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1.
Cytochrome caa3 (cytochrome oxidase) from the thermophilic bacterium PS3 can exhibit full catalytic activity in the presence of ascorbate and TMPD or other electron donors and in the absence of added soluble c-type cytochromes. It appears to possess only a low-affinity and not a high-affinity site for the soluble cytochromes. Proteoliposomal cytochrome caa3 develops an effective membrane potential in the presence of ascorbate and TMPD or PMS, in the absence of added soluble cytochrome c. Reduction of the a3 centre is blocked in the presence of cyanide. During reductive titrations of the cyanide-inhibited enzyme, electrons initially equilibrate among three centres, the c haem, the a haem and one of the associated Cu atoms. During steady-state turnover, electrons probably enter the complex via the bound c haem; the a haem and perhaps an associated CuA atom are reduced next. It is concluded that, despite its size and hydrophobic association with the aa3 complex, the haem c-containing subunit can behave in an analogous way to that of mammalian cytochrome c, bound at the high-affinity site of the eucaryotic enzyme. 相似文献
2.
Haem O and a putative cytochrome bo in a mutant of Bacillus cereus impaired in the synthesis of haem A 总被引:1,自引:0,他引:1
I. P. Del Arenal M. L. Contreras B. B. Svlateorova P. Rangel F. Lledías J. R. Dávila J. E. Escamilla 《Archives of microbiology》1997,167(1):24-31
In a spontaneous mutant (PYM1) of Bacillus cereus impaired in the synthesis of haem A, no haem-A-containing cytochromes were detected spectroscopically. The haem A deficiency
was compensated by high levels of haem O and a CO-reactive cytochrome o in membranes; no other oxidases were detected. In contrast, the wild-type strain had considerable amounts of haem A and negligible
levels of haem O. The mutant PYM1 exhibited normal colony morphology, growth, and sporulation in nonfermentable media, whereas
on fermentable media, the mutant overproduced acid, which led to poor growth and inhibition of sporulation. External control
of the pH of the medium in fermentable media allowed close-to-normal growth and massive sporulation of the mutant. The presence
of membrane-bound cytochrome caa
3
-OII and aa
3
-II subunits in strain PYM1 was confirmed by Western blots and haem C staining (COII subunit). Western blotting also revealed
that in contrast to the wild-type – strain PYM1 contained the membrane-bound subunits caa
3
-COI and aa
3
-I, but in low amounts. The effect of several respiratory inhibitors on the respiratory system of strain PYM1 suggested that
the terminal oxidase is highly resistant to KCN and CO and that a c-type cytochrome might be involved in the electron transfer sequence to the putative cytochrome bo.
Received: 21 June 1996 / Accepted: 9 October 1996 相似文献
3.
D. Boffoli S.C. Scacco R. Vergari G. Solarino G. Santacroce S. Papa 《生物化学与生物物理学报:疾病的分子基础》1994,1226(1):73-82
Mitochondrial respiratory systems have been screened in 63 othopaedic patients of age ranging between 17 and 91 years. The results show a statistically significant definite decrease with ageing of mitochondrial repiratory activity with pyruvate plus malate, succinate and ascorbate plus TMPD. This pattern is associated with an equally significant decrease with age of the enzymatic activity of complex I, II and IV. No significant decrease with age is, on the contrary, observed in the mitochondrial content of cytochromes a+a3, and c+c1. Preliminary Western blot analysis indicates an altered polypeptide pattern in cytochrome c oxidase. This study provides evidence for a decline with age of mitochondrial respiratory activity in human skeletal muscle, affecting complex I, II and IV. 相似文献
4.
