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1.
A temperature dependence of multiheme cytochrome c oxidation induced by a laser pulse was studied in photosynthetic reaction center preparations from Chromatium minutissimum. Absorbance changes and kinetic characteristics of the reaction were measured under redox conditions where one or all of the hemes of the cytochrome subunit are chemically reduced (E h =+300 mV or E h =–20 to -60 mV respectively). In the first case photooxidation is inhibited at temperatures lower than 190–200 K with the rate constant of the photooxidation reaction being practically independent on temperature over the range of 300 to 190 K (k=2.2×105 s-1). Under reductive conditions (E h =–20 to -60 mV) lowering the temperature to 190–200 K causes the reaction to slow from k=8.3×105 s-1 to 2.1×104 s-1. Under further cooling down to the liquid nitrogen temperature, the reaction rate changes negligibly. The absorption amplitude decreases by 30–40% on lowering the temperature. A new physical mechanism of the observed critical effects of temperature on the rate and absorption amplitude of the multiheme cytochrome c oxidation reaction is proposed. The mechanism suggests a close interrelation between conformational mobility of the protein and elementary electron tunneling act. The effect of freezing conformational motion is described in terms of a local diffusion along a random rough potential.  相似文献   

2.
The arrangement and function of the redox centers of the mammalianbc 1 complex is described on the basis of structural data derived from amino acid sequence studies and secondary structure predictions and on the basis of functional studies (i.e., EPR data, inhibitor studies, and kinetic experiments). Two ubiquinone reaction centers do exist—a QH2 oxidation center situated at the outer, cytosolic surface of the cristae membrane (Q0 center), and a Q reduction center (Q i center) situated more to the inner surface of the cristae membrane. The Q0 center is formed by theb-566 domain of cytochromeb, the FeS protein, and maybe an additional small subunit, whereas the Q i center is formed by theb-562 domain of cytochromeb and presumably the 13.4kDa protein (QP-C). The Q binding proteins are proposed to be protein subunits of the Q reaction centers of various multiprotein complexes. The path of electron flow branches at the Q0 center, half of the electrons flowing via the high-potential cytochrome chain to oxygen and half of the electrons cycling back into the Q pool via the cytochromeb path connecting the two Q reaction centers. During oxidation of QH2, 2H+ are released to the cytosolic space and during reduction of Q, 2H+ are taken up from the matrix side, resulting in a net transport across the membrane of 2H+ per e flown from QH2 to cytochromec, the H+ being transported across the membrane as H (H+ + e) by the mobile carrier Q. The authors correct their earlier view of cytochromeb functioning as a H+ pump, proposing that the redox-linkedpK changes of the acidic groups of cytochromeb are involved in the protonation/deprotonation processes taking place during the reduction and oxidation of Q. The reviewers stress that cytochromeb is in equilibrium with the Q pool via the Q i center, but not via the Q0 center. Their view of the mechanisms taking place at the reductase is a Q cycle linked to a Q-pool where cytochromeb is acting as an electron pump.  相似文献   

3.
A spontaneous mutant (R/89) of photosynthetic purple bacterium Rhodobacter sphaeroides R-26 was selected for resistance to 200 M atrazin. It showed increased resistance to interquinone electron transfer inhibitors of o-phenanthroline (resistance factor, RF=20) in UQo reconstituted isolated reaction centers and terbutryne in reaction centers (RF=55) and in chromatophores (RF=85). The amino acid sequence of the QB binding protein of the photosynthetic reaction center (the L subunit) was determined by sequencing the corresponding pufL gene and a single mutation was found (IleL229 Met). The changed amino acid of the mutant strain is in van der Waals contact with the secondary quinone QB. The binding and redox properties of QB in the mutant were characterized by kinetic (charge recombination) and multiple turnover (cytochrome oxidation and semiquinone oscillation) assays of the reaction center. The free energy for stabilization of QAQB with respect to QA QB was GAB=–60 meV and 0 meV in reaction centers and GAB=–85 meV and –46 meV in chromatophores of R-26 and R/89 strains at pH 8, respectively. The dissociation constants of the quinone UQo and semiquinone UQo in reaction centers from R-26 and R/89 showed significant and different pH dependence. The observed changes in binding and redox properties of quinones are interpreted in terms of differential effects (electrostatics and mesomerism) of mutation on the oxidized and reduced states of QB.Abbreviations BChl bacteriochlorophyll - Ile isoleucine - Met methionin - P primary donor - QA primary quinone acceptor - QB secondary quinone acceptor - RC reaction center protein - UQo 2,3-dimethoxy-5-methyl benzoquinone - UQ10 ubiquinone 50 This work is dedicated to the memory of Randall Ross Stein (1954–1994) and is, in a small way, a testament to the impact which Randy's ideas have had on the development of the field of competitive herbicide binding.  相似文献   

