A comparative study of the thermal stability of plastocyanin, cytochrome c6 and Photosystem I in thermophilic and mesophilic cyanobacteria

Cytochrome c₆ (Cyt) from the thermophilic cyanobacterium Phormidium laminosum has been purified and characterized. It is a mildly acidic protein, with physicochemical properties very similar to those of plastocyanin (Pc). This is in agreement with the functional interchangeability of the two metalloproteins as electron donors to Photosystem I (PS I). The kinetic analyses of the interaction of Pc and Cyt with Photosystem I show that both metalloproteins reduce PS I with similar efficiencies, according to an oriented collisional kinetic model involving repulsive electrostatic interactions. The thermostability study of the Phormidium Pc/PS I system compared with those from mesophilic cyanobacteria (Synechocystis, Anabaena and Pseudanabaena) reveals that Pc is the partner limiting the thermostability of the Phormidium couple. The cross-reactions between Pc and PS I from different organisms demonstrate not only that Phormidium Pc enhances the stability of the Pc/PS I system using PS I from mesophilic cyanobacteria, but also that Phormidium PS I possesses a higher thermostability than the other photosystems. This revised version was published online in June 2006 with corrections to the Cover Date.     

Electrogenic reduction of the primary electron donor P700 by plastocyanin in photosystem I complexes

An electrometric technique was used to investigate electron transfer between spinach plastocyanin (Pc) and photooxidized primary electron donor P700 in photosystem I (PS I) complexes from the cyanobacterium Synechocystis sp. PCC 6803. In the presence of Pc, the fast unresolvable kinetic phase of membrane potential generation related to electron transfer between P700 and the terminal iron–sulfur acceptor F_B was followed by additional electrogenic phases in the microsecond and millisecond time scales, which contribute approximately 20% to the overall electrogenicity. These phases are attributed to the vectorial electron transfer from Pc to the protein-embedded chlorophyll dimer P700⁺ within the PsaA/PsaB heterodimer. The observed rate constant of the millisecond kinetic phase exhibited a saturation profile at increasing Pc concentration, suggesting the formation of a transient complex between Pc and PS I with the dissociation constant K_d of about 80 μM. A small but detectable fast electrogenic phase was observed at high Pc concentration. The rate constant of this phase was independent of Pc concentration, indicating that it is related to a first-order process.     

A study on the dynamic features of photosystem stoichiometry: Accomplishments and problems for future studies

Our study on the regulation by light of photosystem stoichiometry in cyanophytes is briefly reviewed here. It can be summarized as follows: Adjustment of photosystem stoichiometry results in optimization of photosynthetic efficiency under the prevailing light-growth conditions. Photosystem (PS) I is primarily affected, increasing or decreasing, relative to PS II. The regulation of PS I synthesis appears to be at the translation or the post-translational level. The regulation is probably governed by changes in the redox level of the electron transport components, most probably at Cyt b₆in the Cyt b₆–f complex. Our results to date are at the physiological level, but they have raised many questions and provided suggestions for future directions in exploring the mechanism of regulation, some of which are discussed in this perspective.     

A large fraction of PsaF is nonfunctional in photosystem I complexes lacking the PsaJ subunit

PsaJ is a small hydrophobic subunit of the photosystem I complex (PSI) whose function is not yet fully understood. Here we describe mutants of the green alga Chlamydomonas reinhardtii, in which the psaJ chloroplast gene has been inactivated either in a wild-type or in a PsaF-deficient nuclear background. Cells lacking one or both subunits grow photoautotrophically and contain normal levels of PSI. Flash-absorption spectroscopy performed with isolated PSI particles isolated from the PsaJ-deficient strain indicates that only 30% of the PSI complexes oxidize plastocyanin (Pc) or cytochrome c6 (Cyt c6) with kinetics identical to wild type, whereas the remaining 70% follow slow kinetics similar to those observed with PsaF-deficient PSI complexes. This feature is not due to partial loss of PsaF, as the PsaJ-less PSI complex contains normal levels of the PsaF subunit. The N-terminal domain of PsaF can be cross-linked to Pc and Cyt c6 indicating that in the absence of PsaJ, this domain is exposed in the lumenal space. Therefore, the decreased amount of functional PsaF revealed by the electron-transfer measurements is best explained by a displacement of the N-terminal domain of PsaF which is known to provide the docking site for Pc and Cyt c6. We propose that one function of PsaJ is to maintain PsaF in a proper orientation which allows fast electron transfer from soluble donor proteins to P700(+).     