Several cytochromes c2 from the Rhodospirillaceae show a pH dependence of redox potential in the physiological pH range which can be described by equations involving an ionisation in the oxidised form (pKo) and one in the reduced form (pKr). These cytochromes fall into one of two groups according to the degree of separation of pKo and pKr. In group A, represented here by the Rhodomicrobium vannielii cytochrome c2, the separation is approx. one pH unit and the ionisation is that of a haem propionic acid. Members of this group are unique among both cytochromes c2 and mitochondrial cytochromes c in lacking the conserved residue Arg-38. We propose that the role of Arg-38 is to lower the pK of the nearby propionic acid, so that it lies out of the physiological pH range. Substitution of this residue by an uncharged amino acid leads to a raised pK for the propionic acid. In group B, represented here by Rhodopseudomonas viridis cytochrome c2, the separation between pKo and pKr is approx. 0.4 pH unit and the ionisable group is a histidine at position 39. This was established by NMR spectroscopy and confirmed by chemical modification. Only a few other members of the cytochrome c2/mitochondrial cytochrome c family have a histidine at this position and of these, both Crithidia cytochrome c-557 and yeast cytochrome c were found to have a pH-dependent redox potential similar to that of Rps. viridis cytochrome c2. Using Coulomb's law, it was found that the energy required to separate pKo and pKr could be accounted for by simple electrostatic interactions between the haem iron and the ionisable group. 相似文献
5.
Catarina M. Paquete Patrícia M. Pereira Teresa Catarino David. L. Turner Ricardo O. Louro 《BBA》2007,1767(2):178-188
Type I cytochrome c3 is a key protein in the bioenergetic metabolism of Desulfovibrio spp., mediating electron transfer between periplasmic hydrogenase and multihaem cytochromes associated with membrane bound complexes, such as type II cytochrome c3. This work presents the NMR assignment of the haem substituents in type I cytochrome c3 isolated from Desulfovibrio africanus and the thermodynamic and kinetic characterisation of type I and type II cytochromes c3 belonging to the same organism. It is shown that the redox properties of the two proteins allow electrons to be transferred between them in the physiologically relevant direction with the release of energised protons close to the membrane where they can be used by the ATP synthase. 相似文献
6.
Lígia M. Saraiva Carlos A. Salgueiro Jean LeGall Walter M. A. M. van Dongen António V. Xavier 《Journal of biological inorganic chemistry》1996,1(6):542-550
Reduction of the haems in tetrahaem cytochromes c
3 is a cooperative process, i.e., reduction of each of the haems depends on the redox states of the other haems. Furthermore,
electron transfer is coupled to proton transfer (redox-Bohr effect). Two of its haems and a strictly conserved nearby phenylalanine
residue, F20, in Desulfovibrio vulgaris (Hildenborough) cytochrome c
3 form a structural motif that is present in all cytochromes c
3 and also in cytochrome c oxidase. A putative role for this phenylalanine residue in the cooperativity of haem reduction was investigated. Therefore,
this phenylalanine was replaced, with genetic techniques, by isoleucine and tyrosine in D. vulgaris (Hildenborough) cytochrome c
3. Cyclic voltammetry studies revealed a small increase (30 mV) in one of the macroscopic redox potentials in the mutated cytochromes.
EPR showed that the main alterations occurred in the vicinity of haem I, the haem closest to residue 20 and one of the haems
responsible for positive cooperativities in electron transfer of D. vulgaris cytochrome c
3. NMR studies of F20I cytochrome c
3 demonstrated that the haem core architecture is maintained and that the more affected haem proton groups are those near the
mutation site. NMR redox titrations of this mutated protein gave evidence for only small changes in the relative redox potentials
of the haems. However, electron/electron and proton/electron cooperativity are maintained, indicating that this aromatic residue
has no essential role in these processes. Furthermore, chemical modification of the N-terminal amino group of cytochrome c
3 backbone, which is also very close to haem I, had no effect on the network of cooperativities.
Received: 25 June 1996 / Accepted: 26 August 1996 相似文献
7.