4.
Kinetics of electron transfer from soluble cytochrome c2 to the tetraheme cytochrome c have been measured in isolated reaction centers and in membrane fragments of the photosynthetic purple bacterium Rhodopseudomonas viridis by time-resolved flash absorption spectroscopy. Absorbance changes kinetics in the region of cytochrome -bands (540–560 nm) were measured at 21 °C under redox conditions where the two high-potential hemes (c-559 and c-556) of the tetraheme cytochrome were chemically reduced. After flash excitation, the heme c-559 donates an electron to the special pair of bacteriochlorophylls and is then re-reduced by heme c-556. The data show that oxidized heme c-556 is subsequently re-reduced by electron transfer from reduced cytochrome c2 present in the solution. The rate of this reaction has a non-linear dependence on the concentration of cytochrome c2, suggesting a (minimal) two-step mechanism involving the f ormation of a complex between cytochrome c2 and the reaction center, followed by intracomplex electron transfer. To explain the monophasic character of the reaction kinetics, we propose a collisional mechanism where the lifetime of the temporary complex is short compared to electron transfer. The limit of the halftime of the bimolecular process when extrapolated to high concentrations of cytochrome c2 is 60 ± 20 s. There is a large ionic strength effect on the kinetics of electron transfer from cytochrome c2 to heme c-556. The pseudofirst-order rate constant decreases from 1.1 × 107 M-1 s-1 to 1.3 × 106 M-1 s-1 when the ionic strength is increased from 1 to 1000 mM. The maximum rate (1.1 × 107 M-1 s-1) was obtained at about 1 mM ionic strength. This dependence of the rate on ionic strength s uggests that attractive electrostatic interactions contribute to the binding of cytochrome c2 with the tetraheme cytochrome. On the basis of our data and of previous molecular modelling, it is proposed that cytochrome c2 docks close to the low-potential heme c-554 and reduces heme c-556 via c-554.  相似文献   

5.
The reaction between membrane-bound cytochrome c and the reaction center bacteriochlorophyll g dimer P798 was studied in the whole cells and isolated membranes of Heliobacterium gestii. In the whole cells, the flash-oxidized P798+ was rereduced in multiple exponential phases with half times (t 1/2s) of 10 s, 300 s and 4 ms in relative amplitudes of 40, 35 and 25%, respectively. The faster two phases were in parallel with the oxidation of cytochrome c. In isolated membranes, a significantly slow oxidation of the membrane-bound cytochrome c was detected with t 1/2 = 3 ms. This slow rate, however, again became faster with the addition of Mg2+. The rate showed a high temperature dependency giving apparent activation energies of 88.2 and 58.9 kJ/mol in the whole cells and isolated membranes, respectively. Therefore, membrane-bound cytochrome c donates electrons to the P798+ in a collisional reaction mode like the reaction of water-soluble proteins. The rereduction of the oxidized cytochrome c was suppressed by the addition of stigmatellin both in the whole cells and isolated membranes. This indicates that the electron transfer from the cytochrome bc complex to the photooxidized P798+ is mediated by the membrane-bound cytochrome c. The multiple flash excitation study showed that 2–3 hemes c were connected to the P798. By the heme staining after the SDS-PAGE analysis of the membraneous proteins, two cytochromes c were detected on the gel indicating apparent molecular masses of 17 and 30 kDa, respectively. The situation resembles the case in green sulfur bacteria, that is, the membrane-bound cyotochrome c z couples electron transfer between the cytochrome bc complex and the P840 reaction center complex.This revised version was published online in October 2005 with corrections to the Cover Date.  相似文献   