Deciphering the 820 nm signal: redox state of donor side and quantum yield of Photosystem I in leaves

By recording leaf transmittance at 820 nm and quantifying the photon flux density of far red light (FRL) absorbed by long-wavelength chlorophylls of Photosystem I (PS I), the oxidation kinetics of electron carriers on the PS I donor side was mathematically analyzed in sunflower (Helianthus annuus L.), tobacco (Nicotiana tabacum L.) and birch (Betula pendula Roth.) leaves. PS I donor side carriers were first oxidized under FRL, electrons were then allowed to accumulate on the PS I donor side during dark intervals of increasing length. After each dark interval the electrons were removed (titrated) by FRL. The kinetics of the 820 nm signal during the oxidation of the PS I donor side was modeled assuming redox equilibrium among the PS I donor pigment (P700), plastocyanin (PC), and cytochrome f plus Rieske FeS (Cyt f + FeS) pools, considering that the 820 nm signal originates from P700⁺ and PC⁺. The analysis yielded the pool sizes of P700, PC and (Cyt f + FeS) and associated redox equilibrium constants. PS I density varied between 0.6 and 1.4 μmol m⁻². PS II density (measured as O₂ evolution from a saturating single-turnover flash) ranged from 0.64 to 2.14 μmol m⁻². The average electron storage capacity was 1.96 (range 1.25 to 2.4) and 1.16 (range 0.6 to 1.7) for PC and (Cyt f + FeS), respectively, per P700. The best-fit electrochemical midpoint potential differences were 80 mV for the P700/PC and 25 mV for the PC/Cyt f equilibria at 22 °C. An algorithm relating the measured 820 nm signal to the redox states of individual PS I donor side electron carriers in leaves is presented. Applying this algorithm to the analysis of steady-state light response curves of net CO₂ fixation rate and 820 nm signal shows that the quantum yield of PS I decreases by about half due to acceptor side reduction at limiting light intensities before the donor side becomes oxidized at saturating intensities. Footnote: This revised version was published online in August 2006 with corrections to the Cover Date.     

Coregulation of electron transport through PS I by Cyt b 6 f,excitation capture by P700 and acceptor side reduction. Time kinetics and electron transport requirement

Regulation of electron transport rate through Photosystem I (PS I) was investigated in intact sunflower leaves. The rate constant of electron donation via the cytochrome b ₆ f complex (k_q, s^–1) was obtained from the postillumination P700⁺ reduction rate, measured as the exponential decay of the light-dark difference (D830) of the 830 nm transmission signal. D830 corresponding to maximum oxidisable P700 (D830_m) was obtained by applying white light flashes of different intensity and extrapolating the plot of the quantum yield Y vs. D830 to the axis of abscissae (Y->0). Maximum quantum yield of PS I at completely reduced P700 (Y_m) was obtained by extrapolating the same plot to the axis of ordinates (D830->0). Regulation of k_q, D830_m and Y_m under rate-limiting CO₂ and O₂ concentrations applied after air (21% O₂, 310 ppm CO₂) was investigated. The amplitude of the downregulation of k_q (photosynthetic control) was maximal when electron transport rate (ETR) was limited to about 3 nmol cm^–2 s^–1 and decreased when ETR was higher or lower. Downregulation did not occur in the absence of CO₂ and O₂. These gases acted only as substrates of ribulosebisphosphate carboxylase-oxygenase, no high-affinity reaction of O₂ leading to enhanced photosynthetic control (e.g. Mehler reaction) was detected. After the transition, D830_m at first decreased and then increased again, showing that the reduction of the PS I acceptor side disappeared as a result of the downregulation of k_q. The variation of Y_m had two reasons, PS I acceptor side reduction and variable excitation capture efficiency by P700. It is concluded that electron transport through PS I is coregulated by the rate of plastoquinol oxidation at Cyt b ₆ f, excitation capture efficiency by P700, and by acceptor side reduction.Abbreviations Cyt b ₆ f cytochrome b ₆ f complex - D830 difference of the 830 nm signal from the dark level - ETR electron transport rate - PAD photon absorption density nmol cm^–2 s^–1 - PFD incident photon flux density, nmol cm^–2 s^–1 - PS I Photosystem I - PS II Photosystem II - PQH₂ plastoquinol - P700 Photosystem I donor pigment - Y quantum yield of PS I electron transport, rel. un.     

