Structural and functional organization of the photosystems in spinach chloroplasts. Antenna size,relative electron-transport capacity,and chlorophyll composition

The structural and functional organization of the spinach chloroplast photosystems (PS) I, II_α and II_β was investigated. Sensitive absorbance difference spectrophotometry in the ultraviolet (?A₃₂₀) and red (?A₇₀₀) regions of the spectrum provided information on the relative concentration of PS II and PS I reaction centers. The kinetic analysis of PS II and PS I photochemistry under continuous weak excitation provided information on the number (N) of chlorophyll (Chl) molecules transferring excitation energy to PS II_α, PS II_β and PS I. Spinach chloroplasts contained almost twice as many PS II reaction centers compared to PS I reaction centers. The number N_α of chlorophyll (Chl) molecules associated with PS II_α was 234, while N_β = 100 and N_{PS I} = 210. Thus, the functional photosynthetic unit size of PS II reaction centers was different from that of PS I reaction centers. The relative electron-transport capacity of PS II was significantly greater than that of PS I. Hence, under light-limiting green excitation when both Chl a and Chl b molecules are excited equally, the limiting factor in the overall electron-transfer reaction was the turnover of PS I. The Chl composition of PS I, PS II_α and PS II_β was analyzed on the basis of a core Chl a reaction center complex component and a Chl $\text{a}\text{b-}\text{LHC}$ component. There is a dissimilar Chl $\text{a}\text{b-}\text{LHC}$ composition in the three photosystems with 77% of total Chl b associated with PS II_α only. The results indicate that PS II_α, located in the membrane of the grana partition region, is poised to receive excitation from a wider spectral window than PS II_β and PS I.     

Development of Photosystem II in dark grown Chlamydomonas reinhardtii. A light-dependent conversion of PS IIβ, QB-nonreducing centers to the PS IIα, QB-reducing form

The green alga Chlamydomonas reinhardtii is a facultative heterotroph and, when cultured in the presence of acetate, will synthesize chlorophyll (Chl) and photosystem (PS) components in the dark. Analysis of the thylakoid membrane composition and function in dark grown C. reinhardtii revealed that photochemically competent PS II complexes were synthesized and assembled in the thylakoid membrane. These PS II centers were impaired in the electron-transport reaction from the primary-quinone electron acceptor, Q_A, to the secondary-quinone electron acceptor, Q_B (Q_B-nonreducing centers). Both complements of the PS II Chl a–b light harvesting antenna (LHC II-inner and LHC II-peripheral) were synthesized and assembled in the thylakoid membrane of dark grown C. reinhardtii cells. However, the LHC II-peripheral was energetically uncoupled from the PS II reaction center. Thus, PS II units in dark grown cells had a -type Chl antenna size with only 130 Chl (a and b) molecules (by definition, PS II units lack LHC II-peripheral). Illumination of dark grown C. reinhardtii caused pronounced changes in the organization and function of PS II. With a half-time of about 30 min, PS II centers were converted froma Q_B-nonreducing form in the dark, to a Q_B-reducing form in the light. Concomitant with this change, PS II units were energetically coupled with the LHC II-peripheral complement in the thylakoid membrane and were converted to a PS II form. The functional antenna of the latter contained more than 250 Chl(a+b) molecules. The results are discussed in terms of a light-dependent activation of the Q_A-Q_B electron-transfer reaction which is followed by association of the PS II unit with a LHC II-peripheral antenna and by inclusion of the mature form of PS II (PS II) in the membrane of the grana partition region.Abbreviations Chl chlorophyll - PS photosystem - Q_A primary quinone electron acceptor of PS II - Q_B secondary quinone electron acceptor of PS II - LHC light harvesting complex - F₀ non-variable fluorescence yield - F_plf intermediate fluorescence yield plateau leyel - F_max maximum fluorescence yield - F_i initial fluorescence yield increase from F₀ to F_pl (F_pl–F₀) - F_v total variable fluorescence yield (F_m–F0) - DCMU dichlorophenyl-dimethylurea     

The quenching characteristics of potassium iridic chloride and their meaning for the origin of chlorophyll fluorescence components