6.
(1) Two populations of reaction centers in the chromatophore membrane can be distinguished under some conditions of initial redox poise (300 mV < Eh < 400 mV): those which transfer a reducing equivalent after the first flash from the secondary quinone (QII) of the reaction center to cytochrome b of the ubiquinone-cytochrome c2 oxidoreductase; and those which retain the reducing equivalent on Q?II until a second flash is given. These two populations do not exchange on a time scale of tens of seconds. (2) At redox potentials higher than 400 mV, Q?II generated after the first flash is no longer able to reduce cytochrome b-560 even in those reaction centers associated with an oxidoreductase. Under these conditions, doubly reduced QII generated by a second flash is required for cytochrome b reduction, so that the QII effectively functions as a two-electron gate into the oxidoreductase at these high potentials. (3) At redox potentials below 300 mV, although the two populations of QII are no longer distinguishable, cytochrome b reduction is still dependent on only part of the reaction center population. (4) Proton binding does not oscillate under any condition tested.  相似文献   

7.
Both the soluble cytochrome c2 and the membrane-bound cytochrome cy act as secondary electron carriers in photoinduced cyclic electron transfer chain of Rhodobacter capsulatus [Jenney and Daldal (1993) EMBO J 12: 1283–1292]. In this work, we have studied the kinetics of electron transfer between these secondary electron donors and the reaction center in intact cells of two mutants, MT-G4/S4 and MT-GS18 deleted in cytochrome c2 and in cytochrome c2 plus cytochrome bc1 complex, respectively. In the MT-G4/S4 mutant, only about one third of the primary electron donor is reduced by cytochrome cy in less than five ms. The remaining fraction is reduced in several seconds, although about 90% of the photoxidized cytochrome cy is reduced in less than 10 ms by the cytochrome bc1 complex. This implies that cytochrome cy is not in thermodynamic equilibrium with the large fraction of primary donors which are slowly reduced. As shown by energy transfer measurements, the reaction centers connected to cytochrome cy and the disconnected reaction centers are localized in the same membrane region. We propose that the movement of cyt cy is restricted to a small membrane domain which includes a single cytochrome bc1 complex. The kinetics of cytochrome cy photooxidation in the MT-G4/S4 mutant in the presence of myxothiazol presents a fast phase (t1/2 3 µs) followed by a slower phase (t1/2 20 µs). In the case of the double mutant MT-GS18, the kinetics of electron transfer between cytochrome cy and the reaction center is highly multiphasic and much slower than those observed for the MT-G4/S4 mutant. In particular, the amplitude of the fast phase is decreased by more than a factor 2 and the 20-µs phase is not observed. This implies an important structural role of the cytochrome bc1 complex in the interaction between reaction center and cytochrome cy, and their formation in supercomplex. The more problable stoichiometry of electron carriers in this supercomplex is 2 reaction centers, 2 cytochrome cy and 1 cytochrome bc1 complex.  相似文献   

8.
1. The reduced minus oxidized extinction coefficients (Δred-ox) of reaction center P605 when in the chromatophore is about 20% smaller than in the detergent-isolated state. Presumably the coupling of the reaction center protein to the antenna bacteriochlorophylls and carotenoids causes this hypochromism. The chromatophore values for P605 are 19.5 mM−1 · cm−1 with the spectrophotometer on single beam mode at 605 nm, and 29.8 mM−1 · cm−1 on dual wavelength mode set at 605 – 540 nm. Cytochrome c2, which is not affected by detergent, has a Δred-ox value at 550-540 nm of 19.0 mM−1 · cm−1.2. The total bacteriochlorophyll to reaction center bacteriochlorophyll protein (P) ratio is about 100 : 1. The cytochrome c2: reaction center protein ratio approaches 2. In current French press chromatophore preparations, about 70% of the reaction centers are each associated on a rapid kinetic basis with two cytochrome c2 molecules (intact P-c2 units). The remaining reaction center proteins are not associated with cytochrome c2 on a kinetically viable basis and may be the result of damage incurred during mechanical rupture of the cells.3. The half-reduction potential of cytochrome c2 in the isolated state is 345 mV. In the chromatophore, two electrochemical species of cytochrome c2 are recognized. The majority has a value of approx. 295 mV and is identifiable with cytochrome c2 in a reaction center protein-associated state (kinetically active, intact P-c2 units); the remainder has an approx. 350 mV half-reduction potential and is probably cytochrome c2 in the “free” or reaction center-dissociated state (possibly from damaged P-c2 units). It appears that there is no exchange of cytochrome c2 between the reaction center-associated and the reaction center-dissociated state.4. The half-reduction potential of cytochrome c2 is pH independent (from pH 5 to 9) whether measured in the free state or when associated with the chromatophore membrane. This shows that a proton is not involved in the oxidation and reduction of cytochrome c2 in the physiological pH range.5. The kinetics of the intact reaction center, P, and cytochrome c2 units in chromatophores and whole cells of Rhodopseudomonas spheroides are described. The two cytochrome c2 molecules which are associated with one P exhibit similar oxidation kinetics; both are biphasic. The fast phase is estimated to be 20–40 μs in half time. The second slower phase is variable depending on the ionic strength of the medium used for the preparation of the chromatophores; it varies from 0.3 to 8 ms.6. An equilibrium for cytochrome c2 and the reaction center and/or the membrane is suggested. The two states of the equilibrium are described by a population of cytochrome c2 functionally “close” to the P+, and a population functionally distant from the P+, which might be physically off the binding site, or orientated unfavorably to the P+. The former population is identified by the 20–40 μs oxidation rate; the latter variable and somewhat slower oxidation (0.3–8 ms) is that whose rate is governed by the diffusional processes of the equilibrium which brings the cytochrome to the close position.7. Carotenoid bandshifts are kinetically compatible (a) with the P oxidation which is too fast to measure, and (b) with the two phases of cytochrome c2 oxidation. These are interpreted as arising from local electric field alterations occurring during the electron transfer events in the reaction center and cytochrome c2.  相似文献   

