The photochemical activities and electron carriers of developing barley leaves

The development of photochemical activities in isolated barley plastids during illumination of dark-grown plants has been studied and compared with the behaviour of plastocyanin, cytochromes f, b-559_LP, b-563 and b-559_HP and pigments P546 (C550) and P700. Electron-transport activity dependent on Photosystem 1 and cyclic photophosphorylation dependent on N-methylphenazonium methosulphate (phenazine methosulphate) were very active relative to the chlorophyll content after only a few minutes of illumination of etiolated leaves, and then rapidly declined during the first few hours of greening. By contrast, Photosystem 2 activity (measured with ferricyanide as electron acceptor) and non-cyclic photophosphorylation were not detectable during the first 2½h of greening, but then increased in total amount in parallel with chlorophyll. The behaviour of the electron carriers suggested their association with either Photosystem 1 or 2 respectively. In the first group were plastocyanin, cytochrome f and cytochrome b-563, whose concentrations in the leaf did not change during greening, and cytochrome b-559_LP whose concentration fell to one-half its original value, and in the second group were cytochrome b-559_HP and pigment P546, the concentrations of which closely followed the activities of Photosystem 2. Pigment P700 could not be detected during the first hour, during which time some other form of chlorophyll may take its place in the reaction centre of Photosystem 1. The plastids started to develop grana at about the time that Photosystem 2 activity became detectable.     

Fluorescence emission spectra of Photosystem I,Photosystem II and the light-harvesting chlorophyll a/b complex of higher plants

Fluorescence emission spectra excited at 514 and 633 nm were measured at ?196 °C on dark-grown bean leaves which had been partially greened by a repetitive series of brief xenon flashes. Excitation at 514 nm resulted in a greater relative enrichment of the 730 nm emission band of Photosystem I than was obtained with 633 nm excitation. The difference spectrum between the 514 nm excited fluorescence and the 633 nm excited fluorescence was taken to be representative of a pure Photosystem I emission spectrum at ?196 °C. It was estimated from an extrapolation of low temperature emission spectra taken from a series of flashed leaves of different chlorophyll content that the emission from Photosystem II at 730 nm was 12% of the peak emission at 694 nm. Using this estimate, the pure Photosystem I emission spectrum was subtracted from the measured emission spectrum of a flashed leaf to give an emission spectrum representative of pure Photosystem II fluorescence at ?196 °C. Emission spectra were also measured on flashed leaves which had been illuminated for several hours in continuous light. Appreciable amounts of the light-harvesting chlorophyll a/b protein, which has a low temperature fluorescence emission maximum at 682 nm, accumulate during greening in continuous light. The emission spectra of Photosystem I and Photosystem II were subtracted from the measured emission spectrum of such a leaf to obtain the emission spectrum of the light-harvesting chlorophyll a/b protein at ?196 °C.     

Functional and structural organization of chlorophyll in the developing photosynthetic membranes of Euglena gracilis Z. I. Formation of System II photosynthetic units during greening under optimal light intensity

The relationships between light-harvesting chlorophyll and reaction centers in Photosystem II were analyzed during the chloroplast development of dark-grown, non-dividing Euglena gracilis Z. Comparative measurements included light saturation of photosynthesis, oxygen evolution under flashing-light and fluorescence induction. The results obtained can be summarized as follows: (1) Photosystem II photocenters are formed in parallel with chlorophyll synthesis, but after a longer lag phase. (2) As a consequence, the chlorophyll: reaction center ratio (Emerson's type photosynthetic unit) decreases during greening. (3) This decrease is accompanied by considerable changes in the energy transfer and trapping properties of Photosystem II. Most of the initially synthesized chlorophylls are inactive in the transfer of excitations to active photochemical centers and are shared among newly formed Photosystem II photocenters; as a consequence, the number of chlorophylls functionally connected to each Photosystem II photocenter decreases and cooperativity between these centers appears. Results are discussed in terms of chlorophyll organization in developing photosynthetic membranes with reference to the lake or puddle models of photosynthetic unit organization.     