To understand the origins of the different lifetime components of photosystem 2 (PS2) chlorophyll (Chl) fluorescence we have studied their susceptibility to potassium iridic chloride (K₂IrCl₆) which has been shown to bleach antenna pigments of photosynthetic bacteria (Loach et al. 1963). The addition of K₂IrCl₆ to PS2 particles gives rise to a preferential quenching of the variable Chl fluorescence (F_v). At concentrations lower than 20 M, this is brought about mainly by a decrease in the yield, but not in the lifetime, of the slowest component when all the PS2 reaction centres are closed (F_M). The yield of the middle and fast decays are not significantly altered. This type of quenching is not seen with DNB. The iridate-induced quenching of the initial fluorescence level (F₀) is due to a proportional decrease in the yield and lifetime of the three components and correlates with the observed modification in the relative quantum yield of oxygen evolution. In this concentration range a bleaching of Chl a is seen. At higher iridate levels, greater than 20 M, a proportional decrease in the lifetimes and yields of the three kinetic components is seen at F_M. These changes are associated with a carotenoid bleaching. In isolated light harvesting Chl a/b complexes of PS2 (LHC2), iridate addition converts a 4 ns decay into a 200 ps emission and both types of bleaching are observed. By also measuring the rate of PS2 trap closure versus iridate concentration, we have discussed the results in terms of excitation energy transfer.Abbreviations DNB m-dinitrobenzene - F_M maximum Chl fluorescence - F₀ initial fluorescence - F_v variable fluorescence - I pheophytin a primary electron acceptor of PS2 - P₆₈₀ chlorophyll a of photochemical centre - PS2 photosystem 2 - Q_A primary stable electron acceptor of PS2 - Chl chlorophyll - LHC2 light harvesting Chl a/b complex of PS2 - MES 2(N-morpholino) ethanesulfonic acid - DCMU 3-(3-4-dichlorophenyl) 1-1 dimethylurea - PPBQ phenyl-p-benzo-quinone - BBY PS2-enriched membranes prepared as in Berthold et al. (1981) - Q₄₀₀ PS2 electron acceptor with a midpoint potential of 400 mV     

Photosynthetic characteristics and effects of exogenous glycine of Chorispora bungeana under drought stress

We investigated the photosynthetic characteristics of Chorispora bungeana under conditions of drought stress caused by different concentrations of polyethylene glycol-6000 (PEG; 0, 5, 20, and 40%) and various concentrations of exogenous glycine (0, 5, 10, and 20 mM) with 20% PEG. We showed that moderate and severe drought stress of PEG reduced the chlorophyll (Chl) content (both Chl a and b), maximal quantum yield of PSII photochemistry (F_v/F_m), actual photochemical efficiency of PSII in light (Y_II), and quantum yield of regulated energy dissipation (Y_NPQ), while Chl a/b and quantum yield of nonregulated energy dissipation (Y_NO) increased. The low and moderate drought stress increased Mg²⁺ and Fe³⁺ contents, while a decrease in Mg²⁺ and Fe³⁺ was found under severe drought stress. Compared to sole PEG stress, the addition of exogenous 10 mM glycine increased Chl, Mg²⁺ and Fe³⁺ contents, F_v/F_m, Y_II, and Y_NPQ, and reduced Y_NO. On the contrary, 20 mM glycine showed an opposite effect, except for Y_NO. Our results proved that Chl contents and fluorescence parameters are reliable indicators for drought tolerance of C. bungeana. We suggest that a proper glycine content can relieve the effect of drought stress on C. bungeana.     

Quantum efficiency and antenna size of Photosystems IIα, IIβ and I in tobacco chloroplasts

Reaction center concentrations were determined in chloroplasts of tobacco, cv John William's Broadleaf, and its mutants Su/su and Su/su var. Aurea. Quantum yields of the primary reactions of Photosystems I, II_α and II_β (Melis, A. and Homann, P.H. (1975) Photochem. Photobiol. 21, 431–437) were obtained by measurement of their rate constants and the absorbed energy, under conditions where all three photosystems operated simultaneously and produced almost irreversibly a single charge separation.The concentrations and reaction rates of the photosystems were different in chloroplasts from the wild type and the mutants, but in chloroplasts of each type of plant used essentially all quanta absorbed by chlorophyll caused a charge separation in PS I, PS II_α or PS II_β. Since the quantum efficiency of each photosystem was close to one, kinetic differences between the photosystems and between different kinds of chloroplasts were only due to differences in antenna size. From the rate constants the number of chlorophyll molecules in the antenna of each photosystem could be calculated. It is argued that PS II_α and PS II_β must be different, independent structures.     