Chemical evolution of peroxidase — amino acid pentacyanoferrate (II) complexes as model

Complexes of the type [Fe(II)(CN)₅(L)] ^n– (wheren=3, or 4;L = glycine, histidine, imidazole, and triglycine) are proposed as evolutionary model of peroxidases. Detailed kinetic investigation for disproportionation of hydrogen peroxide catalysed by [Fe(II)(CN)₅(L)] ^n– complexes at 40°C and pH 9.18 are discussed. Decomposition of hydrogen peroxide catalysed by above complexes conforms to Michaelis-Menten type kinetics.     

Regulation of Photosystem Stoichiometry in the Photosynthetic System of the Cyanophyte Synechocystis PCC 6714 in Response to Light-Intensity

Light-induced changes in stoichiometry among three thylakoidcomponents, PS I, PS II and Cyt b₆-f complexes, were studiedwith the cyanophyte Synechocystis PCC 6714. Special attentionwas paid to two aspects of the stoichiometric change; first,a comparison of the patterns of regulation in response to differencesin light-intensity with those induced by differences in light-quality,and second, the relationship between regulation of the stoichiometryand the steady state of the electron transport system. Resultsfor the former indicated that (1) the abundance of PS I on aper cell basis was reduced under white light at the intensityas high as that for light-saturation of photosynthesis, butPS I per cell was increased under low light-intensity, (2) PSII and Cyt b₆-f complexes remained fairly constant, and (3)changes in the abundance of PS I depended strictly on proteinsynthesis. The pattern was identical with that of chromaticregulation. For the second problem, the redox steady-statesof Cyt f and P700 under white light of various intensities weredetermined by flash-spectroscopy. Results indicated that (1)Cyt f and P700 in cells grown under low light-intensity [highratio of PS I to PS II (PS I/PS II)] were markedly oxidizedwhen the cells were exposed to high light-intensity, while theyremained in the reduced state under low light-intensity. (2)After a decrease in the abundance of PS I, most of P700 remainedin the reduced state even under high light-intensity, whilethe level of reduced Cyt f remained low. (3) Both Cyt f andP700 in cells of low PS I/PS II were fully reduced under lowlight-intensity, and Cyt f reduction following the flash wasrapid, which indicates that the turnover of PS I limits theoverall rate of electron flow. After an increase in the abundanceof PS I, the electron transport recovered from the biased state.(4) The redox steady-state of the Cyt b₆-f complex correlatedwell with the regulation of PS I/PS II while the state of thePQ pool did not. Based on these results, a working model ofthe regulation of assembly of the PS I complex, in which theredox steady-state of the Cyt b₆-f complex is closely relatedto the primary signal, is proposed. (Received August 2, 1990; Accepted December 10, 1990)     

Spectral and Kinetic Analysis of the Energy Coupling in the PS I–LHC I Supercomplex from the Green Alga Chlamydomonas reinhardtii at 77 K

