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1.
Ken-ichiro Takamiya  Shigemi Obata 《BBA》1986,852(2-3):198-202
The photosynthetic membranes from Rhodopseudomonas palustris contained one species of membrane-bound c-type cytochrome, presumably cytochrome c1, and a b-type cytochrome with two heme centers. The molecular weight and midpoint potential of cytochrome c1 were 30000 and 275 mV, respectively. The peak of the reduced-minus-oxidized difference spectrum of cytochrome c1 was at 552 nm. Molecular weight of the b-type cytochrome was 32000 and the cytochrome had two midpoint potentials of 60 mV and −55 mV. The peaks of the reduced-minus-oxidized difference spectra of the high and low midpoint potential heme centers were at 560 and 562 nm, respectively. These results suggested that there was a cytochrome b-c1 complex in Rps. palustris.  相似文献   

2.
The orientations of high potential cytochromes with respect to photosynthetic membranes was investigated in spinach chloroplasts and in Rhodopseudomonas viridis. The general approach consists in detection with polarized light of photoinduced absorbance changes related to the oxidation of the cytochromes. The orientation of cytochrome c-558 was measured at room temperature in chromatophores and whole cells of Rps. viridis, oriented on glass slides and in a magnetic field, respectively. The orientation of cytochrome b-559 of green plants was detected at 77 K in magnetically oriented chloroplasts. In both cases the dichroic ratio for the band shows that the heme plane makes an angle greater than 35°C with the membrane plane. Moreover, the dichroic ratio is not constant throughout the and β bands, for both cytochrome c-558 and b-559. Linear dichroism spectra of oriented pure horse heart cytochrome c and cytochrome c2 of Rhodopseudomonas sphaeroides in stretched polyvinyl alcohol films show that the variations of the dichroic ratio in the and β bands can be explained by the occurrence of x- and y-polarized transitions absorbing at slightly different wavelengths.  相似文献   

3.
Bayard T. Storey 《BBA》1973,292(3):592-602

1. 1. Cycles of oxidation followed by reduction at pH 7.2 have been induced in uncoupled anaerobic mung bean mitochondria treated with succinate and malonate by addition of oxygen-saturated medium. Under the conditions used, cytochromes b557, b553, c549 (corresponding to c1 in mammalian mitochondria) and ubiquinone are completely oxidized in the aerobic state, but become completely reduced in anaerobiosis.

2. 2. The time course of the transition from fully oxidized to fully reduced in anaerobiosis was measured for cytochromes c549, b557, and b553. The intramitochondrial redox potential (IMPh) was calculated as a function of time for each of the three cytochromes from the time course of the oxidized-to-reduced transition and the known midpoint potentials of the cytochromes at pH 7.2. The three curves so obtained are superimposable, showing that the three cytochromes are in redox equilibrium under these conditions during the oxidized-to-reduced transition.

3. 3. This result shows that the slow reduction of cytochrome b557 under these conditions, heretofore considered anomalous, is merely a consequence of its more negative midpoint potential of +42 mV at pH 7.2, compared to +75 mV for cytochrome b553 and +235 mV for cytochrome c549. Cytochrome b557 is placed on the low potential side of coupling site II and transfers electrons to cytochrome c549 via the coupling site.

4. 4. The time course of the transition from fully oxidized to fully reduced was also measured for ubiquinone. Using the change in intramitochondrial potential IMPh with time obtained from the three cytochromes, the change in redox state of ubiquinone with IMPh was calculated. When replotted as IMPh versus the logarithm of the ratio (fraction oxidized)/(fraction reduced), two redox components with n = 2 were found. The major component is ubiquinone with a midpoint potential Em7.2 = + 70 mV. The minor component has a midpoint potential Em7.2 = − 12 mV; its nature is unknown.