Energy transfer and the distribution of excitation energy in the photosynthetic apparatus of spinach chloroplasts

Equations are derived from our model of the photochemical apparatus of photosynthesis to show that the yield of energy transfer from Photosystem II to Photosystem I, ?_T(II→Iz), can be obtained from measurements on an individual sample of chloroplasts frozen to ?196 °C by comparing the sum of two specifically defined fluorescence excitation spectra with the absorption spectrum of the sample. Then, given that value of ?_T(II→I), the fraction of the quanta absorbed by the photochemical apparatus which is distributed initially to Photosystem I, α, can be determined as a function of the wavelength of excitation from the same fluorescence excitation spectra. The results obtained in this study of individual samples of chloroplasts frozen to ?196 °C in the absence of divalent cations, namely, that ?_T(II→I) varies from a minimum value of 0.10 when the Photosystem II reaction centers are all open to a maximum value of 0.25 when the centers are all closed and that α has a value of about 0.30 which is almost independent of wavelength for wavelengths shorter than 675 nm (α increases rapidly toward unity at wavelengths longer than 675 nm), agrees quite well with results obtained previously from comparative measurements of chloroplasts frozen to ?196 °C in the presence and absence of divalent cations.     

Energy transfers from Photosystem II to Photosystem I in Cryptomonas rufescens (Cryptophyceae)

In Cryptomonas rufescens (Cryptophyceae), phycoerythrin located in the thylakoid lumen is the major accessory pigment. Oxygen action spectra prove phycoerythrin to be efficient in trapping light energy.The fluorescence excitation spectra at ?196°C obtained by the method of Butler and Kitajima (Butler, W.L. and Kitajima, M. (1975) Biochim. Biophys. Acta 396, 72–85) indicate that like in Rhodophycease, chlorophyll a is the exclusive light-harvesting pigment for Photosystem I.For Photosystem II we can observe two types of antennae: (1) a light-harvesting chlorophyll complex connected to Photosystem II reaction centers, which transfers excitation energy to Photosystem I reaction centers when all the Photosystem II traps are closed. (2) A light-harvesting phycoerythrin complex, which transfers excitation energy exclusively to the Photosystem II reaction complexes responsible for fluorescence at 690 nm.We conclude that in Cryptophyceae, phycoerythrin is an efficient light-harvesting pigment, organized as an antenna connected to Photosystem II centers, antenna situated in the lumen of the thylakoid. However, we cannot afford to exclude that a few parts of phycobilin pigments could be connected to inactive chlorophylls fluorescing at 690 nm.     

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.     

Photoinhibition of Reaction Centers of Photosystems I and II in Intact Bryopsis Chloroplasts under Anaerobic Conditions

Illumination of intact Bryopsis corticulans chloroplasts under anaerobic conditions induced a decline of chlorophyll fluorescence and photoinhibition of Photosystems I and II. The time course of the light-induced decline of chlorophyll fluorescence and the decreases of activities of reactions sensitized by Photosystems I and II were compared. Photosystem I activity decreased in parallel with the disappearance of active P700. The time course of the destruction of the reaction center of Photosystem II was similar to that of photoinhibition of 2,6-dichlorophenolindophenol-Hill reaction.

It appears that the initial events in photoinhibition are the destruction of the reaction centers of Photosystems I and II and that the reaction centers that are inhibited become quenchers of chlorophyll fluorescence.

Effects of inhibitors of electron transfer and of an electron donor to Photosystem I showed that photoinhibition was related to Photosystem I activity.