Energy transfer processes in the chlorophyll antenna of the PS I–LHCI supercomplexes from the green alga Chlamydomonas reinhardtii have been studied at 77 K using transient absorption spectroscopy with multicolor excitation in the 640–670 nm region. Comparison of the kinetic data obtained at low and room temperatures indicates that the slow ∼ ∼100 ps excitation equilibration phase that is characteristic of energy coupling of the LHCI peripheral antenna to the PS I core at physiological temperatures (Melkozernov AN, Kargul J, Lin S, Barber J and Blankenship RE (2004) J Phys Chem B 108: 10547–10555) is not observed in the excitation dynamics of the PS I–LHCI supercomplex at 77 K. This suggests that at low temperatures the peripheral antenna is energetically uncoupled from the PS I core antenna. Under these conditions the observed kinetic phases on the time scales from subpicoseconds to tens of picoseconds represent the superposition of the processes occurring independently in the PS I core antenna and the Chl a/b containing LHCI antenna. In the PS I–LHCI supercomplex with two uncoupled antennas the excitation is channeled to the excitation sinks formed at low temperature by clusters of red pigments. A better spectral resolution of the transient absorption spectra at 77 K results in detection of two ΔA bands originating from the rise of photobleaching on the picosecond time scale of two clearly distinguished pools of low energy absorbing Chls in the PS I–LHCI supercomplex. The first pool of low energy pigments absorbing at 687 nm is likely to originate from the red pigments in the LHCI where the Lhca1 protein is most abundant. The second pool at 697 nm is suggested to result either from the structural interaction of the LHCI and the PS I core or from other Lhca proteins in the antenna. The kinetic data are discussed based on recent structural models of the PS I–LHCI. It is proposed that the uncoupling of pigment pools may be a control mechanism that regulates energy flow in Photosystem I.     

Kinetic analyses of the OJIP chlorophyll fluorescence rise in thylakoid membranes

N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD) was previously used to study the kinetics of the OJIP chlorophyll fluorescence rise. The present study is an attempt to elucidate the origin of TMPD-induced delay and quenching of the I–P step of fluorescence rise. For this purpose, we analyzed the kinetics of OJIP rise in thylakoid membranes in which electron transport was modified using ascorbate, methyl viologen (MV), and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB). In the absence of TMPD, the OJIP kinetics of fluorescence induction (FI) was not altered by ascorbate. However, ascorbate eliminated the I–P rise delay caused by high concentrations of TMPD. On the other hand, neither ascorbate nor DBMIB, which blocks the electron release from Photosystem II (PS II) at the cytochrome b₆/f complex, could prevent the quenching of I–P rise by TMPD. In control thylakoids, MV suppressed the I–P rise of FI by about 60. This latter effect was completely removed if the electron donation to MV was blocked by DBMIB unless TMPD was present. When TMPD intercepted the linear electron flow from PS II, re-oxidation of TMPD by photosystem I (PS I) and reduction of MV fully abolished the I–P rise. The above is in agreement with the fact that TMPD can act as an electron acceptor for PS II. With MV, the active light-driven uptake of O₂ during re-oxidation of TMPD by PS I contributes towards an early decline in the I–P step of the OJIP fluorescence rise.     

Time-resolved chlorophyll fluorescence studies on photosynthetic mutants of Chlamydomonas reinhardtii: origin of the kinetic decay components

The room temperature chlorophyll fluorescence decay kinetics of photosynthetic mutants of Chlamydomonas reinhardtii have been measured as a function of Photosystem 2 (PS2) trap closure, DNB-induced quenching at F_M, and time-resolved emission spectra. The overall decays have been analyzed in terms of three or four kinetic components where necessary. A comparison of the characteristics of the decay components exhibited by the mutants with the wild-type has been carried out to elucidate the precise origins of the different emissions in relation to the observed pigment-protein complexes. It is shown that a) charge recombination in PS2 is not necessary for the presence of long-lived decay components, b) there are two rapid PS1-associated emissions (=30 and 150–200 ps), c) a slow PS1 decay is observed (=1.73 ns) in the absence of PS1 reaction centres, d) the two variable components (=0.25–1.2 and 0.5–2.2 ns) observed in the wild-type arise from LHC2 and e) a rapid (=50–250 ps) decay is associated with the PS2 core antenna (CP3 and CP4). These results show that the intact thylakoid membrane system is too complex to distinguish all of the individual kinetic components.Abbreviations Aexp preexponential factor (Amplitude) - chl chlorophyll - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - DNB m, dinitrobenzene - F_M maximum chl fluorescence level - F₀ initial chl fluorescence level - F_v variable chl fluorescence (F_M–F₀) - LHC light harvesting chl a/b protein complex - PS photosystem - Q_A primary stable electron acceptor of PS2     