Chilling stress and chilling tolerance of sweet potato as sensed by chlorophyll fluorescence

We studied changes in the chlorophyll (Chl) fluorescence components in chilling-stressed sweet potato (Ipomoea batatas L. Lam) cv. Tainung 57 (TN57, chilling-tolerant) and cv. Tainung 66 (TN66, chilling-susceptible). Plants under 12-h photoperiod and 400 μmol m⁻² s⁻¹ irradiance at 24/20 °C (day/night) were treated by a 5-d chilling period at 7/7 °C. Compared to TN66, TN57 exhibited a significantly greater basic Chl fluorescence (F₀), maximum fluorescence (F_m), maximum fluorescence yield during actinic irradiation (F_m′ ), and the quantum efficiency of electron transport through photosystem 2, PS2 (Φ_PS2). Chilling stress resulted in decrease in the potential efficiency of PS2 (F_v/F_m), Φ_PS2, non-photochemical fluorescence quenching (NPQ), non-photochemical quenching (q_N), and the occurrence of chilling injury in TN66. Chilling increased the likelihood of photoinhibition, characterized by a decline in the Chl fluorescence of both cultivars, and photoinhibition during low temperature stress generally occurred more rapidly in TN66.     

Chlorophyll fluorescence and chlorophyll content in field-grown potato as affected by nitrogen supply, genotype, and plant age

Field experiments were conducted in Sicily (south Italy) to assess chlorophyll (Chl) fluorescence parameters in response of potato crop to nitrogen dose, to variation in genotype and in plant age, and to detect relationships between Chl content, fluorescence parameter F_v/F_m, and tuber yield. The experiment included five nitrogen doses (0, 10, 20, 30, and 40 g m⁻²) and four genotypes (Spunta, Sieglinde, Daytona, and Igea). Chl fluorescence parameters (initial fluorescence, F₀, maximum fluorescence, F_m, variable fluorescence, F_v, F_v/F_m, T_max (the time required to reach F_m), and Chl content were measured weekly between the appearance of the fifth and sixth leaves and the onset of plant senescence. A positive linear relationship was established between nitrogen supply and Chl content, F₀, and T_max. Nitrogen supply up to 10 g m⁻² also had a positive effect on F_m and F_v, but above this rate it reduced F_v/F_m. Spunta had the highest Chl content, F_m, F_v, and F_v/F_m, but the lowest F₀, whereas Sieglinde had the lowest Chl content, F_v, F_v/F_m, and T_max and the highest F₀. The cvs. Igea and Daytona exhibited intermediate Chl fluorescence parameters. Chl content and T_max decreased with increasing plant age, whereas F₀, F_m, and F_v increased until complete canopy development and thereafter declined until crop maturity. Tuber and plant dry matter yield were significantly correlated with Chl content, F₀, and T_max. Thus Chl fluorescence and content detect differences in the response of potato to N supply, can discriminate between genotypes, predict plant age, and yield performance under field conditions.     

The effect of redox potential on the kinetics of fluorescence induction in Photosystem II particles from Phormidium laminosum. Sigmoidicity,energy transfer and the slow phase

Fluorescence induction curves in 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU)-inhibited Photosystem (PS) II particles isolated from the blue-green alga Phormidium laminosum have been analysed as a function of redox potential. Redox titration of the initial fluorescence indicated a single component with E_m,7.5 = +30 mV (n = 1) (Bowes, J., Horton, P. and Bendall, D.S. (1981) FEBS Lett. 135, 261–264). Despite this simplified electron acceptor system and the small number of chlorophylls per reaction centre, a sigmoidal induction curve was nevertheless seen. Sigmoidicity decreased as Q was reduced potentiometrically prior to induction such that the induction was exponential when the ratio $\text{F}{}_{\mathrm{<mtext>i</mtext>}}\text{F}{}_{\mathrm{<mtext>m</mtext>}}\text{= 0.64}$. These particles also showed a slow (β) phase of induction which titrated with an E_m value slightly more positive than that of the major quencher. It is concluded that the sigmoidal shape of the fluorescence induction curve observed in Phormidium PS II particles is not a consequence of a requirement for two photons to close the PS II reaction centre, but is generated as a result of energy transfer between photosynthetic units comprising one reaction centre per approx. 50 chlorophylls. Also, the existence of PS II heterogeneity (PS II_α, PS II_β centres) does not require a structurally differentiated chloroplast, but may only indicate the extent of aggregation of PS II centres.     