     

Energy transfer in the photochemical apparatus of flashed bean leaves

Fluorescence and energy transfer properties of bean leaves greened by brief, repetitive xenon flashes were studied at −196 °C. The bleaching of P-700 has no influence on the yield of fluorescence at any wavelength of emission. The light-induced fluorescence yield changes which are observed in both the 690 and 730 nm emission bands in the low temperature fluorescence spectra are due to changes in the state of the Photosystem II reaction centers. The fluorescence yield changes in the 730 nm band are attributed to energy transfer from Photosystem II to Photosystem I. Such energy transfer was also confirmed by measurements of the rate of photooxidation of P-700 at −196 °C in leaves in which the Photosystem II reaction centers were either all open or all closed. It is concluded that energy transfer from Photosystem II to Photosystem I occurs in the flashed bean leaves which lack the light-harvesting chlorophyll a/b protein.     

Development of Photochemical Activity and the Appearance of the High Potential Form of Cytochrome b-559 in Greening Barley Seedlings

The development of photochemical activity during the greening of dark-grown barley seedlings (Hordeum vulgare L. cv. Svalöfs Bonus) was studied in relation to the formation of the high potential form of cytochrome b-559 (cytochrome b-559_HP). Photosynthetic oxygen evolution from leaves was detected at 30 minutes of illumination. The rate of oxygen evolution per gram fresh weight of leaf was as high at 2 to 2.5 hours of greening as at 24 hours or in fully greened leaves. On a chlorophyll basis, the photosynthetic rate at 90 minutes of greening was 80-fold greater than the rate at 45 hours. It is concluded that the majority of photosynthetic units are functional at an early stage of greening, and that chlorophyll synthesis during greening serves to increase the size of the units.     

Development of photochemical activity in relation to pigment and membrane protein accumulation in chloroplasts of barley and its virescens mutant

The development of photochemical activity in relation to pigment and membrane protein accumulation in chloroplasts of greening wild-type barley (Hordeum vulgare L. cv. Gateway) and its virescens mutant were studied. The rate of chlorophyll accumulation per plastid was faster in the wild-type than in the mutant seedlings upon illumination after 6 days of etiolation, but was not different after 8 days. Although the protein content per plastid did not vary during greening, there was a change in the sodium dodecyl sulfate-polyacrylamide gel polypeptide profiles. High molecular weight proteins of 96,000 and 66,000 decreased whereas those at 34,000, 27,000 and 22,000 increased in relative quantity as a function of greening. The fully greened mutant seedlings were not deficient in the light-harvesting chlorophyll protein complex (LHC) or the reaction centers of photosystem I and photosystem II. Photosystem I-associated photochemical activities appeared within the first hour of plastid development and photosystem II associated activities and O₂ evolution within the next 6 hours. In all cases, the developmental rates per unit protein were slower in the mutant following 6 days of etiolation, but no differences between the two genotypes could be seen after 8 days due to a decrease in the developmental rate of the wild-type chloroplasts. An increase in photosynthetic unit size associated with plastid morphogenesis was faster in the wild-type seedlings after 6 days, but again the difference was negligible after 8 days. It was concluded that no single measured photochemical parameter is affected by this mutation, but rather, all aspects of chloroplast development are affected similarly by an overall reduction in the rate of chloroplast morphogenesis. This mutant, therefore, undergoes the normal pattern of proplastid to chloroplast development, but at a markedly reduced rate.     

Greening of etiolated bean leaves in far red light

Eight-day-old dark-grown bean leaves were greened by prolonged irradiation with far red light. Growth, chlorophyll content, oxygen-evolving capacity, photophosphorylation capacity, chloroplast structure (by electron microscopy), and in vivo forms of chlorophyll (by low temperature absorption and derivative spectroscopy on intact leaves) were followed during the greening process. Chlorophyll a accumulated slowly but continuously during the 7 days of the experiment (each day consisted of 12 hours of far red light and 12 hours of darkness). Chlorophyll b was not detected until the 5th day. The capacity for oxygen evolution and photophosphorylation began at about the 2nd day. Electron microscopy showed little formation of grana during the 7 days but rather unfused stacks of primary thylakoids. The thylakoids would fuse to give grana if the leaves were placed subsequently in white light. The low temperature spectroscopy of intact leaves showed that the chlorophyll a was differentiated into three forms with absorption maxima near 670, 677, and 683 nanometers at −196 C during the first few hours and that these forms accumulated throughout the greening process. Small amounts of two longer wavelength forms with maxima near 690 and 698 nanometers appeared at about the same time as photosynthetic activity.     