Isabel B. Coutinho David L. Turner Ming Y. Liu Jean LeGall António V. Xavier 《Journal of biological inorganic chemistry》1996,1(4):305-311
The trihaem cytochrome c
551.5, formerly known as cytochrome c
7, from the organism Desulfuromonas
acetoxidans, has been studied in the reduced state by 2D proton NMR. The haem proton resonances were assigned, and several nuclear Overhauser
enhancements (NOEs) between resonances arising from different haems were detected and assigned. The relative orientations
of the three haems were calculated by fitting both the intensities of the interhaem NOEs and the magnitudes of the ring current
shifts of the haem resonances, following the strategy previously used by the authors to reassess the X-ray structure of the
haem core in tetrahaem cytochrome c
3 from Desulfumicrobium baculatum.
It is concluded that, although the comparison of the protein sequence with those of the tetrahaem cytochromes c
3 shows that in cytochrome c
551.5 about 40% of the sequence is deleted, including the region involved in the attachment of the second of the four haems, this
does not induce any significant rearrangement of the remaining three haems other than a slight decrease in the iron-iron distance
between two of the haems, namely those corresponding to haems I and IV of cytochrome c
3.
Received: 2 February 1996 / Accepted: 26 March 1996 相似文献
8.
1. The steady-state oxidation of ferrocytochrome c by dioxygen catalyzed by cytochrome c oxidase, is inhibited non-competitively towards cytochrome c by methanethiol, ethanethiol, 1-propanethiol and 1-butanethiol with Ki values of 4.5, 91, 200 and 330 μM, respectively.2. The inhibition constant Ki of ethanethiol is found to be constant between pH 5 and 8, which suggests that only the neutral form of the thiol inhibits the enzyme.3. The absorption spectrum of oxidized cytochrome c oxidase in the Soret region shows rapid absorbance changes upon addition of ethanethiol to the enzyme. This process is followed by a very slow reduction of the enzyme. The fast reaction, which represents a binding reaction of ethanethiol to cytochrome c oxidase, has a k1 of 33 M?1 · s?1 and dissociation constant Kd of 3.9 mM.4. Ethanethiol induces fast spectral changes in the absorption spectrum of cytochrome c, which are followed by a very slow reduction of the heme. The rate constant for the fast ethanethiol reaction representing a bimolecular binding step is 50 M?1 · s?1 and the dissociation constant is about 2 mM. Addition of up to 25 mM ethanethiol to ferrocytochrome c does not cause spectral changes.5. EPR (electron paramagnetic resonance) spectra of cytochrome c oxidase, incubated with methanethiol or ethanethiol in the presence of cytochrome c and ascorbate, show the formation of low-spin cytochrome a3-mercaptide compounds with g values of 2.39, 2.23, 1.93 and of 2.43, 2.24, 1.91, respectively. 相似文献
9.
The respiratory pigments of Prototheca zopfii include seven cytochromes: two c-type cytochromes, a soluble c(549) and a membrane bound c(551); three b-type cytochromes, b(555), b(559) and b(564); and cytochromes a and a3. Cytochromes a and a3 could be resolved spectrally in the α-band region by reducing the cells in the presence of methanol and cyanide. Methanol shifted the absorption maximum of cytochrome a from 598 to 603 nanometers and permitted dithionite (or substrate) to reduce the cyanide-cytochrome a3 complex to give a well defined 595-nanometer absorption band. Methanol did not interfere with CO binding by cytochrome a3, and CO did not alter the methanol effect on cytochrome a. Azide and cyanide, which partially inhibited exogenous respiration, stimulated endogenous respiration. Frozen steady states of the electron transport chain in the presence of cyanide and azide indicated that the stimulation by these inhibitors was due to an increased autooxidation of one of the b-type cytochromes, possibly b(564). 相似文献
10.
