Tetranitromethane modification of photosystem 2

Inhibition of photosystem 2 by the peptide-modification reagent, tetranitromethane, has been investigated with spinach digitonin particles. In the presence of tetranitromethane, (1) the initial fluoresence yield is suppressed with a concomitant elimination of the variable component of fluorescence; (2) the optical absorption transient at 820 nm, attributed to P680⁺, is greatly attenuated; (3) diphenylcarbazide-supported photoreduction of dichlorophenol indophenol is abolished; and (4) electron spin resonance Signal 2f and Signal 2s are eliminated. These results are consistent with multiple sites of modification in photosystem 2 by tetranitromethane, and suggest further that this reagent can inhibit charge stabilization in the reaction center.Abbreviations D₁ electron donor to P680⁺ in oxygen-inhibited photosystem 2 preparations - DPIP 2,6-dichlorophenol indophenol - esr electron spin resonance - F_i initial chlorophyll a fluorescence yield - F_max maximum chlorophyll a fluorescence yield - F_v variable chlorophyll a fluorescence yield - FWHM full width at half maximum - Mes 2-(N-morpholino)ethanesulfonic acid - P680 primary electron donor chlorophyll of photosystem 2 - Ph pheophytin - PS 2-photosystem 2 - Q_a primary quinone electron acceptor - Q_b secondary quinone acceptor - Tricine N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine - TNM tetranitromethane     

The photochemical and fluorescence properties of whole cells,spheroplasts and spheroplast particles from the blue-green alga Phormidium luridum

The photochemical activities and fluorescence properties of cells, spheroplasts and spheroplast particles from the blue-green alga Phormidium luridum were compared. The photochemical activities were measured in a whole range of wavelengths and expressed as quantum yield spectra (quantum yield vs. wavelength). The following reactions were measured: Photosynthesis (O₂ evolution) in whole cells; Hill reaction (O₂ evolution) with Fe(CN)₆^3? and NADP as electron acceptors (Photosystem II and Photosystem II+Photosystem I reactions); electron transfer from reduced 2,6-dichlorophenolindophenol to diquat (Photosystem I reaction). The fluorescence properties were emission spectra, quantum yield spectra and the induction pattern.On the basis of comparison between the quantum yield spectra and the pigments compositions the relative contribution of each pigment to each photosystem was estimated. In normal cells and spheroplasts it was found that Photosystem I (Photosystem II) contains about 90 % (10 %) of the chlorophyll a, 90 % (10 %) of the carotenoids and 15 % (85 %) of the phycocyanin. In spheroplast particles there is a reorganization of the pigments: they loose a certain fraction (about half) of the phycocyanin but the remaining phycocyanin attaches itself exclusively to Photosystem I (!). This is reflected by the loss of Photosystem II activity, a flat quantum yield vs. wavelength dependence and a loss of the fluorescence induction.The fluorescence quantum yield spectra conform qualitatively to the above conclusion. More quantitative estimation shows that only a fraction (20–40 %) of the chlorophyll of Photosystem II is fluorescent. Total emission spectrum and the ratio of variable to constant fluorescence are in agreement with this conclusion.The fluorescence emission spectrum shows characteristic differences between the constant and variable components. The variable fluorescence comes exclusively from chlorophyll a; the constant fluorescence is contributed, in addition to chlorophyll a, by phycocyanine and an unidentified long wavelength component.The variable fluorescence does not change in the transition from whole cells to spheroplasts. However, the constant fluorescence increases considerably. This indicates the release of a small fraction of pigments from the photosynthetic photochemical apparatus which then become fluorescent.     