The efficiency of electron transfer from QA − to the donor side of Photosystem II decreases during induction of photosynthesis: Evidences from chlorophyll fluorescence and photoacoustic techniques

The amplitudes ratio of the fast and slow phases (A_fast/A_slow) in the kinetics of the dark relaxation of variable chlorophyll fluorescence (F_V) was studied after various periods of illumination of dark-adapted primary barley leaves. Simultaneously, photosynthetic activity was monitored using the photoacoustic technique and the photochemical and non-photochemical fluorescence quenching parameters. The ratio A_fast/A_slow changed with the preceding illumination time in a two-step manner. During the first stage of photosynthetic induction (0–20 s of illumination), characterized by a drop in O₂-dependent photoacoustic signal following an initial spike and by a relatively stable small value of photochemical F_V quenching, the ratio A_fast/A_slow remained practically unaltered. During the second stage (20–60 s of illumination), when both the rate of O₂ evolution and the photochemical F_V quenching were found to be sharply developed, a marked increase in the above ratio was also observed. A linear correlation was found between the value of the photochemical quenching and the ratio A_fast/A_slow during the second phase of photosynthetic induction. It is concluded that the slow phase appearing in the kinetics of F_V dark relaxation is not due to the existence of Photosystem II reaction centres lacking the ability to reduce P700⁺ with high rates, but is instead related to the limitation of electron release from Photosystem I during the initial stage of the induction period of photosynthesis. This limitation keeps the intersystem electron carriers in the reduced state and thus increases the probability of back electron transfer from Q_A ^– to the donor side of Photosystem II.Abbreviations A_fast/A_slow the ratio of magnitudes between the fast and slow phases of dark relaxation of variable fluorescence - F_O initial level of chlorophyll fluorescence - F_V variable chlorophyll fluorescence (F-F_O) - (F_V)_S the yield of variable chlorophyll fluorescence under saturating pulse in illuminated leaves - (F_V)_M the yield of variable chlorophyll fluorescence under saturating pulse in dark-adapted leaves - PA photoacoustic - PSI Photosystem I - PS II Photosystem II - qN non-photochemical quenching - qQ photochemical quenching     

Reduced levels of cytochrome b 6/f in transgenic tobacco increases the excitation pressure on Photosystem II without increasing sensitivity to photoinhibition in vivo

We have examined tobacco transformed with an antisense construct against the Rieske-FeS subunit of the cytochromeb ₆ f complex, containing only 15 to 20% of the wild-type level of cytochrome f. The anti-Rieske-FeS leaves had a comparable chlorophyll and Photosystem II reaction center stoichiometry and a comparable carotenoid profile to the wild-type, with differences of less than 10% on a leaf area basis. When exposed to high irradiance, the anti-Rieske-FeS leaves showed a greatly increased closure of Photosystem II and a much reduced capacity to develop non-photochemical quenching compared with wild-type. However, contrary to our expectations, the anti-Rieske-FeS leaves were not more susceptible to photoinhibition than were wild-type leaves. Further, when we regulated the irradiance so that the excitation pressure on photosystem II was equivalent in both the anti-Rieske-FeS and wild-type leaves, the anti-Rieske-FeS leaves experienced much less photoinhibition than wild-type. The evidence from the anti-Rieske-FeS tobacco suggests that rapid photoinactivation of Photosystem II in vivo only occurs when closure of Photosystem II coincides with lumen acidification. These results suggest that the model of photoinhibition in vivo occurring principally because of limitations to electron withdrawal from photosystem II does not explain photoinhibition in these transgenic tobacco leaves, and we need to re-evaluate the twinned concepts of photoinhibition and photoprotection.Abbreviations Chl chlorophyll - DCMU 3-(3,4-dichlophenyl)-1,-dimethylurea - Fo and Fo minimal fluorescence when all PS II reaction centers are open in dark- and light-acclimated leaves, respectively - Fm and Fm maximal fluorescence when all PS II reaction centers are closed in dark- and light-acclimated leaves, respectively - Fv variable fluorescence (Fm-Fo) in dark acclimated leaves - Fv variable fluorescence (Fm-Fo) in lightacclimated leaves - NPQ non-photochemical quenching of fluorescence - PS I and PS II Photosystem I and II - P680 primary electron donor of the reaction center of PS II - PFD photosynthetic flux density - Q_A primary acceptor quinone of PS II - q_p photochemical quenching of fluorescence - V+A+Z violaxanthin+antheraxanthin+zeaxanthin     