Abbreviations: IMPh, intramitochondrial potential, referred to the normal hydrogen electrode; Em7.2, midpoint potential at pH 7.2  相似文献   


4.
M  rten K. F. Wikstr  m  Jan A. Berden 《BBA》1972,283(3):403-420
1. The effect of oxidizing equivalents on the redox state of cytochrome b in the presence of antimycin has been studied in the presence and absence of various redox mediators.

2. The antimycin-induced extra reduction of cytochrome b is always dependent on the initial presence of an oxidant such as oxygen. After removal of the oxidant this effect remains or is partially (under some conditions even completely) abolished depending on the redox potential of the substrate used and the leak through the antimycin-inhibited site.

3. The increased reduction of cytochrome b induced by oxidant in the presence of antimycin involves all three spectroscopically resolvable b components (b-562, b-566 and b-558.

4. Redox mediators with an actual redox potential of less than 100–170 mV cause the oxidation of cytochrome b reduced under the influence of antimycin and oxidant.

5. Redox titrations of cytochrome b with the succinate/fumarate couple were performed aerobically in the presence of cyanide. In the presence of antimycin two b components are separated potentiometrically, one with an apparent midpoint potential above 80 mV (at pH 7.0), outside the range of the succinate/fumurate couple, and one with an apparent midpoint potential of 40 mV and an n value of 2. In the absence of antimycin cytochrome b titrates essentially as one species with a midpoint potential of 39 mV (at pH 7.0) and n = 1.14.

6. The increased reducibility of cytochrome b induced by antimycin plus oxidant is considered to be the result of two effects: inhibition of oxidation of ferrocytochrome b by ferricytochrome c1 (the effect of antimycin), and oxidation of the semiquinone form of a two-equivalent redox couple such as ubiquinone/ubiquinol by the added oxidant, leading to a decreased redox potential of the QH2/QH couple and reduction of cytochrome b.  相似文献   


5.
Three c-type cytochromes (c-551, c-553, c-555) have been isolated and characterized from a strain of the green photosynthetic bacterium Chlorobium thiosulfatophilum. These cytochromes are atypical when compared to horse heart cytochrome c in many properties, among them: oxidation-reduction potential at pH 7.0 (c-551, 135 mV; c-553, 98 mV; c-555, 145 mV), molecular weight (c-551, 45000–60000; c-553, 50000; c-555, 10000) and isolelectric point (c-551, 6.0; c-553, 6.7). No protoheme was detected in whole cells or cell-free extracts.  相似文献   

6.
Considerable progress was made over the last few years in understanding the mechanism of folding of cytochrome c551, a small acidic hemeprotein from Pseudomonas aeruginosa. Comparison of our results with those obtained by others on horse heart cytochrome c allows to draw some general conclusions on the structural features that are common determinants in the folding of members of the cytochrome c family.  相似文献   

7.
(1) Cells of Rhodopseudomonas capsulata (wild-type) were grown photoheterotrophically in a turbidostat under very high and very low light intensity. Membranes were isolated from cells adapted to the respective light conditions and fractionated by sucrose density centrifugation. The molar ratios of ubiquinone and cytochromes c2, c1, b-561 and b-566 per reaction center were 3-fold to 5-fold higher in high-light than in low-light membranes. (2) Most of the Cyt(c1 + c2) and Cyt b-561 detected in dark redox titrations undergoes light-induced redox changes, both in high- and in low-light membranes. (3) The fractions of the total photooxidizable reaction center and Cyt(c1 + c2) oxidized under continuous light in the absence of antimycin are higher in membranes from low-light- than from high-light-grown cells. (4) From these data and results of kinetic studies it is proposed that cyclic electron flow under saturating light intensities is faster in high-light-grown cells.  相似文献   

8.
1. In membranes prepared from dark grown cells of Rhodopseudomonas capsulata, five cytochromes of b type (E0 at pH 7.0 +413±5, +270±5, +148±5, +56±5 and −32±5 mV) can be detected by redox titrations at different pH values. The midpoint potentials of only three of these cytochromes (b148, b56, and b−32) vary as a function of pH with a slope of 30 mV per pH unit.