Zinc-inhibited Electron Transport of Photosynthesis in Isolated Barley Chloroplasts

In isolated barley chloroplasts, the presence of 2 millimolar ZnSO₄ inhibits the electron transport activity of photosystem II, as measured by photoreduction of dichlorophenolindophenol, O₂ evolution, and chlorophyll a fluorescence. The inhibition of photosystem II activity can be restored by the addition of the electron donor hydroxylamine or diphenylcarbazide, but not by benzidine and MnCl₂. These observations suggest that Zn inhibits electron flow at the oxidizing side of photosystem II at a site prior to the electron donating site(s) of hydroxylamine and diphenylcarbazide. No inhibition of photosystem I-dependent electron transport by 3 millimolar ZnSO₄ is observed. However, with concentrations of ZnSO₄ above 5 millimolar, photosystem I activity is partially inactivated. Washing Zn²⁺-treated chloroplasts partially restores the O₂-evolving activity.     

Electron transport to oxygen mitigates against the photoinactivation of Photosystem II in vivo

The role of electron transport to O₂ in mitigating against photoinactivation of Photosystem (PS) II was investigated in leaves of pea (Pisum sativum L.) grown in moderate light (250 mol m^–2 s^–1). During short-term illumination, the electron flux at PS II and non-radiative dissipation of absorbed quanta, calculated from chlorophyll fluorescence quenching, increased with increasing O₂ concentration at each light regime tested. The photoinactivation of PS II in pea leaves was monitored by the oxygen yield per repetitive flash as a function of photon exposure (mol photons m^–2). The number of functional PS II complexes decreased nonlinearly with increasing photon exposure, with greater photoinactivation of PS II at a lower O₂ concentration. The results suggest that electron transport to O₂, via the twin processes of oxygenase photorespiration and the Mehler reaction, mitigates against the photoinactivation of PS II in vivo, through both utilization of photons in electron transport and increased nonradiative dissipation of excitation. Photoprotection via electron transport to O₂ in vivo is a useful addition to the large extent of photoprotection mediated by carbon-assimilatory electron transport in 1.1% CO₂ alone.Abbreviations Fm, Fo, Fv- maximal, initial (corresponding to open PS II traps) and variable chlorophyll fluorescence yield, respectively - NPQ- non-photochemical quenching - PS- photosystem - Q_A- primary quinone acceptor - qP- photochemical quenching coefficient     

Photochemical properties of mesophyll and bundle sheath chloroplasts of maize

Several photochemical and spectral properties of maize (Zea mays) bundle sheath and mesophyll chloroplasts are reported that provide a better understanding of the photosynthetic apparatus of C₄ plants. The difference absorption spectrum at 298 K and the fluorescence excitation and emission spectra of chlorophyll at 298 K and 77 K provide new information on the different forms of chlorophyll a in bundle sheath and mesophyll chloroplasts: the former contain, relative to short wavelength chlorophyll a forms, more long wavelength chlorophyll a form (e.g. chlorophyll a 693 and chlorophyll a 705) and less chlorophyll b than the latter. The degree of polarization of chlorophyll a fluorescence is 6% in bundle sheath and 4% in mesophyll chloroplasts. This result is consistent with the presence of relatively high amounts of oriented long wavelength forms of chlorophyll a in bundle sheath compared to mesophyll chloroplasts. The relative yield of variable, with respect to constant, chorophyll a fluorescence in mesophyll chloroplasts is more than twice that in bundle sheath chloroplast. Furthermore, the relative yield of total chlorophyll a fluorescence is 40% lower in bundle sheath compared to that in mesophyll chloroplasts. This is in agreement with the presence of the higher ratio of the weakly fluorescent pigment system I to pigment system II in bundle sheath than in mesophyll chloroplast. The efficiency of energy transfer from chlorophyll b and carotenoids to chlorophyll a are calculated to be 100 and 50%, respectively, in both types of chloroplasts. Fluorescence quenching of atebrin, reflecting high energy state of chloroplasts, is 10 times higher in mesophyll chloroplasts than in bundle sheath chloroplasts during noncyclic electron flow but is equal during cyclic flow. The entire electron transport chain is shown to be present in both types of chloroplasts, as inferred from the antagonistic effect of red (650 nm) and far red (710 nm) lights on the absorbance changes at 559 nm and 553 nm, and the photoreduction of methyl viologen from H₂O. (The rate of methyl viologen photoreduction in bundle sheath chloroplasts was 40% of that of mesophyll chloroplasts.)     