Effect of Supra-High Irradiation on the Photosynthetic System of the Cyanophyte Synechocystis PCC 6714

Stability of thylakoid components under supra-high irradiancewas studied with the cyanophyte Synechocystis PCC 6714. Theactivity of overall photosynthesis was quickly inactivated (T_1/2=20min) under supra-high irradiance (300 W m^–2, white light).In parallel with the inactivation of photosynthesis, QA in PSII was also inactivated. Both inactivations were acceleratedby chloramphenicol (CAP) addition. The reactivation of PS IIrequired weak irradiation and was suppressed by CAP. However,PS I measured as P700 was very stable. The level of PS I measuredas P700 was not significantly reduced by the irradiation for12 h even in the presence of CAP while the level of Cyt b₅₅₉,component of PS II, was decreased markedly. The function ofPS I before and after supra-high irradiation with CAP was examinedby comparing sizes of P700 oxidation induced by a short flash,by a continuous light, and by determination of O₂-and ferredoxin-reduction.No difference was observed in PS I actions before and afterthe irradiation treatment. These results indicate that the PSI complex is very tolerant of supra-high irradiation. However,the cells grown under supra-high irradiance contained much fewerPS I and PS II complexes than Cyt b₆–f complexes. Theformer levels were reduced to a half to one fourth of thosebefore growth while the level of Cyt b₆–f complex wasnot reduced so much. A possible mechanism for changes in thylakoidcomposition under supra-high irradiation was discussed. (Received February 16, 1991; Accepted June 12, 1991)     

Short-term responses of Photosystem I to heat stress

When 23°C-grown potato leaves (Solanum tuberosum L.) were exposed for 15 min to elevated temperatures in weak light, a dramatic and preferential inactivation of Photosystem (PS) II was observed at temperatures higher than about 38°C. In vivo photoacoustic measurements indicated that, concomitantly with the loss of PS II activity, heat stress induced a marked gas-uptake activity both in far-red light (>715 nm) exciting only PS I and in broadband light (350–600 nm) exciting PS I and PS II. In view of its suppression by nitrogen gas and oxygen and its stimulation by high carbon-dioxide concentrations, the bulk of the photoacoustically measured gas uptake by heat-stressed leaves was ascribed to rapid carbon-dioxide solubilization in response to light-modulated stroma alkalization coupled to PS I-driven electron transport. Heat-induced gas uptake was observed to be insensitive to the PS II inhibitor diuron, sensitive to the plastocyanin inhibitor HgCl₂ and saturated at a rather high photon flux density of around 1200 E m^–2 s^–1. Upon transition from far-red light to darkness, the oxidized reaction center P700⁺ of PS I was re-reduced very slowly in control leaves (with a half time t_1/2 higher than 500 ms), as measured by leaf absorbance changes at around 820 nm. Heat stress caused a spectacular acceleration of the postillumination P700⁺ reduction, with t_1/2 falling to a value lower than 50 ms (after leaf exposure to 48°C). The decreased t_1/2 was sensitive to HgCl₂ and insensitive to diuron, methyl viologen (an electron acceptor of PS I competing with the endogenous acceptor ferredoxin) and anaerobiosis. This acceleration of the P700⁺ reduction was very rapidly induced by heat treatment (within less than 5 min) and persisted even after prolonged irradiation of the leaves with far-red light. After heat stress, the plastoquinone pool exhibited reduction in darkness as indicated by the increase in the apparent Fo level of chlorophyll fluorescence which could be quenched by far-red light. Application (for 1 min) of far-red light to heat-pretreated leaves also induced a reversible quenching of the maximal fluorescence level Fm, suggesting formation of a pH gradient in far-red light. Taken together, the presented data indicate that PS I responded to the heat-induced loss of PS II photochemical activity by catalyzing an electron flow from stromal reductants. Heat-stress-induced PS I electron transport independent of PS II seems to constitute a protective mechanism since block of this electron pathway in anaerobiosis was observed to result in a dramatic photoinactivation of PS I.Abbreviations PFD photon flux density - PS Photosystem - Apt and Aox amplitude of the photothermal and photobaric components of the photoacoustic signal, respectively - P700 reaction center pigment of PS I - Fo and Fm initial and maximal levels of chlorophyll fluorescence, respectively - Fv=Fm Fo-variable chlorophyll fluorescence - Q_A primary (stable) electron acceptor of PS II - DCMU (diuron) 3-(3,4-dichlorophenyl)-1,1-dimethylurea - Cyt cytochrome     