Characterization of potato genotypes by chlorophyll fluorescence during plant aging in a Mediterranean environment

Field experiments were conducted in Sicily (south Italy) during two seasons to characterize by chlorophyll (Chl) fluorescence four genotypes (Spunta, Sieglinde, Daytona, and Ninfa) of potato (Solanum tuberosum L.) for off-season production during plant aging and to analyse the possible relation between Chl parameters and tuber yield. Chl fluorescence parameters [initial fluorescence (F₀), maximum fluorescence (F_m), F_v/F_m, time in which maximal fluorescence occurs (T_max)] gained from Kautsky kinetics and Chl content were measured weekly, from 5^th to 6^th leaf appearance to beginning of plant senescence in the first season and to full plant senescence in the second season. F₀ and F_v/F_m were the most reliable Chl fluorescence parameters for the definition of genotypic differences while Chl content and T_max were the most reliable Chl parameters to predict plant aging. Tuber yield was highly correlated with Chl content, T_max, F₀, and F_m.     

Characterization of chlorophyll fluorescence quenching in chloroplasts by fluorescence spectroscopy at 77 K I. ΔpH-dependent quenching

The nature of the light-induced ΔpH-dependent decline of chlorophyll a fluorescence in intact and broken spinach chloroplasts was investigated. Fluorescence spectra at 77 K of chloroplasts frozen in the low-fluorescent (high ΔpH) state showed increased ratios of the band peak at 735 nm (Photosystem (PS) I fluorescence) to the peak at 695 nm (PS II fluorescence). The increase in the $\text{F}{}_{735}\text{F}{}_{695}$ ratio at 77 K was related to the extent of fluorescence quenching at room temperature. Normalization of low-temperature spectra with fluorescein as an internal standard revealed a lowering of F₆₉₅ that was not accompanied by an increase in F₇₃₅: preillumination before freezing decreased both F₆₉₅ and, to a lesser extent, F₇₃₅ in the spectra recorded at 77 K. Fluorescence induction of chloroplasts frozen in the low-fluorescent state showed a markedly decreased variable fluorescence (F_v) of PS II, but no concomitant increase in initial fluorescence (F₀) of PS I. Thus, the buildup of a proton gradient at the thylakoid membrane, as reflected by fluorescence quenching at room temperature, affects low-temperature fluorecence emission in a manner entirely different from the effect of removal of Mg²⁺, which is thought to alter the distribution of excitation energy in favor of PS I. The ΔpH-dependent quenching therefore cannot be caused by such change in energy distribution and is suggested to reflect increased thermal deactivation.     

Analysis of xanthophyll cycle carotenoids and chlorophyll fluorescence in light intensity-dependent chlorophyll-deficient mutants of wheat and barley