The incorporation of radioactive amino acids and of δ-amino[2,3-3H2]laevulinate into rat liver cytochromes b5 and c and cytochrome oxidase has been examined with and without protein-synthesis inhibitors. Cycloheximide promptly inhibits labelling of both haem and protein for cytochrome c in parallel fashion. Although incorporation of 14C-labelled amino acid into microsomal cytochrome b5 is also rapidly inhibited, cycloheximide incompletely inhibits haem labelling of cytochrome b5 and cytochrome a+a3, and inhibition occurs only after repeated antibiotic injections. The possibility of apo-protein pools, or of haem exchange, with a rapidly renewed `free' haem pool, is considered. Consistent with this model is the observation of non-enzymic haem exchange in vitro between cytochrome b5 and methaemoglobin. Chloramphenicol, injected intravenously over 5h, results in a 20–40% decrease in incorporation of δ-amino[2,3-3H2]laevulinate into haem a+a3 and haem of cytochromes b5 and c. With the dosage schedule of chloramphenicol studied, amino acid labelling of total liver protein and of cytochrome c was not inhibited. Similarly, ferrochelatase activity was not decreased. 相似文献
11.
The antibiotic funiculosin mimics the action of antimycin in several ways. It inhibits the oxidation of NADH and succinate, but not TMPD+ascorbate. The titer for maximal inhibition in Mg2+-ATP particles (0.4–0.6 nmol/mg protein) is close to the concentrations of cytochromes b and cc1. Funiculosin also induces the oxidation of cytochromes cc1 and an extra reduction of cytochrome b in the aerobic steady state, and it inhibits duroquinol-cytochrome c reductase activity in isolated Complex III. The location of the funiculosin binding site is clearly similar to that of antimycin. In addition, funiculosin, like antimycin, prevents electron transport from duroquinol to cytochrome b in isolated Complex III if the complex is pre-reduced with ascorbate. Funiculosin and antimycin differ, however, in the manner in which they modulate the reduction of cytochrome b by ascorbate+TMPD. 相似文献
12.
Irene Díaz-Moreno Rinske Hulsker Pavol Skubak Johannes M. Foerster Davide Cavazzini Michelina G. Finiguerra Antonio Díaz-Quintana Blas Moreno-Beltrán Gian-Luigi Rossi G. Matthias Ullmann Navraj S. Pannu Miguel A. De la Rosa Marcellus Ubbink 《BBA》2014
The rapid transfer of electrons in the photosynthetic redox chain is achieved by the formation of short-lived complexes of cytochrome b6f with the electron transfer proteins plastocyanin and cytochrome c6. A balance must exist between fast intermolecular electron transfer and rapid dissociation, which requires the formation of a complex that has limited specificity. The interaction of the soluble fragment of cytochrome f and cytochrome c6 from the cyanobacterium Nostoc sp. PCC 7119 was studied using NMR spectroscopy and X-ray diffraction. The crystal structures of wild type, M58H and M58C cytochrome c6 were determined. The M58C variant is an excellent low potential mimic of the wild type protein and was used in chemical shift perturbation and paramagnetic relaxation NMR experiments to characterize the complex with cytochrome f. The interaction is highly dynamic and can be described as a pure encounter complex, with no dominant stereospecific complex. Ensemble docking calculations and Monte-Carlo simulations suggest a model in which charge–charge interactions pre-orient cytochrome c6 with its haem edge toward cytochrome f to form an ensemble of orientations with extensive contacts between the hydrophobic patches on both cytochromes, bringing the two haem groups sufficiently close to allow for rapid electron transfer. This model of complex formation allows for a gradual increase and decrease of the hydrophobic interactions during association and dissociation, thus avoiding a high transition state barrier that would slow down the dissociation process. 相似文献
13.