Energy transfer between Photosystem II and Photosystem I in chloroplasts

A model for the photochemical apparatus of photosynthesis is presented which accounts for the fluorescence properties of Photosystem II and Photosystem I as well as energy transfer between the two photosystems. The model was tested by measuring at ?196 °C fluorescence induction curves at 690 and 730 nm in the absence and presence of 5 mM MgCl₂ which presumably changes the distribution of excitation energy between the two photosystems. The equations describing the fluorescence properties involve terms for the distribution of absorbed quanta, α, being the fraction distributed to Photosystem I, and β, the fraction to Photosystem II, and a term for the rate constant for energy transfer from Photosystem II to Photosystem I,k_T(II→I). The data, analyzed within the context of the model, permit a direct comparison of α andk_T(II→I) in the absence (?) and presence (+) of Mg²⁺:α/^?α⁺= 1.2andk/^?_T(II→I)k⁺_T(II→I)= 1.9. If the criterion thatα + β = 1 is applied absolute values can be calculated: in the presence of Mg²⁺,a⁺ = 0.27 and the yield of energy transfer,φ⁺_T(II→I) varied from 0.065 when the Photosystem II reaction centers were all open to 0.23 when they were closed. In the absence of Mg²⁺,α^? = 0.32 andφ_T(II→I) varied from 0.12 to 0.28.The data were also analyzed assuming that two types of energy transfer could be distinguished; a transfer from the light-harvseting chlorophyll of Photosystem II to Photosystem I,k_T(II→I), and a transfer from the reaction centers of Photosystem II to Photosystem I,k_t(II→I). In that caseα/^?α⁺= 1.3,k/^?_T(II→I)k⁺_T(II→I)= 1.3 andk/^?_t(II→I)k⁺_(tII→I)= 3.0. It was concluded, however, that both of these types of energy transfer are different manifestations of a single energy transfer process.     

Polypeptides of chloroplastic and cytoplastic origin required for development of Photosystem II activity,and chlorophyll-protein complexes,in Euglena gracilis Z chloroplast membranes