Three light intensity-dependent Chl b-deficient mutants, two in wheat and one in barley, were analyzed for their xanthophyll cycle carotenoids and Chl fluorescence characteristics under two different growth PFDs (30 versus 600 mol photons·m^–2 s^–1 incident light). Mutants grown under low light possessed lower levels of total Chls and carotenoids per unit leaf area compared to wild type plants, but the relative proportions of the two did not vary markedly between strains. In contrast, mutants grown under high light had much lower levels of Chl, leading to markedly greater carotenoid to Chl ratios in the mutants when compared to wild type. Under low light conditions the carotenoids of the xanthophyll cycle comprised approximately 15% of the total carotenoids in all strains; under high light the xanthophyll cycle pool increased to over 30% of the total carotenoids in wild type plants and to over 50% of the total carotenoids in the three mutant strains. Whereas the xanthophyll cycle remained fairly epoxidized in all plants grown under low light, plants grown under high light exhibited a considerable degree of conversion of the xanthophyll cycle into antheraxanthin and zeaxanthin during the diurnal cycle, with almost complete conversion (over 90%) occurring only in the mutants. 50 to 95% of the xanthophyll cycle was retained as antheraxanthin and zeaxanthin overnight in these mutants which also exhibited sustained depressions in PS II photochemical efficiency (F_v/F_m), which may have resulted from a sustained high level of photoprotective energy dissipation activity. The relatively larger xanthophyll cycle pool in the Chl b-deficient mutant could result in part from the reported concentration of the xanthophyll cycle in the inner antenna complexes, given that the Chl b-deficient mutants are deficient in the peripheral LHC-II complexes.Abbreviations A antheraxanthin - Chl chlorophyll - F_o and F_m minimal yield (at open PS II reaction centers) and maximal yield (at closed centers) of chlorophyll fluorescence in darkness - F level of fluorescence during illumination with photosynthetically active radiation - F_m maximal yield (at closed centers) of chlorophyll fluorescence during illumination with photosynthetically active radiation - (F_m–F)/F_m actual efficiency of PS II during illumination with photosynthetically active radiation - F_v/F_m+(F_m–F_o)/F_m intrinsic efficiency of PS II in darkness - LHC_II light-harvesting chlorophyll-protein complex of Photosystem II - PFD photon flux density (between 400 and 700 nm) - PS I Photosystem I - PS II Photosystem II - V violaxanthin - Z zeaxanthin     

Regulation of the Hill reaction by cations and its abolishment by uncouplers

Concurrent measurements of P-700 turnover and the reduction of K₃Fe(CN)₆ revealed an identical relative quantum yield for both reactions in isolated pea chloroplasts as well as chloroplast particles from wild type Scenedesmus. On the other hand, chloroplast particles of wild type Scenedesmus showed the same relative quantum yield for the Hill reaction as those of the P-700-free mutant No. 8, indicating that P-700 is not required for ferricyanide reduction.Several metal ions, such as Mg²⁺, Ca²⁺, Na⁺ and K⁺ stimulated the reduction of K₃Fe(CN)₆. In short wavelength light, the stimulation was a function of light intensity, varying in magnitude from an approximate doubling of the yield in low intensities to only a slight increase at light saturation. P-700 was totally unaffected by the cations.The effect of the metal salts was abolished in the presence of uncouplers of photophosphorylation.The data reconcile several divergent results concerning the effect of divalent cations on the reduction of ferricyanide which have been reported in the recent literature. On the whole the experiments suggest that the Hill acceptor can be reduced at two sites. The stimulation of the Hill reaction by metal ions is proposed to be due to an activation of Photosystem II and a more efficient utilization of quanta at the expense of radiationless de-excitation.     

Photosynthesis in leaves of <Emphasis Type="Italic">Nicotiana tabacum</Emphasis> L. infected with tobacco mosaic virus

In tobacco leaves inoculated with tobacco mosaic virus (TMV), changes in chlorophyll (Chl) and carotenoid contents, parameters of slow Chl fluorescence kinetics, i.e. the maximum quantum yield of photosystem (PS2) photochemistry F_v/F_m, the effective quantum yield of photochemical energy conversion in PS2 Φ₂, ratio of quantum yields of photochemical and concurrent non-photochemical processes in PS2 F_v/F₀, non-photochemical quenching (NPQ), and photochemical activities of isolated chloroplasts from systemically infected tobacco leaves were investigated. We compared two successive stages of infection, the first in the stage of vein clearing at 9^th day post inoculation (dpi) and the second at 22^nd dpi when two different regions, i.e. light- (LGI) or dark-green (DGI) islands in the infected leaf were apparent and symptoms were fully developed. These two different regions were measured separately. The Chl and carotenoid contents in infected leaves decreased with a progression of infection and were lowest in LGI in the second stage. Also the ratio of Chl a/b declined in similar manner. The maximum quantum yield of PS2 photochemistry F_v/F_m, was decreased in the following order: first stage, DGI, and LGI. The same is true for the ratio F_v/F₀. The decrease of Φ₂ in infected leaves declined as compared to their controls. On the contrary, NPQ increased in infected leaves, the highest value was found in the first infection stage. Photochemical activities of the whole electron transport chain in isolated chloroplasts dramatically declined with the progression of symptoms, the lowest value was in LGI. Similarly, but to a lesser extent, the activity of PS2 in isolated chloroplasts decreased in infected leaves. Generally, the most marked impairment of the photosynthetic apparatus was manifested in the LGI of infected leaves.     