A membrane-bound cytochrome oxidase from Azobacter vinelandii was purified 20-fold using a detergent-solubilization procedure. Activity was monitored using an ascorbate-TMPD oxidation assay. The oxidase was ‘solubilized’ from a sonic-type electron-transport particle (R3 fraction) using Triton X-100 and deoxycholate. Low detergent concentrations first solubilized the flavoprotein oxidoreductases, then higher concentrations of Triton X-100 and KCl solubilized the oxidase, which was precipitated at 27–70% (NH4)2SO4. The highly purified cytochrome oxidase has a V of 60–78 μgatom O consumed/min per mg protein. TMPD oxidation by the purified enzyme was inhibited by CO, KCN, NaN3 and NH2OH; NaNO2 (but not NaNO3) also had a potent inhibitory effect. Spectral analyses revealed two major hemoproteins, the c-type cytochrome c4 and cytochrome o; cytochromes a1 and d were not detected. The Azotobacter cytochrome oxidase is an integrated cytochrome c4?o complex, TMPD-dependent cytochrome oxidase activity being highest in preparations having a high c-type cytochrome content. This TMPD-dependent cytochrome oxidase serves as a major oxygen-activation site for the A. vinelandii respiratory chain. It appears functionally analogous to cytochrome a+a3 oxidase of mammalian mitochondria. 相似文献
14.
Günter A. Peschek 《BBA》1981,635(3):470-475
The cytochrome content of membrane fragments prepared from the bluegreen alga (cyanobacterium) Anacystis nidulans was examined by difference spectrophotometry. Two b-type cytochromes and a hitherto unknown cytochrome a could be characterized. In the reduced-minus-oxidised difference spectra the a-type cytochrome showed an α-band at 605 nm and a γ-band at 445 nm. These bands shifted to 590 and 430 nm, respectively, in CO difference spectra. NADPH, NADH and ascorbate reduced the cytochrome through added horse heart cytochrome c as electron mediator. In presence of KCN the reduced-minus-oxidised spectrum showed a peak at 600 nm and a trough at 604 nm. Photoaction spectra of O2 uptake and of horse heart cytochrome c oxidation by CO-inhibited membranes showed peaks at 590 and 430 nm. These findings are consistent with cytochrome aa3 being the predominant respiratory cytochrome c oxidase in Anacystis nidulans. 相似文献
15.
1. In the absence of cytochrome c, ferrocyanide or ferrous sulphate reduces cytochrome c oxidase (EC 1.9.3.1), but no continuous oxygen uptake ensues, as it does with N,N,N′,N′-Tetramethyl-p-phenylenediamine or reduced phenazine methosulphate as reductants, unless a substoichiometric amount of cytochrome c or an excess of clupein is present. Cytochrome c cannot be replaced by porphyrin cytochrome c.2. Cytochrome c, porphyrin cytochrome c and clupein all stimulate the reduction of cytochrome aa3 by ferrocyanide.3. A model is proposed to explain these findings in which a high-affinity site for cytochrome c on the oxidase regulates the access of hydrophilic electron donors to a low-affinity site, and reduction via the high-affinity site is required for continuous oxygen uptake.4. Furthermore, it is shown that upon reaction of oxidase with ferrocyanide, cyano-oxidase is formed. 相似文献
16.
(1) Cells of ThiobacillusA2 grown chemoautotrophically on thiosulfate or heterotrophically on succinate with oxygen contained b-, c-, o-, a- and a3-type cytochromes. The amount of cytochrome per mg of cell protein was much greater in thiosulfate-grown cells and differences in the relative concentrations of cytochromes were observed for the different growth conditions. (2) The half-reduction potentials at pH 7.0 (Em,7.0) and spectral maxima of c-, b-, a- and a3-type cytochromes were similar in cells grown aerobically with thiosulfate or with succinate as the growth substrate. (3) The half-reduction potential of the ‘invisible’, or high-potential copper, as determined from the potentiometric behavior of the carbon monoxide-reduced cytochrome a3 complex at pH 8.0, was 365 mV. (4) Reducing equivalents from thiosulfate appear to enter the respiratory chain at the cytochrome c level; however, studies in cell-free extracts were limited due to a loss in respiratory activity with thiosulfate as a substrate upon cell disruption. 相似文献
17.