Inhibition of photosynthesis in current-year needles of unfertilized and fertilized Norway spruce [Picea abies (L.) Karst.] during autumn and early winter

Inhibition of photosynthesis was followed during autumn and early winter in current-year sun and shade needles of unfertilized and fertilized Norway spruce [Picea abies (L.) Karst.] by simultaneous measurements of photosynthetic O₂ evolution and chlorophyll a fluorescence at 20 °C. The CO₂-saturated rate of O₂ evolution was generally higher in sun needles of fertilized trees than in those of unfertilized trees over a wide range of incident photon flux densities (PFDs). Furthermore, the maximum photo-chemical efficiency of photosystem (PS) II, as indicated by the ratio of variable to maximum fluorescence (F_V/F_M) was generally higher for sun needles of fertilized trees. The depression of f_v/f_m during frost periods was more pronounced in sun needles than in shade needles, indicating that winter inhibition in Norway spruce is strongly light-dependent. However, the inhibition of the rate of O₂ evolution at high PFDs in needles of fertilized trees during early winter was partly independent of the light regime experienced by those needles in the field, which appeared to result in a pronounced decrease in the proportion of oxidized PS II reaction centres in shade needles. A nearly identical linear relationship between the quantum yield of PS II electron transport determined by chlorophyll fluorescence and the quantum yield of O₂ evolution (gross rate of O₂ evolution/PFD) was obtained for the investigated types of needles during autumn and early winter. Except for shade needles of fertilized trees, this appeared to be largely achieved by adjustments in thermal energy dissipation within PS II.     

Development of the Primary Photochemical Apparatus of Photosynthesis during Greening of Etiolated Bean Leaves

Seven-day-old dark-grown bean leaves were greened under continuous light. The amount of chlorophyll, the ratio of chlorophyll a to chlorophyll b, the O₂ evolving capacity and the primary photochemical activities of Photosystem I and Photosystem II were measured on the leaves after various times of greening. The primary photochemical activities were measured as the photo-oxidation of P₇₀₀, the photoreduction of C-550, and the photo-oxidation of cytochrome b₅₅₉ in intact leaves frozen to −196 C. The results indicate that the reaction centers of Photosystem I and Photosystem II begin to appear within the first few minutes and that Photosystem II reaction centers accumulate more rapidly than Photosystem I reaction centers during the first few hours of greening. The very early appearances of the primary photochemical activity of Photosystem II was also confirmed by light-induced fluorescence yield measurements at −196 C.     

Analysis of chlorophyll fluorescence induction kinetics exhibited by DCMU-inhibited thylakoids and the origin of α and β centres

Analyses of room-temperature chlorophyll fluorescence curves from DCMU-inhibited thylakoids were used to investigate the proposed PS II structural heterogeneity of α and β centres. The kinetics of the area growth curves, representative of Q_A photoreduction, could be modified in the presence of DCMU by exogenous electron acceptors and by added reductants of the PQ pool. The effect of altered DCMU levels (range 0.2–100 μM) on the induction curve kinetics was to modify preferentially the slow-β component, while having only a very small effect on the total variable fluorescence yield. Over the DCMU concentration range used, the unnormalized area of the induction curve (A_max) decreased with increasing herbicide concentration by approx. 45%, indicating that less quanta were required to reduce Q_A. It was found that the dark reoxidation of Q_A in the presence of DCMU and Ant 2p after a light pretreatment regenerated the slow kinetic component. When chlorophyll fluorescence emission at 685 and 731 nm was measured, no difference was observed in the kinetics of the induction curve. The analysis of PS II-enriched, oxygen-evolving membranes indicated the presence of both the fast and slow kinetic components, although this type of preparation showed a modified fast phase. The above observations led to the conclusion that several of the previously proposed characteristics of PS II_α and PS II_β centres do not hold and that a type of PS II heterogeneity involving different degrees of DCMU inhibition is sufficient to explain many of the observations made.     