Photosynthesis and protective mechanisms during ageing in transgenic tobacco leaves with over-expressed cytokinin oxidase/dehydrogenase and thus lowered cytokinin content

The content of cytokinins (CKs), the plant inhibitors of the final phase of plant development, senescence, is effectively controlled by irreversible degradation catalysed by cytokinin oxidase/dehydrogenase (CKX). In transgenic tobacco, denoted as AtCKX, with over-expressed CKX causing lowered CK content, we investigated changes in the time courses of chlorophyll (Chl) and xanthophyll (violaxanthin, antheraxanthin, zeaxanthin, neoxanthin, and lutein) contents. We also determined parameters of slow Chl fluorescence kinetics such as minimum Chl fluorescence yield in the darkadapted state F₀, maximum quantum yield of PS2 photochemistry (F_v/F_m), maximum ratio of quantum yields of photochemical and concurrent non-photochemical processes in photosystem 2 (PS2), F_v/F₀, non-photochemical quenching (NPQ), and effective quantum yield of photochemical energy conversion in PS2 (Φ₂). We used three different developmental leaf stages, old, mature, and young, and compared this with time courses of these characteristics in leaves with natural CK levels. The parameters F_v/F_m, F_v/F₀, and Φ₂ were unchanged during ageing in AtCKX plants in contrast to control ones where a significant decrease in old leaves was found. In control plants F₀ increased during ageing, but in the oldest leaf a considerable decrease was observed. This could indicate progressive damage to PS2 reaction centres and then detachment and rapid degradation of Chl. This is in agreement with time course of Chl content. NPQ decreased with age and was similar in both plant types. We observed a decline of xanthophyll contents in the oldest leaves in both plant types, but the contents were enhanced in AtCKX compared to control plants, especially of neoxanthin. The higher xanthophyll contents in the transgenic plants contribute to a better photoprotection and the fluorescence parameters indicated that photosynthetic apparatus was in better condition compared to control and it consequently postponed the onset of leaf senescence.     

Analysis of chlorophyll fluorescence induction curves in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea as a function of magnesium concentration and NADPH-activated light-harvesting chlorophyll ab-protein phosphorylation

The effect of Mg²⁺ concentration and phosphorylation of light-harvesting chlorophyll $\text{a}\text{b-}\text{protein}$ on various chlorophyll fluorescence induction parameters of isolated pea thylakoids has been studied. (1) Lowering the Mg²⁺ concentration from 3 to 0.4 mM decreases only the variable fluorescence (F_v) and the area above the induction curve while at the same time increasing the slow exponential component of the rise (β_max). (2) A further decrease in Mg²⁺ concentration from 0.4 to 0 mM decreases the initial (F₀) fluorescence level such that the ratio $\text{F}{}_{\mathrm{<mtext>v</mtext>}}\text{F}{}_{\mathrm{<mtext>m</mtext>}}$ increases slightly as does the area above the induction curve and β_max. (3) Thylakoid membranes, phosphorylated at 5 mM Mg²⁺, show an equal decrease in F_v and F₀, no change in the area above the induction curve and an increase in β_max. At 2 mM Mg²⁺, however, phosphorylation induced a more extensive quenching of F_v so that the $\text{F}{}_{\mathrm{<mtext>v</mtext>}}\text{F}{}_{\mathrm{<mtext>m</mtext>}}$ ratio was lowered and the area above the induction curve decreased while β_max increased. (4) When phosphorylated membranes were subsequently suspended in an Mg²⁺-free medium the effect on F₀ due to phosphorylation was found to be additive to that due to the absence of Mg²⁺. The effect of membrane phosphorylation on fluorescence is discussed in relation to the control of excitation energy distribution and shows that different mechanisms operate depending on the background Mg²⁺ levels. At high Mg²⁺ the phosphorylation seems to affect the absorption cross-section of Photosystem II while at lower Mg²⁺ levels there is an additional effect of increased spillover from Photosystem II to I.     