9.
Soluble cytochrome c-554 (M r 10 kDa) is purified from the green sulfur bacterium Chlorobium tepidum. Its midpoint redox potential is determined to be +148 mV from redox titration at pH 7.0. The kinetics of cytochrome c-554 oxidation by a purified reaction center complex from the same organism were studied by flash absorption spectroscopy at room temperature, and the results indicate that the reaction partner of cytochrome c-554 is cytochrome c-551 bound to the reaction center rather than the primary donor P840. The second-order rate constant for the electron donation from cytochrome c-554 to cytochrome c-551 was estimated to be 1.7×107 M–1 s–1. The reaction rate was not significantly influenced by the ionic strength of the reaction medium.This revised version was published online in October 2005 with corrections to the Cover Date.  相似文献   

10.
The reactions of Rhodopseudomonas viridis cytochrome c2 and horse cytochrome c with Rps. viridis photosynthetic reaction centers were studied by using both single- and double-flash excitation. Single-flash excitation of the reaction centers resulted in rapid photooxidation of cytochrome c-556 in the cytochrome subunit of the reaction center. The photooxidized cytochrome c-556 was subsequently reduced by electron transfer from ferrocytochrome c2 present in the solution. The rate constant for this reaction had a hyperbolic dependence on the concentration of cytochrome c2, consistent with the formation of a complex between cytochrome c2 and the reaction center. The dissociation constant of the complex was estimated to be 30 microM, and the rate of electron transfer within the 1:1 complex was 270 s-1. Double-flash experiments revealed that ferricytochrome c2 dissociated from the reaction center with a rate constant of greater than 100 s-1 and allowed another molecule of ferrocytochrome c2 to react. When both cytochrome c-556 and cytochrome c-559 were photooxidized with a double flash, the rate constant for reduction of both components was the same as that observed for cytochrome c-556 alone. The observed rate constant decreased by a factor of 14 as the ionic strength was increased from 5 mM to 1 M, indicating that electrostatic interactions contributed to binding. Molecular modeling studies revealed a possible cytochrome c2 binding site on the cytochrome subunit of the reaction center involving the negatively charged residues Glu-93, Glu-85, Glu-79, and Glu-67 which surround the heme crevice of cytochrome c-554.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