2. In the presence of a Co/N2 mixture, the apparent E0 of cytochrome b270 shifts markedly towards higher potentials (+355 mV); a similar but less pronounced shift is apparent also for cytochrome b150. The effect of CO on the midpoint potential of cytochrome b270 is absent in the respiration deficient mutant M6 which possesses a specific lesion in the CO-sensitive segment of the branched respiratory chain present in the wild type strain.

3. Preparations of spheroplasts with lysozyme digestion lead to the release of a large amount of cytochrome c2 and of virtually all cytochrome cc′. These preparations show a respiratory chain impaired in the electron pathway sensitive to low KCN concentration, in agreement with the proposed role of cytochrome c2 in this branch; on the contrary, the activity of the CO-sensitive branch remains unaffected, indicating that neither cytochrome c2 nor the CO-binding cytochrome cc′ are involved in this pathway.

4. Membranes prepared from spheroplasts still possess a CO-binding pigment characterized by maxima at 420.5, 543 and 574 nm and minima at 431, 560 nm in CO-difference spectra and with an band at 562.5 nm in reduced minus oxidized difference spectra. This membrane-bound cytochrome, which is coincident with cytochrome b270, can be classified as a typical cytochrome “o” and considered the alternative CO-sensitive oxidase.  相似文献   


9.
T.E. Meyer  S.J. Kennel  S.M. Tedro  M.D. Kamen 《BBA》1973,292(3):634-643
Four out of five soluble electron transport iron proteins of Thiocapsa pfennigii plus the particulate cytochromes have been found to be analogous to those of Chromatium vinosum strain D. In addition to ferredoxin, high potential iron-sulfur protein. cytochrome c′, and cytochrome c-552(550), T. pfennigii contains a cytochrome c-552(545) not previously isolated from photosynthetic bacteria. It is concluded that T. pfennigii is more closely related to C. vinosum than to Rhodopseudomonas viridis, the only other known bacterial species having bacteriochlorophyll b.  相似文献   

10.
P.Muir Wood 《BBA》1974,357(3):370-379
The rate of electron transfer between reduced cytochrome ƒ and plastocyanin (both purified from parsley) has been measured as k = 3.6 · 107 M−1 · s−1, at 298 °K and pH 7.0, with activation parameters ΔH = 44 kJ · mole−1 and ΔS = +46 J · mole−1 · °K−1. Replacement of cytochrome ƒ with red algal cytochrome c-553, Pseudomonas cytochrome c-551 and mammalian cytochrome c gave rates at least 30 times slower: k = 5 · 105, 7.5 · 105 and 1.0 · 106 M−1 · s−1, respectively.

Similar measurements made with azurin instead of plastocyanin gave k = 6 · 106 and approx. 2 · 107 M−1 · s−1 for reaction of reduced azurin with cytochrome ƒ and algal cytochrome respectively.

Rate constants of 115 and 80 M−1 · s−1 were found for reduction of plastocyanin by ascorbate and hydroquinone at 298 °K and pH 7.0. The rate constants for the oxidation of plastocyanin, cytochrome ƒ, Pseudomonas cytochrome c-551 and red algal cytochrome c-553 by ferricyanide were found to be between 3 · 104 and 8 · 104 M−1 · s−1.

The results are discussed in relation to photosynthetic electron transport.  相似文献   


11.
Roger C. Prince  P.Leslie Dutton 《BBA》1977,462(3):731-747
We have examined the thermodynamic properties of the physiological electron donor to ferricytochrome c2 in chromatophores from the photosynthetic bacterium Rhodopseudomonas sphaeroides. This donor (Z), which is capable of reducing the ferri-cytochrome with a halftime of 1–2 ms under optimal conditions, has an oxidation-reduction midpoint potential of close to 150 mV at pH 7.0, and apparently requires two electrons and two protons for its equilibrium reduction.