The development of photosynthetic activity and synthesis of chloroplast membrane polypeptides was studied during greening of Euglena gracilis Z in alternate light-dark-light cycles. The results show: (a) The development of both Photosystem II and Photosystem I can be dissociated from chlorophyll synthesis. (b) Most of the polypeptides required for development of Photosystem I are already synthesized during the initial light period (10–12 h); the further rise in Photosystem I activity in the dark is not inhibited by cycloheximide nor by chloramphenicol. (c) The development of Photosystem II requires continuous de novo synthesis of polypeptides and is inhibited by chloramphenicol. The water-splitting activity already present at the end of the first light period decays in the presence of chloramphenicol while that of 1,5-diphenylcarbazide oxidation is only partially retained. The activity can be repaired in the absence of chlorophyll synthesis and is correlated with the de novo synthesis of polypeptides of 50 000–60 000 daltons. The synthesis of these polypeptides and associated repair of Photosystem II activity is not inhibited by cycloheximide. (d) The chloroplast membranes can be resolved into about 40 distinct polypeptides, among them several in the molecular weight range 50 000–60 000, 20 000–35 000 and 10 000–15 000, which are major membrane constitutents. (e) The synthesis of two major polypeptides (M_r = 20 000–30 000) required for the formation of chlorophyll-protein complex(es) containing chlorophyll a and traces of chlorophyll b (CPII?) is light-dependent and cycloheximide-inhibited. It is concluded that the synthesis and addition to the growing membrane of chlorophyll and polypeptides required for the formation of Photosystem II and Photosystem I complexes can be dissociated in time. The H₂O-splitting enzyme(s) and possibly other components of Photosystem II complex are of chloroplastic origin and turn over in the dark while at least some of the chlorophyll binding polypeptides are of cytoplastic origin and their synthesis is light-controlled.     

Inactive Photosystem II Complexes in Leaves : Turnover Rate and Quantitation

The flash-induced electrochromic shift, measured by the amplitude of the rapid absorbance increase at 518 nanometers (ΔA518), was used to determine the amount of charge separation within photosystems II and I in spinach (Spinacia oleracea L.) leaves. The recovery time of the reaction centers was determined by comparing the amplitudes of ΔA518 induced by two flashes separated by a variable time interval. The recovery of the ΔA518 on the second flash revealed that 20% of the reaction centers exhibited a recovery half-time of 1.7 ± 0.3 seconds, which is 1000 times slower than normally active reaction centers. Measurements using isolated thylakoid membranes showed that photosystem I constituted 38% of the total number of reaction centers, and that the photosystem I reaction centers were nearly fully active, indicating that the slowly turning over reaction centers were due solely to photosystem II. The results demonstrate that in spinach leaves approximately 32% of the photosystem II complexes are effectively inactive, in that their contribution to energy conversion is negligible. Additional evidence for inactive photosystem II complexes in spinach leaves was provided by fluorescence induction measurements, used to monitor the oxidation kinetics of the primary quinone acceptor of photosystem II, Q_A, after a short flash. The measurements showed that in a fraction of the photosystem II complexes the oxidation of Q_A⁻ was slow, displaying a half-time of 1.5 ± 0.3 seconds. The kinetics of Q_A⁻ oxidation were virtually identical to the kinetics of the recovery of photosystem II determined from the electrochromic shift. The key difference between active and inactive photosystem II centers is that in the inactive centers the oxidation rate of Q_A⁻ is slow compared to active centers. Measurements of the electrochromic shift in detached leaves from several different species of plants revealed a significant fraction of slowly turning over reaction centers, raising the possibility that reaction centers that are inefficient in energy conversion may be a common feature in plants.     

Development of Photosystem I and Photosystem II Activities in Leaves of Light-grown Maize (Zea mays)

To compare chloroplast development in a normally grown plant with etiochloroplast development, green maize plants (Zea mays), grown under a diurnal light regime (16-hour day) were harvested 7 days after sowing and chloroplast biogenesis within the leaf tissue was examined. Determination of total chlorophyll content, ratio of chlorophyll a to chlorophyll b, and O₂-evolving capacity were made for intact leaf tissue. Plastids at different stages of development were isolated and the electron-transporting capacities of photosystem I and photosystem II measured. Light saturation curves were produced for O₂-evolving capacity of intact leaf tissue and for photosystem I and photosystem II activities of isolated plastids. Structural studies were also made on the developing plastids. The results indicate that the light-harvesting apparatus becomes increasingly efficient during plastid development due to an increase in the photosynthetic unit size. Photosystem I development is completed before that of photosystem II. Increases in O₂-evolving capacity during plastid development can be correlated with increased thylakoid fusion. The pattern of photosynthetic membrane development in the light-grown maize plastids is similar to that found in greening etiochloroplasts.     