11.
Transitions in growth irradiance level from 92 to 7 Em-2 s-1 and vice versa caused changes in the pigment contents and photosynthesis of Oscillatoria agardhii. The changes in chlorophyll a and C-phycocyanin contents during the transition from high to low irradiance (HL) were reflected in photosynthetic parameters. In the LH transition light utilization efficiencies of the cells changed faster than pigment contents. This appeared to be related to the lowering of light utilization efficiencies of photosynthesis. As a possible explanation it was hypothesized that excess photosynthate production led to feed back inhibition of photosynthesis. Time-scales of changes in the maximal rate of O2 evolution were discussed as changes in the number of reaction centers of photosystem II in relation to photosynthetic electron transport. Parameters that were subject to change during irradiance transitions obeyed first order kinetics, but hysteresis occurred when comparing HL with LH transients. Interpretation of first order kinetic analysis was discussed in terms of adaptive response vs changes in growth rate.Non-standard abbreviations Chla chlorophyll a - CPC C-phycocyanin - PS II photosystem II - PS I photosystem I - RC II reaction center of photosystem II - P photosynthetic O2-evolution - I irradiance, Em-2 s-1 - light utilization efficiency of cells, mmol O2·mg dry wt-1·h-1/Em-2 s-1 - light utilization efficiency of photosynthetic apparatus, mol O2·mol Chla -1·h-1/Em-2 s-1 - Pmax maximal rate of O2 evolution by cells, mol O2·mg dry wt-1·h-1 - Pmax maximal rate of O2 evolution by photosynthetic apparatus, mol O2·mol·Chla -1·h-1 - LL low light, E m-2 s-1 - HL high light, E m-2 s-1 - LH low to high light transition - HL high to low light transition - k specific rate of adaptation, h-1 - specific growth rate, h-1 - Q pool size of cell constituent, mol·mg dry wt-1 - q net synthesis rate of cell constituent, mol·mg dry wt-1·h-1  相似文献   

12.
《BBA》1987,893(2):232-240
The spectroscopic and thermodynamic properties of the electron-transport components of the photosynthetic bacterium Heliobacterium chlorum were studied by means of absorbance-difference spectroscopy. Upon flash illumination of membranes of H. chlorum photooxidation of the primary electron donor, P-798, was observed. In about 15% of the reaction centers P-798+ was reduced by cytochrome c-553, while in the remaining reaction centers P-798+ reduction occurred via a back reaction with a reduced electron acceptor. Titration experiments indicated a midpoint potential of −440 mV for the electron acceptor. At low redox potentials the formation of the triplet of P-798 was observed after a flash. The triplet was formed in about 30 ns by a back reaction with a reduced electron acceptor and decayed with a time constant of 35 μs. The yield of triplet formed in a flash was 30%. Upon continuous illumination at low redox potentials the accumulation in the reduced state of an electron acceptor was observed. The difference spectrum of this acceptor indicates that it is an iron-sulfur center. The yield of triplet formation was independent of the redox state of the iron-sulfur center, which indicates that the center is not located in the main electron-transport chain. A scheme with three acceptors in the main electron-transport chain is presented to accomodate our results and those of others.  相似文献   

13.
14.
Membrane preparation from the bacteriochlorophyll-containing cells of a facultative methylotroph, Protaminobacter ruber strain NR-1, contained reaction center bacteriochlorophyll similar to those in many species of purple bacteria and contained a few cytochrome species. -Peak of the reduced-minus-oxidized difference spectrum of one of the cytochromes was at 554 nm. The midpoint potential of the cytochrome at pH 7 (Em7) was 350 mV. Two other cytochromes had the same reduced-minus-oxidized difference spectra with a split -band at 557 and 566 nm, but had two different Em7s' of 130 mV and 0 mV.On flash or continuous light the reaction center bacteriochlorophyll and the cytochrome with -peak at 554 nm were reversibly oxidized. Redox titration of the light-induced cytochrome oxidation gave an Em7 value of 356 mV. Under continuous illumination the membrane preparation reversibly took up protons, and formed ATP in the presence of ADP and inorganic phosphate. The ATP formation activity on the bacteriochlorophyll basis was one-third to one-fifth that in chromatophores from Rhodospirillum rubrum under similar experimental conditions. These results clearly indicated that the membrane preparation from P. ruber which contained bacteriochlorophyll had a cyclic photosynthetic electron transfer system and coupled ATP formation activity.Abbreviations Bchl (only in figure legends) bacteriochlorophyll - CCCP carbonylcyanide-m-chlorophenylhydrazone - Eh the ambient redox potential - Em7 the midpoint potential at pH 7 - PMS N-methylphenazonium methosulfate - MES morpholinoethanesulfonic acid - MOPS morpholinopropanesulfonic acid  相似文献   

15.
A photosystem I reaction center has been isolated fromChlamydomonas chloroplasts and compared with the photosystem I reaction center from higher plants. While the higher plant reaction center is active in cytochrome 552 photooxidation, theChlamydomonas preparation was not active unless salts were included in the assay medium or the pH was lowered to 5. Subunit III-depleted photosystem I reaction center from higher plants is also inactive in cytochrome 552 photooxidation in the absence of salts. As with theChlamydomonas reaction center, salts induced its activity. Subunit I of the photosystem I reaction center has tentatively been identified as the binding site of cytochrome 552.  相似文献   