The state of reduction of Z, which may be a quinone · protein complex near the inner (cytochrome c2) side of the membrane, appears to govern the rate at which the cyclic photosynthetic electron transport system can operate. If Z is oxidized prior to the flash-oxidation of cytochrome c2, the re-reduction of the cytochrome takes hundreds of milliseconds and no third phase of the carotenoid bandshift occurs. In contrast if Z is reduced before flash activation, the cytochrome is rereduced within milliseconds and the third phase of the carotenoid bandshift occurs. The prior reduction of Z also has a dramatic effect on the uncoupler sensitivity of the rate of electron flow; if it is oxidized prior to activation, uncoupler can stimulate the cytochrome re-reduction after several turnovers by less than tenfold, but if it is reduced prior to activation, the stimulation after several turnovers can be as dramatic as a thousandfold. The results suggest that Z plays a central role in controlling electron and proton movements in the ubiquinone cytochrome b-c2 oxido-reductase.  相似文献   


12.
I. Y. Lee  E. C. Slater 《BBA》1972,283(3):395-402
Under anaerobic conditions cytochrome b in beef-heart mitochondria is partially reduced in the presence of NADH, whereas other cytochromes are completely reduced. Addition of antimycin together with oxygen under these conditions causes an immediate reduction of cytochromes b-558, b-562 and b-566 and oxidation of cytochrome c. During the subsequent transient aerobic steady state cytochromes b-558 and b-566 are rapidly re-oxidized without changes in redox state of cytochrome c, but cytochrome b-562 remains reduced. When oxygen is consumed by the leak through or around the antimycin-inhibition site, cytochrome b-562 becomes oxidized with concomitant reduction of cytochrome c.

The cytochromes b in lyophilized beef-heart mitochondria are more readily accessible to electrons from NADH, and in the presence of antimycin and NADH a complete and stable reduction is obtained under both aerobic and anaerobic conditions. Gradual addition of rotenone under these conditions causes re-oxidation of cytochromes b in which oxidation of cytochromes b-558 and b-566 precedes that of cytochrome b-562.

It is concluded that (1) the effect of antimycin in the presence of oxygen involves all three cytochromes b, (2) the reducibility of the cytochromes b in the aerobic steady state of antimycin-treated mitochondria is dependent upon the potential of the substrate redox couple registered on the cytochromes, and (3) the midpoint potential of cytochrome b-562 in the presence of antimycin is higher than that of cytochrome b-558 or b-566.  相似文献   


13.
The kinetics of the reaction of hydrated electron (eaq) and carboxyl anion radical (CO2) with Pseudomonas aeruginosa ferricytochrome c-551 were studied by pulse radiolysis. The rate of reaction of eaq with the negatively charged ferricytochrome c-551 (17 nM−1 · s−1) is significantly slower than the larger positively charged horse heart ferricytochrome c (70 nM · s). This difference cannot be explained solely by electrostatic effects on the diffusion-controlled reactions. After the initial encounter of eaq with the protein, ferricytochrome c-551 is less effective in transferring an electron to the heme which may be due to the negative charge on the protein. The charge on ferricytochrome c-551 is estimated to be −5 at pH 7 from the effect of ionic strength on the reaction rate. A slower relaxation (2 · 104 s−1) observed after fast eaq reduction is attributed to a small conformational change. The rate of reaction of CO2 with ferricytochrome c-551 (0.7 nM−1 · s) is, after electrostatic correction, the same as ferricytochrome c, indicating that the steric requirements for reaction are similar. This reaction probably takes place through the exposed heme edge.  相似文献   

