 4.1.3 Metal Detection and ICP Technology An inductively coupled plasma (ICP) is a type of plasma source in which the energy is supplied by electric currents which are produced by electromagnetic induction, that is, by time- varying magnetic fields. When a time-varying electric current is passed through the coil, it creates a time-varying magnetic field around it, which in turn induces azimuthal electric currents in the rarefied gas, leading to the formation of plasma. Argon is one example of a commonly- used rarefied gas. Plasma temperatures can range between 6 000 K and 10 000 K, comparable to the surface of the sun. ICP discharges are of relatively high electron density, on the order of 1015 cm −3 . As a result, ICP discharges have wide applications where high-density plasma is needed. Another benefit of ICP discharges is that they are relatively free of contamination because the electrodes are completely outside the reaction chamber. By contrast, in capacitively coupled plasma (CCP), the electrodes are often placed inside the reactor and are thus exposed to the plasma and subsequent reactive chemical species.
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