Characterization of chlorophyll fluorescence quenching in chloroplasts by fluorescence spectroscopy at 77 K II. ATP-dependent quenching

In intact, uncoupled type B chloroplasts from spinach, added ATP causes a slow light-induced decline ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{≈ 3}\text{min}$) of chlorophyll a fluorescence at room temperature. Fluorescence spectra were recorded after fast cooling to 77 K and normalized with fluorescein as an internal standard. Related to the fluorescence quenching at room temperature, an increase in Photosystem (PS) I fluorescence (F₇₃₅) and a decrease in PS II fluorescence (F₆₉₅) were observed in the low-temperature spectra. The change in the $\text{F}{}_{735}\text{F}{}_{695}$ ratio was abolished by the presence of methyl viologen. Fluorescence induction at 77 K of chloroplasts frozen in the quenched state showed lowered variable (F_v) and initial (F₀) fluorescence at 690 nm and an increase in F₀ at 735 nm. The results are interpreted as indicating an ATP-dependent change of the initial distribution of excitation energy in favor of PS I, which is controlled by the redox state of the electron-transport chain and, according to current theories, is caused by phosphorylation of the light-harvesting complex.     

Leaf characteristics and diurnal variation of chlorophyll fluorescence in leaves of the ‘bana’ vegetation of the amazon region

In six dominant species of the Amazonian ‘Bana’ vegetation, leaf blade characteristics, pigment composition, and chlorophyll (Chl) fluorescence parameters were measured in young and mature leaves under field conditions. Leaf δ¹³C was comparable in the six species, which suggested that both expanding and expanded leaves contained organic matter fixed under similar intercellular and ambient CO₂ concentration (C _i/C _a). High leaf C/N and negative δ¹⁵N values found in this habitat were consistent with the extreme soil N-deficiency. Analysis of Chl and carotenoids showed that expanding leaves had an incomplete development of photosynthetic antenna when compared to adult leaves. Dynamic inactivation of photosystem 2 (PS2) at midday was observed at both leaf ages as F_v/F_m decreased compared to predawn values. Adult leaves reached overnight F_v/F_m ratios typical of healthy leaves. Overnight recovery of F_v/F_m in expanding leaves was incomplete. F₀ remained unchanged from midday to predawn and F_v tended to increase from midday to predawn. The recovery from midday depression observed in adult leaves suggested an acclimatory down-regulation associated with photo-protection and non-damage of PS2.     

State Transitions in the Green Alga Scenedesmus Obliquus Probed by Time-Resolved Chlorophyll Fluorescence Spectroscopy and Global Data Analysis

Decay-associated fluorescence spectra of the green alga Scenedesmus obliquus have been measured by single-photon timing with picosecond resolution in various states of light adaptation. The data have been analyzed by applying a global data analysis procedure. The amplitudes of the decay-associated spectra allow a determination of the relative antenna sizes of the photosystems. We arrive at the following conclusions: (a) The fluorescence kinetics of algal cells with open PS II centers (F₀ level) have to be described by a sum of three exponential components. These decay components are attributed to photosystem (PS) I (τ ≈ 85 ps, λ_max^em ≈ 695-700 nm), open PS II α-centers (τ ≈ 300 ps, λ_max^em = 685 nm), and open PS II β-centers (τ ≈ 600 ps, λ_max^em = 685 nm). A fourth component of very low amplitude (τ ≈ 2.2-2.3 ns, λ_max^em = 685 nm) derives from dead chlorophyll. (b) At the F_max level of fluorescence there are also three decay components. They originate from PS I with properties identical to those at the F₀ level, from closed PS II α-centers (τ ≈ 2.2 ns, λ_max^em = 685 nm) and from closed PS β-centers (τ ≈ 1.2 ns, λ_max^em = 685 nm). (c) The major effect of light-induced state transitions on the fluorescence kinetics involves a change in the relative antenna size of α- and β-units brought about by the reversible migration of light-harvesting complexes between α-centers and β-centers. (d) A transition to state II does not measurably increase the direct absorption cross-section (antenna size) of PS I. Our data can be rationalized in terms of a model of the antenna organization that relates the effects of state transitions and light-harvesting complex phosphorylation with the concepts of PS II α,β-heterogeneity. We discuss why our results are in disagreement with those of a recent lifetime study of Chlorella by M. Hodges and I. Moya (1986, Biochim. Biophys. Acta., 849:193-202).     