16.
A newly-developed field-portable multi-flash kinetic fluorimeter for measuring the kinetics of the microsecond to millisecond reactions of the oxidizing and reducing sides of photosystem 2 in leaves of intact plants is described and demonstrated. The instrumental technique is a refinement of that employed in the double-flash kinetic fluorimeter (Joliot 1974 Biochim Biophys Acta 357: 439–448) where a low-intensity short-duration light pulse is used to measure the fluorescence yield changes following saturating single-turnover light pulses. The present instrument uses a rapid series of short-duration (2 s) pulses to resolve a complete microsecond to millisecond time-scale kinetic trace of fluorescence yield changes after each actinic flash. Differential optics, using a matrix of optical fibers, allow very high sensitivity (noise levels about 0.05% Fmax) thus eliminating the need for signal averaging, and greatly reducing the intensity of light required to make a measurement. Consequently, the measuring pulses have much less actinic effect and an entire multi-point trace (seven points) excites less than 1% of the reaction centers in a leaf. In addition, bu combining the actinic and measuring pulse light in the optical fiber network, the tail of the actinic flash can be compensated for, allowing measurements of events as rapidly as 20 s after the actinic flash. This resolution makes practical the routine measurement of the microsecond turnover kinetics of the oxygen evolving complex in leaves of intact plants in the field. The instrument is demonstrated by observing flash number dependency and inhibitor sensitivity of the induction and decay kinetics of flash-induced fluorescence transients in leaves of intact plants. From these traces the period-two oscillations associated with the turnover of the two-electron gate and the period-four oscillations associated with the turnover of the oxygen evolving complex can be observed. Applications of the instrument to extending our knowledge of chloroplast function to the whole plant, the effects on plants of environmental stress, herbicides, etc, and possible applications to screening of mutants are discussed.Abbreviations DCMU 3-(3,4-Dichlorophenol)-1,1-dimethylurea - PS 2 photosystem 2 - PS 1 photosystem 1 - P680 primary electron donor of the PS 2 reaction center - QA primary acceptor quinone of PS 2 - QB secondary acceptor quinone of PS 2 - CCCP carbonyl cyanide-m-chlorophenylhydrazone - Yz donor to P680 + - F0 level of fluorescence with all PS 2 centers open - Fmax maximum level of fluorescence with all PS 2 centers closed - P680QA Open reaction centers with P680 reduced and QA oxidized (low fluorescence) - P680QA - Closed reaction centers, in which P680 is reduced (high fluorescence) - P680 +QA - Closed reaction centers, in which P680 is oxidized (low fluorescence)  相似文献   

17.
A minimal kinetic model of the photocycle, including both quinone (Q-6) reduction at the secondary quinone-binding site and (mammalian) cytochrome c oxidation at the cytochrome docking site of isolated reaction centers from photosynthetic purple bacteria Rhodobacter sphaeroides, was elaborated and tested by cytochrome photooxidation under strong continuous illumination. The typical rate of photochemical excitation by a laser diode at 810 nm was 2.200 s-1, and the rates of stationary turnover of the reaction center (one-half of that of cytochrome photooxidation) were 600 +/- 70 s-1 at pH 6 and 400 +/- 50 s-1 at pH 8. The rate of turnover showed strong pH dependence, indicating the contribution of different rate-limiting processes. The kinetic limitation of the photocycle was attributed to the turnover of the cytochrome c binding site (pH < 6), light intensity and quinone/quinol exchange (6 < pH < 8), and proton-coupled second electron transfer in the quinone acceptor complex (pH > 8). The analysis of the double-reciprocal plot of the rate of turnover versus light intensity has proved useful in determining the light-independent (maximum) turnover rate of the reaction center (445 +/- 50 s-1 at pH 7.8).  相似文献   