Simple method for isolation of oxygen evolving photosystem 2 core complex free of cytochrome <Emphasis Type="Italic">f</Emphasis> contamination

This communication presents a simple method for isolation of oxygen evolving photosystem 2 (PS 2) core complex by solubilisation of PS 2 membranes with the nonionic detergent octyl-β-D-thioglucopyranoside (OTG). This complex was free of cytochrome (Cyt) f contamination and also lacked the 22 and 10 kDa proteins that may not be directly required for primary photochemistry of the PS 2 complex and water oxidation. OTG could also remove the Cyt f contamination from the PS 2-core complex isolated using octyl-β-D-glucopyranoside (OGP). The Cyt f contamination in the PS 2 complexes could potentially interfere with spectrophotometric determinations of redox states of Cyt _b559 and its stoichiometry in the PS 2 complex.     

Evidence for an association of the early light-inducible protein (ELIP) of pea with photosystem II

The precursor to the nuclear-coded 17 kDa early light-inducible protein (ELIP) of pea has been transported into isolated intact chloroplasts. The location of the mature protein in the thylakoid membranes was investigated after using cleavable crosslinkers such as DSP and SAND in conjunction with immuno-fractionation methods and by application of mild detergent fractionation. We show that ELIP is integrated into the membranes via the unstacked stroma thylakoids. After isolation of protein complexes by solubilization of membranes with Triton X-100 and sucrose density-gradient centrifugation the crosslinked ELIP comigrates with the PS II core complex. Using SAND we identified ELIP as a 41–51 kDa crosslinked product while with DSP four products of 80 kDa, 70 kDa, 50–42 kDa and 23–21 kDa were found. The immunoprecipitation data suggested that the D₁-protein of the PS II complex is one of the ELIP partners in crosslinked products.Abbreviations chl chlorophyll - D₁ herbicide-binding protein - DSP dithiobis-(succinimidylpropionate) - ELIP early light-inducible protein - LHC I and LHC II light-harvesting chlorophyll a/b complex associated with photosystem I or II - PAGE polyacrylamide gel electrophoresis - poly(A)-rich RNA polyadenyd mRNA - PS I and PS II photosystems I and II - SAND sulfosuccinimidyl 2-(m-azido-o-nitro-benzamido)-ethyl-1,3-dithiopropionate - Triton X-100 octylphenoxypolyethoxyethanol     

Regulation of Electron Transport Composition in Cyanobacterial Photosynthetic System: Stoichiometry among Photosystem I and II Complexes and Their Light-Harvesting Antennae and Cytochrome b6/fComplex