Cobalt induced changes in photosystem activity in Synechocystis PCC 6803: Alterations in energy distribution and stoichiometry

Adaptive responses to excess (supraoptimal) level of cobalt supplied to the growth medium were studied in the cyanobacterium Synechocystis PCC 6803. Growth of cells in the medium containing 10 M CoCl₂ led to a large stimulation (50%) in O₂-evolution and an overall increase (30%) in the photosynthetic electron transport rates. Analysis of variable Chl a fluorescence yield of PS II and immuno-detection of Photosystem II (PS II) reaction-center protein D1, showed a small increase (15–20%) in the number of PS II units in cobalt-grown cells. Cobalt-grown cells, therefore, had a slightly elevated PS II/PS I ratio compared to control.We observed alteration in the extent of energy distribution between the two photosystems in the eobalt grown cells. Energy was preferentially distributed in favour of PS II accompanied by a reduction in the extent of energy transfer from PS II to PS I in cobalt-grown cells. These cells also showed a smaller PS I absorption cross-section and a smaller size of intersystem electron pool than the control cells. Thus, our results suggest that supplementation of 10 M CoCl₂, to the normal growth medium causes multiple changes involving small increase in PS II to PS I ratio, enhanced funneling of energy to PS II and an increase in PS I electron transport, decrease PS I cross section and reduction in intersystem pool size. The cumulative effects of these alterations cause stimulation in electron transport and O₂ evolution.Abbreviations BCIP 5-bromo-4-chloro-3-indolylphosphate - Chl a Chlorophyll a - Cyt blf Cytochrome blf - DCBQ 2,6-dichlorobenzoquinone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - DCPIP 2,6-dichlorophenol indophenol - DPC Diphenyl carbazide - F_o fluorescence when all reaction centers are open - F_M fluorescence yield when all reaction centers are closed - F_v variable chlorophyll fluorescence - HEPES N-2-hydroxyethyl piperazine-N'-2-ethanesulphonic acid - MV methyl viologen - NBT nitro-blue tetrazolium - pBQ para-benzoquinone - PB somes phycobilisomes - PC Phycocyanin - PQ plastoquinone - PS I Photosystem I - PS II Photosystem II - P700 reaction center Chl a of PS 1 - ST-and MT-flash single turnover and multiple turnover flash     

Reversible inhibition of photochemistry of photosystem II by Ca2+ removal from intact cells of Anacystis nidulans

Depletion of Ca²⁺ from Anacystis nidulans produces an inhibition of O₂ evolution that is accompanied both at 39°C and 77 K by a loss of chlorophyll fluorescence of variable yield. This indicates that Ca²⁺-depletion causes disruption of normal photosystem II function, manifested by the disappearance of photoreduction of Q. Delayed light emission in the ms time range is also eliminated in Ca²⁺-depleted cells, which confirms that Ca²⁺ removal prevents charge separation and recombination in reaction centers of photosystem II. Readdition of Ca²⁺ to depleted cells restores fully the fluorescence of variable yield and delayed light emission, as well as O₂ evolution. Thus, Ca²⁺ may be a required component for photosystem II in A. nidulans.     

Delayed fluorescence observed in the nanosecond time region at 77 K originates directly from the photosystem II reaction center

The excited-state dynamics of delayed fluorescence in photosystem (PS) II at 77 K were studied by time-resolved fluorescence spectroscopy and decay analysis on three samples with different antenna sizes: PS II particles and the PS II reaction center from spinach, and the PS II core complexes from Synechocystis sp. PCC 6803. Delayed fluorescence in the nanosecond time region originated from the 683-nm component in all three samples, even though a slight variation in lifetimes was detected from 15 to 25 ns. The relative amplitude of the delayed fluorescence was higher when the antenna size was smaller. Energy transfer from the 683-nm pigment responsible for delayed fluorescence to antenna pigment(s) at a lower energy level was not observed in any of the samples examined. This indicated that the excited state generated by charge recombination was not shared with antenna pigments under the low-temperature condition, and that delayed fluorescence originates directly from the PS II reaction center, either from chlorophyll a_D1 or P680. Supplemental data on delayed fluorescence from spinach PS I complexes are included.     