18.
The protolytic reactions of PSII membrane fragments were analyzed by measurements of absorption changes of the water soluble indicator dye bromocresol purple induced by a train of 10 s flashes in dark-adapted samples. It was found that: a) in the first flash a rapid H+-release takes place followed by a slower H+-uptake. The deprotonation is insensitive to DCMU but is completely eliminated by linolenic acid treatment of the samples; b) the extent of the H+-uptake in the first flash depends on the redox potential of the suspension. In this time domain no H+-uptake is observed in the subsequent flashes; c) the extent of the H+-release as a function of the flash number in the sequence exhibits a characteristic oscillation pattern. Multiphasic release kinetics are observed. The oscillation pattern can be satisfactorily described by a 1, 0, 1, 2 stoichiometry for the redox transitions Si Si+1 (i=0, 1, 2, 3) in the water oxidizing enzyme system Y. The H+-uptake after the first flash is assumed to be a consequence of the very fast reduction of oxidized Q400(Fe3+) formed due to dark incubation with K3[Fe(CN)6]. The possible participation of component Z in the deprotonation reactions at the PSII donor side is discussed.Abbreviations A protonizable group at the PSII acceptor side - BCP Bromocresol Purple - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - FWHM Full Width at Half Maximum - QA, QB primary and secondary plastoquinone at PSII acceptor side - Q400 redox group at PSII-acceptor side (high spin Fe2+) - P680 Photoactive chlorophyll of PSII reaction center - Si redox states of the catalytic site of water oxidation - Z redox component connecting the catalytic site of water oxidation with the reaction center  相似文献   

19.
Many herbicides that inhibit photosynthesis in plants also inhibit photosynthesis in bacteria. We have isolated three mutants of the photosynthetic bacterium Rhodobacter sphaeroides that were selected for increased resistance to the herbicide terbutryne. All three mutants also showed increased resistance to the known electron transfer inhibitor o-phenanthroline. The primary structures of the mutants were determined by recombinant DNA techniques. All mutations were located on the gene coding for the L-subunit resulting in these changes Ile229 Met, Ser223 Pro and Tyr222 Gly. The mutations of Ser223 is analogous to the mutation of Ser264 in the D1 subunit of photosystem II in green plants, strengthening the functional analogy between D1 and the bacterial L-subunit. The changed amino acids of the mutant strains form part of the binding pocket for the secondary quinone, Q b . This is consistent with the idea that the herbicides are competitive inhibitors for the Q b binding site. The reaction centers of the mutants were characterized with respect to electron transfer rates, inhibition constants of terbutryne and o-phenanthroline, and binding constants of the quinone UQ0 and the inhibitors. By correlating these results with the three-dimensional structure obtained from x-ray analysis by Allen et al. (1987a, 1987b), the likely positions of o-phenanthroline and terbutryne were deduced. These correspond to the positions deduced by Michel et al. (1986a) for Rhodopseudomonas viridis.Abbreviations ATP adenosine 5-triphosphate - Bchl bacteriochlorophyll - Bphe bacteriopheophytin - bp basepair - cyt c2+ reduced form of cytochrome c - DEAE diethylami-noethyl - EDTA ethylenediamine tetraacetic acid - Fe2+ non-heme iron atom - LDAO lauryl dimethylamine oxide - Pipes piperazine-N,N-bis-2-ethane-sulfonic acid - PSII photosystem II - RC reaction center - SDS sodium dodecylsulfate - Tris tris(hydroxy-methyl)aminomethane - UQ0 2,3-dimethoxy-5-methyl benzoquinone - UQ10 ubiquinone 50  相似文献   

20.
Chlorosome-depleted membranes and a reaction center complex with well-defined subunit composition were prepared from the green sulfur bacterium Chlorobium vibrioforme under anaerobic conditions. The reaction center complex contains a 15-kDa polypeptide with the N-terminal amino acid sequence MEPQLSRPETASNQVR/. This sequence is nearly identical to the N-terminus of the pscD gene product from Chlorobium limicola (Hager-Braun et al. (1995) Biochemistry 34: 9617–9624). In the presence of ferredoxin and ferredoxin:NADP+ oxidoreductase, the membranes and the isolated reaction center complex photoreduced NADP+ at rates of 333 and 110 mol (mg bacteriochlorophyll a)–1 h–1, respectively. This shows that the isolated reaction center complex contains all the components essential for steady state electron transport. Midpoint potentials at pH 7.0 of 160 mV for cytochrome c 551 and of 245 mV for P840 were determined by redox titration. Antibodies against cytochrome c 551 inhibit NADP+ reduction while antibodies against the bacteriochlorophyll a-binding Fenna-Matthews-Olson protein do not.Abbreviations FMO protein Fenna-Matthews-Olson protein - TMBZ 3,3,5,5-tetramethylbenzidine  相似文献   

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