Irradiation of cells of Prototheca zopfii with blue light inhibited the respiratory capacity of the cells. The inhibition of respiration was correlated with a photodestruction of cytochrome c(551), cytochrome b(559), and cytochrome a3. Cytochrome c(549), cytochrome b(555), and cytochrome b(564) were unaffected by the irradiation treatment. The α-band of reduced cytochrome a was shifted from 599 to 603 nm by irradiation, an effect similar to that observed when methanol was added to nonirradiated cells. The presence of oxygen was required during irradiation for both photoinhibition of respiration and photodestruction of the cytochromes. Cytochrome a3 was protected against photodestruction by cyanide. Photodestruction of these same cytochromes also occurred when washed mitochondria of P. zopfii were irradiated. 相似文献
18.
Elisabeth C. Lowe Sarah Bydder Robert S. Hartshorne Hannah L. U. Tape Elizabeth J. Dridge Charles M. Debieux Konrad Paszkiewicz Ian Singleton Richard J. Lewis Joanne M. Santini David J. Richardson Clive S. Butler 《The Journal of biological chemistry》2010,285(24):18433-18442
Selenate reductase (SER) from Thauera selenatis is a periplasmic enzyme that has been classified as a type II molybdoenzyme. The enzyme comprises three subunits SerABC, where SerC is an unusual b-heme cytochrome. In the present work the spectropotentiometric characterization of the SerC component and the identification of redox partners to SER are reported. The mid-point redox potential of the b-heme was determined by optical titration (Em + 234 ± 10 mV). A profile of periplasmic c-type cytochromes expressed in T. selenatis under selenate respiring conditions was undertaken. Two c-type cytochromes were purified (∼24 and ∼6 kDa), and the 24-kDa protein (cytc-Ts4) was shown to donate electrons to SerABC in vitro. Protein sequence of cytc-Ts4 was obtained by N-terminal sequencing and liquid chromatography-tandem mass spectrometry analysis, and based upon sequence similarities, was assigned as a member of cytochrome c4 family. Redox potentiometry, combined with UV-visible spectroscopy, showed that cytc-Ts4 is a diheme cytochrome with a redox potential of +282 ± 10 mV, and both hemes are predicted to have His-Met ligation. To identify the membrane-bound electron donors to cytc-Ts4, growth of T. selenatis in the presence of respiratory inhibitors was monitored. The specific quinol-cytochrome c oxidoreductase (QCR) inhibitors myxothiazol and antimycin A partially inhibited selenate respiration, demonstrating that some electron flux is via the QCR. Electron transfer via a QCR and a diheme cytochrome c4 is a novel route for a member of the DMSO reductase family of molybdoenzymes. 相似文献
19.
Cytochrome a-type terminal oxidase was purified from Thiobacillus novellus to an electrophoretically homogeneous state and some of its properties were studied.The enzyme shows absorption peaks at 428 and 602 nm in the oxidized form, and at 442 and 602 nm in the reduced form. The CO compound of the reduced enzyme shows peaks at 431 and 599 nm. The enzyme has 1 mol of haem a and 1 g-atom of copper per 55 600 g and is composed of two kinds of subunit, of 32 000 and 23 000 daltons, respectively.The enzyme reacts rapidly with tuna, bonito and yeast cytochromes c as well as with T. novellus cytochrome c, while it reacts slowly with horse and cow cytochromes c. The reduction product of oxygen catalysed by the enzyme is water. 相似文献
20.
Rhodospirillum rubrum chromatophores associated with a planar phospholipid macromembrane by bivalent cations in the presence of quinone, N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD) and ascorbate generate a transmembrane electrical potential difference in the light. Photoelectrical activity is also observed if chromatophores are preincubated with cytochrome c; the maximum values of responses are reached upon subsequent addition of ascorbate and menadion in the absence of bivalent cations and TMPD. The cytochrome c-dependent responses of the illuminated chromatophores are inhibited by Ca2+ and prevented by quinones. The possibility of cytochrome c (c2) translocation across the chromatophore membrane and the mechanism of charge transfer across the planar phospholipid membrane are discussed. 相似文献