Excitation-energy redistribution in the cryptomonad alga Cryptomonas ovata

We have studied the redistribution of excitation energy in the cryptomonad alga Cryptomonas ovata. Low-temperature fluorescence emission spectra from cells preilluminated with light 1 and light 2 show that preferential excitation of Photosystem II (PS II) leads to decreased fluorescence emission from chlorophyll (Chl) a associated with PS II relative to the emission following the preferential excitation of Photosystem I (PS I). The fluorescence change is indicative of a light-state transition by the cells. However, comparision of measurements of the kinetics of P-700 photooxidation by cells fixed with glutaraldehyde following illumination with light 1 or light 2 shows that the relative activity of PS I is lower in cells fixed in light 2. This is in contrast to the expectation for cells in State 2. Excitation spectra for the fluorescence emission from PS II Chl a show that preferential excitation of PS II leads to a decreased probability for energy transfer from phycoerythrin and Chl c₂ to PS II when compared to cells in which PS I is preferentially excited. This result is in accordance with recent picosecond time-resolved fluorescence studies (Bruce, D., Biggins, J., Charbonneau, S. and Thewalt, M. (1987) in Progress in Photosynthesis Research (Biggins, J., ed.), Vol. II, pp. 777–780, Martinus Nijhoff, Dordrecht) and we, therefore, suggest that C. ovata does not undergo a classical light-state transition. However, preferential excitation of PS II or PS I appears to cause pigment-protein conformational changes which change the probability for energy transfer from phycoerythrin to PS II, and we suggest that this may be a mechanism for photoprotection of PS II. Studies of the kinetics of excitation-energy redistribution, and of the effects of electron-transport inhibitors and uncouplers of photophosphorylation indicate that the mechanism for excitation-energy redistribution in C. ovata and phycobilisome-containing organisms may be similar.     

Active Transport of Inorganic Carbon Increases the Rate of O(2) Photoreduction by the Cyanobacterium Synechococcus UTEX 625

Chlorophyll a fluorescence of Synechococcus UTEX 625 was quenched during the transport of inorganic carbon, even when CO₂ fixation was inhibited by iodoacetamide. Measurements with a pulse modulation fluorometer showed that at least 75% of the quenching was due to oxidation of Q_a, the primary acceptor of photosystem II. Mass spectrometry revealed that transport of inorganic carbon increased the rate of O₂ photoreduction. Hence, O₂ could serve as an electron acceptor to allow oxidation of Q_a even in the absence of CO₂ fixation.     

Photoinactivation of the reactivation capacity of photosystem II in pea subchloroplast particles after a complete removal of manganese

After a complete removal of Mn from pea subchloroplast photosystem-II (PS II) preparations the electron phototransfer and oxygen evolution are restored upon addition of Mn²⁺ and Ca²⁺. Pre-illumination of the sample in the absence of Mn²⁺ leads to photoinhibition (PI) — irreversible loss of the capability of PS II to be reactivated by Mn²⁺. The effect of PI is considerably decreased in the presence of Mn²⁺ (4 Mn atoms per reaction center of PS II) and it is increased in the presence of ferricyanide or p-benzoquinone revealing the oxidative nature of the photoeffect. PI results in suppression of oxygen evolution, variable fluorescence, photoreduction of 2,6-dichlorophenol indophenol from either water or diphenylcarbazide. However, photooxidation of chlorophyll P₆₈₀, the primary electron donor of PS II as well as dark and photoinduced EPR signal II (ascribed to secondary electron donors D ₁ and Z) are preserved. PI is accompanied by photooxidation of 2–3 carotenoid molecules per PS II reaction center (RC) that is accelerated in the presence of ferricyanide and is inhibited upon addition of Mn²⁺ or diuron. The conclusion is made that PI in the absence of Mn leads to irreversible oxidative inactivation of electron transfer from water to RC of PS II which remains photochemically active. A loss of functional interaction of RC with the electron transport chain as a common feature for different types of PS II photoinhibition is discussed.Abbreviations A photoinduced absorbance changes - DPC diphenylcarbazide - DPIP 2,6-dichlorophenol indophenol - F _o constant fluorescence of chlorophyll - F photoinduced changes of Chl fluorescence yield - Mn manganese - P₆₈₀ the primary electron donor in PS II - PI photoinhibition - PS II photosystem II - Q the primary (quinone) electron acceptor in PS II - RC reaction center     

