scholarly journals Formation and reduction of a ‘peroxy’ intermediate of cytochrome c oxidase by hydrogen peroxide

1984 ◽  
Vol 217 (3) ◽  
pp. 715-719 ◽  
Author(s):  
J M Wrigglesworth
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Electron Flow ◽  
Soret Band ◽  
Stable Complex ◽  
Direct Reaction ◽  
Alpha Band ◽  
Cyanide Complex ◽  
Absorption Intensity ◽  
Flow Through

In the presence of micromolar concentrations of H2O2, ferric cytochrome c oxidase forms a stable complex characterized by an increased absorption intensity at 606-607 nm with a weaker absorption band in the 560-580 nm region. Higher (millimolar) concentrations of H2O2 result in an enzyme exhibiting a Soret band at 427 nm and an alpha-band of increased intensity in the 589-610 nm region. Addition of H2O2 to ferric cytochrome c oxidase in the presence of cyanide results in absorbance increases at 444nm and 605nm. These changes are not seen if H2O2 is added to the cyanide complex of the ferric enzyme. The results support the idea that direct reaction of H2O2 with ferric cytochrome a 3 produces a ‘peroxy’ intermediate that is susceptible to further reduction by H2O2 at higher peroxide concentrations. Electron flow through cytochrome a is not involved, and the final product of the reaction is the so-called ‘pulsed’ or ‘oxygenated’ ferric form of the enzyme.

Sulphide oxidation and oxidative phosphorylation in the mitochondria of the lugworm

1997 ◽  
Vol 200 (1) ◽  
pp. 83-92 ◽  
Author(s):  
S Vökel ◽  
M K Grieshaber
Keyword(s):  
Oxygen Consumption ◽  
Cytochrome C ◽  
Oxidative Phosphorylation ◽  
Cytochrome C Oxidase ◽  
Electron Flow ◽  
Respiratory Control ◽  
Sulphide Oxidation ◽  
Atp Production ◽  
Sulphide Concentration ◽  
Flow Through

Oxygen consumption, ATP production and cytochrome c oxidase activity of isolated mitochondria from body-wall tissue of Arenicola marina were measured as a function of sulphide concentration, and the effect of inhibitors of the respiratory complexes on these processes was determined. Concentrations of sulphide between 6 and 9 µmol l-1 induced oxygen consumption with a respiratory control ratio of 1.7. Production of ATP was stimulated by the addition of sulphide, reaching a maximal value of 67 nmol min-1 mg-1 protein at a sulphide concentration of 8 µmol l-1. Under these conditions, 1 mole of ATP was formed per mole of sulphide consumed. Higher concentrations of sulphide led to a decrease in ATP production until complete inhibition occurred at approximately 50 µmol l-1. The production of ATP with malate and succinate was stimulated by approximately 15 % in the presence of 4 µmol l-1 sulphide, but decreased at sulphide concentrations higher than 15­20 µmol l-1. Cytochrome c oxidase was also inhibited by sulphide, showing half-maximal inhibition at 1.5 µmol l-1 sulphide. Sulphide-induced ATP production was inhibited by antimycin, cyanide and oligomycin but not by rotenone or salicylhydroxamic acid. The present data indicate that sulphide oxidation is coupled to oxidative phosphorylation solely by electron flow through cytochrome c oxidase, whereas the alternative oxidase does not serve as a coupling site. At sulphide concentrations higher than 20 µmol l-1, oxidation of sulphide serves mainly as a detoxification process rather than as a source of energy.

scholarly journals Hydrazine and hydroxylamine as probes for O2-reduction site of mitochondrial cytochrome c oxidase

1993 ◽  
Vol 292 (2) ◽  
pp. 519-524 ◽  
Author(s):  
T Kubota ◽  
S Yoshikawa
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Soret Band ◽  
Blue Shift ◽  
Alpha Band ◽  
Spectral Change ◽  
Spectral Changes ◽  
O2 Reduction ◽  
Highly Hydrophobic ◽  
Reduced State