14.
Roger C. Prince  P.Leslie Dutton 《BBA》1975,387(3):609-613
In Rhodopseudomonas sphaeroides, following a single-turnover flash of light, cytochrome c2 is oxidized by reaction center bacteriochlorophyll, and a cytochrome b is reduced by the primary electron acceptor, probably via ubiquinone. In this report we show that, in the uncoupled state, the rate of re-oxidation of the cytochrome b is identical to the rate of reduction of the cytochrome c2, a kinetic completion of the cyclic photosynthetic electron transport system.  相似文献   

15.
Fast reaction kinetic experiments on the electron transfer reaction between azurin and cytochrome c551 isolated from Pseudomonas aeruginosa confirmed the existence of two redox forms of reduced azurin previously reported. The pH dependence of the amplitudes of the relaxation processes observed in temperature jump experiments indicate that these two redox forms are in pH dependent equilibrium. The pH independence of the overall equilibrium constant indicates that redox active and inactive forms of cytochrome c551 may also exist. Evidence that reduced cytochrome c551 undergoes a pH transition is given by optical spectrophotometry. The nature of the transition is discussed in the context of recent nmr studies and in terms of the Marcus theory of electron transfer. The metabolic consequences of these transitions are also discussed.  相似文献   

16.
Linda Yu  Jian-Hua Dong  Chang-An Yu 《BBA》1986,852(2-3):203-211
Cytochrome c1 from a photosynthetic bacterium Rhodobacter sphaeroides R-26 has been purified to homogeneity. The purified protein contains 30 nmol heme per mg protein, has an isoelectric point of 5.7, and is soluble in aqueous solution in the absence of detergents. The apparent molecular weight of this protein is about 150 000, determined by Bio Gel A-0.5 m column chromatography; a minimum molecular weight of 30 000 is obtained by sodium dodecylsulfate polyacrylamide gel electrophoresis. The absorption spectrum of this cytochrome is similar to that of mammalian cytochrome c1, but the amino acid composition and circular dichroism spectral characteristics are different. The heme moiety of cytochrome c1 is more exposed than is that of mammalian cytochrome c1, but less exposed than that of cytochrome c2. Ferricytochrome c1 undergoes photoreduction upon illumination with light under anaerobic conditions. Such photoreduction is completely abolished when p-chloromercuriphenylsulfonate is added to ferricytochrome c1, suggesting that the sulfhydryl groups of cytochrome c1 are the electron donors for photoreduction. Purified cytochrome c1 contains 3 ± 0.1 mol of the p-chloromercuriphenylsulfonate titratable sulfhydryl groups per mol of protein. In contrast to mammalian cytochrome c1, the bacterial protein does not form a stable complex with cytochrome c2 or with mammalian cytochrome c at low ionic strength. Electron transfer between bacterial ferrocytochrome c1 and bacterial ferricytochrome c2, and between bacterial ferrocytochrome c1 and mammalian ferricytochrome c proceeds rapidly with equilibrium constants of 49 and 3.5, respectively. The midpoint potential of purified cytochrome c1 is calculated to be 228 mV, which is identical to that of mammalian cytochrome c1.  相似文献   

17.
George D. Case  William W. Parson   《BBA》1973,292(3):677-684
The isoionic pH of Chromatium chromatophores is 5.2±0.1. At pH 7.7, the net charge on the chromatophore is approx. −1·104. If a change in this charge accompanies the oxidation of an electron carrier, the midpoint redox potential (Em) of that carrier should be a function of the solution ionic strength (I). of that carrier should be a function of the solution ionic strength (I).

The Em values of P870 and cytochrome c-555 increase strongly with increasing I at low values of I. The Em of cytochrome c-552 also increases with increasing I, though not so strongly. These effects probably cannot be attributed to an influence of I on the activity coefficient of a dissociable ion. We conclude that, when either P870 or cytochrome c-555 loses an electron, no specific ions (including protons) are bound or released in significant amounts, and the absolute value of the charge on the chromatophore decreases.