Effects of inorganic carbon accumulation on photosynthetic oxygen reduction and cyclic electron flow in the cyanobacterium Synechococcus PCC7942

This paper examines the effect of inorganic carbon transport and accumulation in Synechococcus PCC7942 on fluorescence quenching, photosynthetic oxygen reduction and both linear and cyclic electron flow. The data presented support the previous findings of Miller et al. (1991) that the accumulation of Ci by the CO₂ concentrating mechanism is able to stimulate oxygen photoreduction, particularly so when CO₂ fixation is inhibited by PCR cycle inhibitors such as glycolaldehyde. This effect is found with both high and low-Ci grown cells, but the potential for oxygen photoreduction is about two-fold higher in low-Ci grown cells. This greater potential for O₂ photoreduction is also correlated with a higher ability of low-Ci cells to photoreduce H₂O₂. Experiments with a mutant which transports Ci but does not accumulate it internally, indicates that the stimulation of O₂ photoreduction appears to be a direct effect of the internal accumulation of Ci rather than from its participation in the transport process. In the absence of Ci, no specific partial reactions of photosynthetic electron transport appear to be inhibited, and the PS 1 acceptors PNDA and MV as well as the PS 2 acceptor DMQ can all run electron transport at levels approaching those during active CO₂ fixation. Measurements of P700⁺ show that when the cells are depleted of Ci during photosynthesis, P700 becomes more oxidised. This indicates that the resupply of electrons from the intersystem chain is relatively more restricted under conditions of Ci limitation than is the availability of PS 1 electron acceptors. It is proposed that the accumulated Ci pool can directly stimulate the ability of O₂ to act as a PS 1 acceptor and that the ability of PS 1 acceptors, such as O₂, to relieve restrictions on intersystem electron transfer is perhaps a result of a reduction in cyclic electron flow and a subsequent increase in the oxidation state of the plastoquinone pool.Abbreviations BTP 1,3-bis[tris(hydroxymethyl)-methylaminopropane] - CA carbonic anhydrase' - Ci inorganic carbon (CO₂+HCO₃ ^–+CO₃ ^2–) - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DMQ 2,6-dimethylbenzoquinone - EZ ethoxyzolamide or 6-ethoxy-2-benzothiazole-sulfonamide - FCCP carbonyl cyanide p-trifluoro methoxyphenyl-hydrazone - F steady-state chlorophyll fluorescence - F_m chlorophyll fluorescence during a saturating light pulse - F_o chlorophyll fluorescence in the dark, prior to illumination by actinic light - MV methyl viologen or 1,1-dimethyl-4,4-bipyridinium dichloride - PCR cycle photosynthetic carbon reduction cycle - PNDA N,N-dimethyl-p-nitrosoaniline - _{PS 1} the quantum yield of Photosystem 1 - _{PS 2} the quantum yield of Photosystem 2     

Novel effects of methyl viologen on photosystem II function in spinach leaves

Methyl viologen (MV) is a well-known electron mediator that works on the acceptor side of photosystem I. We investigated the little-known, MV-induced inhibition of linear electron flow through photosystem II (PS II) in spinach-leaf discs. Even a low [MV] decreased the (1) average, light-adapted photochemical efficiency of PS II traps, (2) oxidation state of the primary quinone acceptor Q_A in PS II during illumination, (3) photochemical efficiency of light-adapted open PS II traps, (4) fraction of absorbed light energy dissipated constitutively in a light-independent manner or as chlorophyll (Chl) a fluorescence emission, (5) Chl a fluorescence yield corresponding to dark-adapted open reaction-center traps (F _o) and closed reaction-center traps (F _m), and (6) half-time for re-oxidation of Q_A⁻ in PS II after a single-turnover flash. These effects suggest that the presence of MV accelerates various “downhill” electron-transfer steps in PS II. Therefore, when using the MV to quantify cyclic electron flow, the inhibitory effect of MV on PS II should be taken into account.     