Studies on the functional organization of Photosystem II. The effect of protein-modifying procedures and of ADRY agents on the reaction pattern of photosystem II in Tris-washed chloroplasts

The role of the protein matrix embedding the functionally active redox components of Photosystem II reaction centers has been studied by investigating the effects of procedures which modify the structure of proteins. In order to reduce the influence of the electron transport involving secondary donor and acceptor components, Triswashed chloroplasts were used which are completely deprived of their oxygen-evolving capacity. The functional activity was detected via absorption changes, reflecting at 334 and 690 or 834 nm the turnover of the primary plastoquinone acceptor, X320, and of the photochemically active chlorophyll a complex, Chl a_II, respectively, and at 520 nm the transient formation of a transmembrane electric potential gradient. Under repetitive flash excitation of Tris-washed chloroplasts it was found that: (a) The relaxation kinetics at 690 nm become significantly accelerated in the presence of external electron donors. (b) Trypsin treatment blocks to a high degree the turnover of Chl a_II and X320 unless exogenous acceptors are present, which directly oxidize X320^?, such as K₃Fe(CN)₆. (c) In the presence of K₃Fe(CN)₆ the recovery kinetics of Chl a_II and X320 are retarded markedly by trypsin, followed by a progressive decline in the extent thereof. (d) 2-(3-Chloro-4-trifluoromethyl)anilino-3,5-dinitrothiophene (ANT 2p), known to reduce the lifetime of S₂ and S₃ in normal chloroplasts, significantly accelerates the recovery of Chl a_II. 10 μs kinetics are observed which correspond with the electron-transfer rate from D₁ to Chl a⁺_II. ANT 2p simultaneously retards the decay kinetics of X320^? and of the electrochromic absorption changes. (e) The kinetic pattern of the electrochromic absorption changes is also affected by the salt content of the suspension. Under dark-adapted conditions, the 10 μs relaxation kinetics of the 834 nm absorption change due to the first flash are hardly affected by mild trypsinization of 5–10 min duration, whereas the amplitude decreases by approx. 30%. The data obtained in Tris-washed chloroplasts could consistently be interpreted as a modification of the back reaction between X320^? and Chl a⁺_II which is caused solely by a change in the reactivity of X320 due to trypsin-induced degradation of the native X320-B apoprotein. Furthermore, ADRY agents are inferred to stimulate cyclic electron flow, which leads to reduction of D⁺₁ between the flashes. A simplified scheme is discussed which describes the functional organization of the reaction center complex.     

Detection of photosynthetic energy storage in a photosystem I reaction center preparation by photoacoustic spectroscopy

Thermal emission and photochemical energy storage were examined in photosystem I reaction center/core antenna complexes (about 40 Chl a/P700) using photoacoustic spectroscopy. Satisfactory signals could only be obtained from samples bound to hydroxyapatite and all samples had a low signal-to-noise ratio compared to either PS I or PS II in thylakoid membranes. The energy storage signal was saturated at low intensity (half saturation at 1.5 W m^-2) and predicted a photochemical quantum yield of >90%. Exogenous donors and acceptors had no effect on the signal amplitudes indicating that energy storage is the result of charge separation between endogenous components. Fe(CN)₆ ^-3 oxidation of P700 and dithionite-induced reduction of acceptors F_A-F_B inhibited energy storage. These data are compatible with the hypothesis that energy storage in PS I arises from charge separation between P700 and Fe-S centers F_A-F_B that is stable on the time scale of the photoacoustic modulation. High intensity background light (160 W m^-2) caused an irreversible loss of energy storage and correlated with a decrease in oxidizable P700; both are probably the result of high light-induced photoinhibition. By analogy to the low fluorescence yield of PS I, the low signal-to-noise ratio in these preparations is attributed to the short lifetime of Chl singlet excited states in PS I-40 and its indirect effect on the yield of thermal emission.Abbreviations FFT fast Föurier transform - HA hydroxyapatite - I₅₀ half saturation intensity for energy storage - PA photoacoustic - PS photosystem - PS I-40 photosystem I reaction center/core antenna complex containing about 40 Chl a/P700 - 201-1 photoacoustic energy storage signal - S/N signal-to-noise