Reactions of hydrazine and hydroxylamine with bovine heart cytochrome c oxidase in the fully reduced state were investigated under anaerobic conditions following the visible-Soret spectral change. Hydrazine gave a sharp band at 575 nm with 20% decrease in the alpha band at 603 nm, and hydroxylamine induced a 2 nm blue-shift for the alpha band without any clear splitting. The Soret band at 443 nm was decreased significantly in intensity, with the concomitant appearance of a shoulder with hydrazine or a peak with hydroxylamine, both near 430 nm. The dependence on pH of the affinity of these reagents for the enzyme indicates that only the deprotonated forms of these reagents bind to the enzyme, suggesting a highly hydrophobic environment of the haem ligand-biding site. These spectral changes were largely removed by addition of cyanide or CO. However, detailed analysis of these spectral changes indicates that hydrazine perturbs the shape of the spectral change induced by cyanide and hydroxylamine perturbs that induced by CO. These results suggest that these aldehyde reagents bind to haem a3 iron as well as to a second site which is most likely to be the formyl group on the haem periphery, and that these two sites bind these reagents anti-cooperatively with each other.

scholarly journals 1P-119 Soret band absorption spectra of cytochrome c oxidase in the crystalline state(The 46th Annual Meeting of the Biophysical Society of Japan)

2008 ◽  
Vol 48 (supplement) ◽  
pp. S39
Author(s):  
Masao MOCHIZUKI ◽  
Kazumasa MURAMOTO ◽  
Kyoko SHINZAWA-ITOH ◽  
Tomitake TSUKIHARA ◽  
Shinya YOSHIKAWA
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Annual Meeting ◽  
Absorption Spectra ◽  
Crystalline State ◽  
Soret Band ◽  
Biophysical Society ◽  
Band Absorption

scholarly journals The mechanism of transmembrane ΔμH+ generation in mitochondria by cytochrome c oxidase

1979 ◽  
Vol 182 (1) ◽  
pp. 133-147 ◽  
Author(s):  
M Lorusso ◽  
F Capuano ◽  
D Boffoli ◽  
R Stefanelli ◽  
S Papa
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Respiratory Chain ◽  
Electron Flow ◽  
Aerobic Oxidation ◽  
Outer Side ◽  
Rat Liver Mitochondria ◽  
Proton Release ◽  
Ferrocytochrome C ◽  
Proton Production

In rat liver mitochondria treated with rotenone, N-ethylmaleimide or oligomycin the expected alkalinization caused by proton consumption for aerobic oxidation of ferrocyanide was delayed with respect to ferrocyanide oxidation, unless carbonyl cyanide p-trifluoromethoxyphenylhydrazone was present. 2. When valinomycin or valinomycin plus antimycin were also present, ferricyanide, produced by oxidation of ferrocyanide, was re-reduced by hydrogenated endogenous reductants. Under these circumstances the expected net proton consumption caused by ferrocyanide oxidation was preceded by transient acidification. It is shown that re-reduction of formed ferricyanide and proton release derive from rotenone- and antimycin-resistant oxidation of endogenous reductants through the proton-translocating segments of the respiratory chain on the substrate side of cytochrome c. The number of protons released per electron flowing to ferricyanide varied, depending on the experimental conditions, from 3.6 to 1.5. 3. The antimycin-insensitive re-reduction of ferricyanide and proton release from mitochondria were strongly depressed by 2-n-heptyl-4-hydroxyquinoline N-oxide. This shows that the ferricyanide formed accepts electrons passing through the protonmotive segments of the respiratory chain at the level of cytochrome c and/or redox components of the cytochrome b-c1 complex situated on the oxygen side of the antimycin-inhibition site. Dibromothymoquinone depressed and duroquinol enhanced, in the presence of antimycin, the proton-release process induced by ferrocyanide respiration. Both quinones enhanced the rate of scalar proton production associated with ferrocyanide respiration, but lowered the number of protons released per electron flowing to the ferricyanide formed. 4. Net proton consumption caused by aerobic oxidation of exogenous ferrocytochrome c by antimycin-supplemented bovine heart mitochondria was preceded by scalar proton release, which was included in the stoicheiometry of 1 proton consumed per mol of ferrocytochrome c oxidized. This scalar proton production was associated with transition of cytochrome c from the reduced to the oxidized form and not to electron flow along cytochrome c oxidase. 5. It is concluded that cytochrome c oxidase only mediates vectorial electron flow from cytochrome c at the outer side to protons that enter the oxidase from the matrix side of the membrane. In addition to this consumption of protons the oxidase does not mediate vectorial proton translocation.

scholarly journals The mechanism of proton translocation by the cytochrome system of mitochondria. Characterization of proton-transfer reactions associated with oxidoreductions of terminal respiratory carriers