Period four oscillations in chlorophyll a fluorescence

The discovery of period four oscillations of the fluorescence yield under flashing light demonstrated that not only the redox state of the Photosystem II (PS II) electron acceptor Q_A, but also the oxygen evolving cycle (described by the S states) modulates the fluorescence yield of chlorophyll (Chl). The positive charges accumulated on the donor side of PS II act on the fluorescence yield (measured in the Q_A ⁻ state during a strong flash) through the concentration of the quencher P₆₈₀ ⁺, the oxidized form of PS II reaction center Chl a. However, the period four oscillations of the fluorescence yield detected 1 s after a strong flash (in the P₆₈₀Q_A state) have not yet been fully explained. This revised version was published online in June 2006 with corrections to the Cover Date.     

Photoreduction of pheophytin as a result of electron donation from the water-splitting system to Photosystem-II reaction centers

Reversible photoreduction of pheophytin (Pheo) accompanied by a decrease of chlorophyll-fluorescence yield is observed in subchloroplast oxygen-evolving preparations of Photosystem II (PS II) under anaerobic conditions. This photoreaction is activated at addition of CCCP, inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and reactivated upon subsequent addition of ascorbate. Benzyl viologen as well as methyl viologen accelerates dark oxidation of reduced pheophytin, indicating that they are able to accept an electron from Pheo^⨪. The data on both the photoreduction of pheophytin in the absence of exogenous reductants - when electron donation to reaction centers of PS II occurs only from water - and the inhibition of this photoreaction by DCMU show that the pheophytin photoreduction is sensitized by reaction centers of PS II, and it probably occurs as a result of electron donation from the water-splitting system being in the sate S₃ to P^⨥-680Pheo^⨪Q⁻, producing the long-lived state S₀ P-680Pheo^⨪Q⁻ and O₂. Photoreduction of pheophytin in the presence of ascorbate (and dithionite) evidently occurs as a result of donation of its electrons to P^⨥-680Pheo^⨪Q⁻ by means of the S-states of the water-oxidizing system. It is shown that the photoinduced decrease of fluorescence in chloroplasts under anaerobic conditions is due to two processes: photoreduction of pheophytin in Photosystem II and photooxidation of Q⁻ by Photosystem I. It is suggested that photoreduction of pheophytin takes also place under aerobic conditions when Q is reduced. It may contribute to the P−S fluorescence decrease during fluorescence induction in leaves.     

Cobalt ions inhibit electron-transport activity of Photosystem II without affecting Photosystem I

Investigations on photosynthesis have greatly benefited by the use of specific inhibitors that affect a specific site of inhibition on the electron-transport chain. We show here for the first time that cobalt (Co²⁺) ions can be used specifically to inactivate electron donation to the reaction centre of Photosystem (PS) II without affecting PS I reactions. This conclusion is based on the following observations: (1) addition of exogenous electron donors such as NH₂OH does not relieve Co²⁺-induced inactivation of photoelectron transport or the lowering of steady-state chlorophyll a fluorescence yield; this suggests that the inhibition is beyond the NH₂OH donation site and before the fluorescence quencher Q, i.e., on the reaction centre complex itself. (2) Washing of Co²⁺-pretreated chloroplasts with isolation buffer to remove Co²⁺ does not relieve Co²⁺-induced inhibition of Hill activity, suggesting that the Co²⁺ effect is irreversible. (3) Co²⁺ did not alter the PS I reactions. Thus, Co²⁺-treated chloroplasts can be used to study PS I functions free from PS II reactions in isolated chloroplasts.