The Em values of the primary and secondary electron acceptors, X and Y, do not depend on I. Because these Em values have been shown previously to depend on pH, we conclude that the uptake of a proton keeps the charge on the chromatophore constant when either X or Y accepts an electron. This means that the primary and secondary electron transfer reactions in Chromatium result in a net decrease in the charge on the photosynthetic membrane. They do not result in the translocation of protons across the membrane.

The Em of the soluble flavocytochrome c-552 from Chromatium depends only weakly on I, but depends strongly on the pH. The uptake of a proton appears to keep the net charge on this cytochrome constant upon reduction.  相似文献   


18.
A photosynthetically-incompetent mutant Rhodopseudomonas spheroides that lacks bacteriochlorophyll was isolated. Spectroscopic evidence from CO difference spectra and cyanide difference spectra suggested that a cytochrome oxidase was present in this mutant that contained two components, corresponding to cytochromes a and a3 of mitochondria. Potentiometric titration at 607 nm also showed the presence of two components with oxidation-reduction mid-point potentials of +375 mV and +200 mV. They were present in a ratio close to unity. No cytochrome of the the c-type corresponding to mitochondrial cytochrome c was detected, but a minor c component (near 10% of the total cytochrome c) with an oxidation-reduction mid-point potential of +120 mV was found

Growth of the mutant in medium with low aeration or lacking added copper diminished the concentration of the a-type cytochrome but not the concentrations of cytochromes of the b and c-type.  相似文献   


19.
Desulfovibrio vulgaris Hildenborough cytochrome c3 contains four hemes in a low-spin state with bis-histidinyl coordination. High-spin forms of cytochrome c3 can be generated by protonation of the axial ligands in order to probe spin equilibrium (low-spin/high-spin). The spin alterations occurring at acid pH, the associated changes in redox potentials, as well as the reactivity towards external ligands were followed by the conjunction of square wave voltammetry and UV–visible, CD, NMR and EPR spectroscopies. These processes may be used for modelling the action of enzymes that use spin equilibrium to promote enzyme activity and reactivity towards small molecules.  相似文献   

20.
J.Peter Kusel  Bayard T. Storey 《BBA》1973,305(3):570-580
Highly purified mitochondrial preparations from the trypanosomatid hemoflagellate, Crithidia fasciculata (A.T.C.C. No.11745), were examined by low-temperature difference spectroscopy. The cytochrome a+a3 maximum of hypotonically-treated mitochondria reduced with succinate, was shifted from 605 nm at room temperature to 601 nm at 77 °K. The Soret maximum, found at 445 nm at 23 °C, was split at 77 °K into two approximately equally absorbing species with maxima at 438 and 444 nm. A prominent shoulder observed at 590 nm with hypotonically-treated mitochondria was not present in spectra of isotonic controls.

The cytochrome b maxima observed in the presence of succinate plus antimycin A were shifted from the 431 and 561 nm positions observed at 23 °C to 427 and 557 nm at 77 °K. Multiple b cytochromes were not apparent.

Unlike other soluble c-type cytochromes, the maximum of cytochrome c555 was not shifted at 77 °K although it was split to give a 551 nm shoulder adjacent to the 555 nm maximum. This lack of a low-temperature blue shift was true for partially purified hemoprotein preparations as well as in situ in the mitochondrial membrane.

Using cytochrome c555-depleted mitochondria, a cytochrome c1 pigment was observed with a maximum at 420 nm and multiple maxima at 551, 556, and 560 nm. After extraction of non-covalently bound heme, the pyridine hemochromogen difference spectrum of cytochrome c555-depleted preparations exhibited an maximum at 553 nm at room temperature.

The reduced rate of succinate oxidation by cytochrome c555-depleted mitochondria and the ferricyanide requirement for the reoxidation of cytochrome c1, even in the presence of antimycin, indicated that cytochrome c555-mediated electron transfer between cytochromes c1 and a+a3 in a manner analagous to that of cytochrome c in mammalian mitochondria.  相似文献   


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