1983 ◽  
Vol 216 (2) ◽  
pp. 259-272 ◽  
Author(s):  
S Papa ◽  
F Guerrieri ◽  
G Izzo
Keyword(s):  
Cytochrome C ◽  
Proton Transfer ◽  
Cytochrome C Oxidase ◽  
Electron Flow ◽  
Aerobic Oxidation ◽  
Proton Pumping ◽  
Rat Liver Mitochondria ◽  
Complex Scalar ◽  
Total Electron ◽  
Ionizable Groups

A direct kinetic analysis is presented of rapid proton-releasing reactions at the outer or C-side of the membrane, in ox heart and rat liver mitochondria, associated with aerobic oxidation of reduced terminal respiratory carriers in the presence of antimycin. Valinomycin plus K+ enhances the rate of cytochrome c oxidation and the rate and extent of H+ release. In the presence of valinomycin the leads to H+/e- ratio, computed on the basis of total electron flow from respiratory carriers to oxygen, varies with pH, remaining always lower than 1, and is unaffected by N-ethylmaleimide. 2-Heptyl-4-hydroxyquinoline N-oxide and 5-(n-undecyl)-6-hydroxy-4,7-dioxobenzothiazole, at concentrations which inhibit in the presence of antimycin the oxygen-induced reduction of b cytochromes, cause also a marked depression of the H+ release associated with aerobic oxidation of terminal respiratory carriers. Aerobic oxidation of the cytochrome system in mitochondria and of isolated b-c1 complex and cytochrome c oxidase results in scalar proton release from ionizable groups (redox Bohr effects). In mitochondria and submitochondrial particles, about 70% of the oxidoreductions of the components of the cytochrome system are linked to scalar proton transfer by ionizable groups. In isolated b-c1 complex scalar proton transfer, resulting from redox Bohr effect, amounts to 0.9H+ per Fe-S protein (190 muT). In isolated cytochrome c oxidase, Bohr protons amount to 0.8 per haem a + a3. The results presented indicate that the H+ release from mitochondria during oxidation of terminal respiratory carriers derives from residual antimycin-insensitive electron flow in the quinone-cytochrome c span and from redox Bohr effects in the b-c1 complex and cytochrome c oxidase. There is no sign of proton pumping by cytochrome oxidase during its transition from the reduced to the active ‘pulsed’ state and the first one or two turnovers.

Dibucaine interacts differently with membrane and protein in cytochrome c oxidase systems

1993 ◽  
Vol 71 (1-2) ◽  
pp. 14-21
Author(s):  
Jia He ◽  
Peter Nicholls
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Proton Translocation ◽  
Tight Binding ◽  
Respiratory Control ◽  
Soret Band ◽  
Red Shift ◽  
Ph Gradients ◽  
Low Concentrations ◽  
Low Ionic Strength

Dibucaine acts as a weak protonophore in cytochrome c oxidase proteoliposomes. At low concentrations in the presence of permeant anions, it stimulates turnover and collapses enzyme-generated pH gradients. At higher concentrations, dibucaine inhibits activity of cytochrome c oxidase in proteoliposomes and the isolated enzyme. It also induces a red shift in the resting spectrum, indicating a change at the binuclear centre. This spectroscopic effect is kinetically biphasic. Dibucaine inhibits steady-state oxidase activity, but not the rate of the red shift in the cytochrome a3 Soret band during turnover. It reacts faster with the partially reduced state than with resting enzyme. The inhibition is kinetically biphasic with a noncompetitive Ki ≈ 0.5 mM. Excess dibucaine effects a maximal turnover decline of 80%. At low ionic strength only the total Vmax is affected; tight binding of cytochrome c and turnover at the "tight" site are unaffected. Dibucaine may bind to an anionic site in a hydrophobic pocket, modifying electron transfer from cytochrome a and CuA to cytochrome a3 - CuB and the oxidized spectrum of the latter centre. Stimulation of turnover in cytochrome c oxidase in proteoliposomes is due to a separate membrane-dependent proton translocation catalysed by dibucaine in the presence of permeant anions.Key words: dibucaine, cytochrome c oxidase, proteoliposomes, respiratory control, inhibition.

Structural and mechanistic insights into the electron flow through protein for cytochrome c-tethering copper nitrite reductase

2013 ◽  
Vol 154 (1) ◽  
pp. 51-60 ◽  
Author(s):  
Aiko Tsuda ◽  
Ryosuke Ishikawa ◽  
Hiroyasu Koteishi ◽  
Kosuke Tange ◽  
Yohta Fukuda ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Nitrite Reductase ◽  
Electron Flow ◽  
Flow Through
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