Functional and structural organization of chlorophyll in the developing photosynthetic membranes of Euglena gracilis Z. II. Formation of system II photosynthetic units during greening under optimal light intensity.

The relationships between light-harvesting chlorophyll and reaction centers in Photosystem II were analyzed during the chloroplast development of dark-grown, non-dividing Euglena gracilis Z. Comparative measurements included light saturation of photosynthesis, oxygen evolution under flashing-light and fluorescence induction. The results obtained can be summarized as follows: (1) Photosystem II photocenters are formed in parallel with chlorophyll synthesis, but after a long lag phase. (2) As a consequence, the chlorophyll reaction center ratio (Emerson's type photosynthetic unit) decreases during greening. (3) This decrease is accompanied by considerable changes in the energy transfer and trapping properties of Photosystem II. Most of the initially synthesized chlorophylls are inactive in the transfer of excitations to active photochemical centers and are shared among newly formed Photosystem II photocenters; as a consequence, the number of chlorophylls functionally connected to each Photosystem II photocenter decreases and cooperatively between these centers appears. Results are discussed in terms of chlorophyll organization in developing photosynthetic membranes with reference to the lake or puddle models of photosynthetic unit organization.     

Characterization of a non-detergent PS II-cytochrome b/f preparation (BS)

A non-detergent photosystem II preparation, named BS, has been characterized by countercurrent distribution, light saturation curves, absorption spectra and fluorescence at room and at low temperature (–196°C). The BS fraction is prepared by a sonication-phase partitioning procedure (Svensson P and Albertsson P-Å, Photosynth Res 20: 249–259, 1989) which removes the stroma lamellae and the margins from the grana and leaves the appressed partition region intact in the form of vesicles. These are closed structures of inside-out conformation. They have a chlorophyll a/b ratio of 1.8–2.0, have a high oxygen evolving capacity (295 mol O₂ per mg chl h), are depleted in P700 and enriched in the cytochrome b/f complex. They have about 2 Photosystem II reaction centers per 1 cytochrome b/f complex.The plastoquinone pool available for PS II in the BS vesicles is 6–7 quinones per reaction center, about the same as for the whole thylakoid. It is concluded, therefore, that the plastoquinone of the stroma lamellae is not available to the PS II in the grana and that plastoquinone does not act as a long range electron transport shuttler between the grana and stroma lamellae.Compared with Photosystem II particles prepared by detergent (Triton X-100) treatment, the BS vesicles retain more cytochrome b/f complex and are more homogenous in their surface properties, as revealed by countercurrent distribution, and they have a more efficient energy transfer from the antenna pigments to the reaction center.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - F_v variable fluorescence - LHC light-harvesting complex - PpBQ phenyl-p-benzoquinone - PQ plastoquinone pool - P700 reaction center of PS I - PS I, PS II Photosystem I, II - Q_A first bound plastoquinone accepter - RC reaction centre     

Development of the Two Heterogeneous Photosystem II Units in Etiolated Bean Leaves

The development of the photosystem II units in relation to the heterogeneity of their photochemical centers was studied in etiolated bean leaves (Phaseolus vulgaris var. red kidney) greened under continuous or intermittent light. The study was done in order to see whether grana are the loci of the units with the efficient photosystem II activity (α units), while the stroma thylakoids are the loci of the units with the less efficient photosystem II activity (β units), as it has been proposed. In addition, the interrelations between α and β centers have been investigated. It was found that the α and the β centers of photosystem II were present in the first photosynthetic membranes irrespective of the mode of greening of the leaves. The magnitude of their respective photochemical rate constants, K′_α and K_β, increased with time in continuous light and it reached the steady-state values of the mature chloroplasts within 16 hours, while in intermittent light it remained smaller. The differentiation of the system II units in α and β centers containing units is more evident under conditions of intermittent illumination, i.e. when the rate of chlorophyll biosynthesis is the limiting step for chloroplast development.