This study was done to confirm our previous observation withthe pattern of changes in electron transport composition inducedby an imbalance of the electron transport state. Contents ofphotosystem (PS) I and II complexes and their antennae and Cytb₆/f complex were determined for systems of cyanobacterium SynechocystisPCC 6714 of different PS I/PS II ratios. The results indicatedthat (1) the observed changes in the PS I/PS II ratio are not-dueto regulation of the activities of the respective PS's but tochanges in their contents, (2) the molar ratio between PS IIand Cyt b₆/f complexes was fairly constant when marked changesoccurred in the PS I content, and (3) the PS II and Cyt b₆/fcontents per cell remained fairly constant while the PS I contentchanged markedly. These findings agree with our previous observationwith autotrophic cells of Anacystis nidulans Tx 20 and supportour argument that in cyanobacterial and red algal electron transportsystems, the content of the terminalcomponent(s), such as PSI complex, is regulated in order to maintain a balance betweenthe electron influx by PS II action to the system and the effluxby PS I action from it. (Received June 3, 1987; Accepted September 20, 1987)     

Regulation of the Stoichiometry of Thylakoid Components in the Photosynthetic System of Cyanophytes: Model Experiments Showing that Control of the Synthesis or Supply of Chl a can Change the Stoichiometric Relationship between the Two Photosystems

Regulation of the assembly of the photosystem I (PS I) complexin response to the light regime in the photosynthetic systemof cyanophytes was studied in Synechocystis PCC 6714. The relationshipbetween the assembly of the PS I complex and synthesis of Chla was examined by model experiments in which synthesis of Chla was controlled by two inhibitors, gabaculine (GAB) and 2,2'-dipyridyl(DP). Both inhibitors caused a change to a lower ratio of PSI to PS II even under light that normally induces a high ratioof PS I to PS II. The change in stoichiometry induced by theseinhibitors was suppressed when protein synthesis was inhibitedby chloram-phenicol, similarly to the change in the stoichiometryinduced by light that excites mainly PS I (PS I light). Comparisonof the levels of PS I, PS II and Cyt b₆-f complexes per cellindicated that a selective suppression of the assembly of thePS I complex was induced by the inhibitors: the stoichiometricrelationship among PS I, PS II and Cyt b₆-f complexes was identicalto that induced by PS I light or white light of high intensity.GAB induced a decrease in size of the phycobilisome also, whileDP did not, similarly to PS I light. The results indicate thatthe ratio of PS I to PS II can be changed by the control ofsynthesis of Chl a. They also suggest that control of the synthesisor supply of Chl a probably exerted at site(s) in or after theprocess of the Mg-protoporphyrin branch, is involved in themechanism of regulation of the assembly of the PS I complexin cyanophytes. (Received September 7, 1989; Accepted November 20, 1989)     

Steady State of Photosynthetic Electron Transport in Cells of the Cyanophyti Synechocystis PCC 6714 Having Different Stoichiometry between PS I and PS II: Analysis of Flash-Induced Oxidation-Reduction of Cytochrome f and P700 under Steady State of Photosynthesis

The steady state of photosynthetic electron transport drivenby two photosystems was studied with cells of the cyanophyteSynechocystis PCC 6714 by analyzing the flash-induced oxidation-reductionof Cyt f and P700 under continuous background illumination.We first analyzed the spectra and the kinetics of flash-inducedabsorption changes in the 400 to 440 nm wavelength region anddefined the absorption changes due to oxidation-reduction ofCyt f and P700. Results indicated that the flash-induced absorptionchanges at 420 and 435 nm are due to the oxidation-reductionof Cyt f and P700, respectively. Determination of the steadystate of Cyt f (420 nm) and P700 (435 nm) was made for the cellsgrown under a weak orange light exciting mainly PS II (PS IIlight) and having a high ratio of PS I to PS II (PS I/PS II),and those grown under a weak red light exciting preferentiallyPS I (PS I light) and having a low PS I/PS II. The steady stateof electron transport in cells of the two types were comparedunder PS I and PS II lights. The results indicated that: (1)under the light conditions used for growth (both red and orangelight), the intermediate electron pool between the two photosystemsremained in a redox state so as to keep both photosystems inthe open state. (2) When shifted to PS I light, the intermediatepool and PS I in cells of high PS I/PS II became extremely electron-poor,and so most of the PS I reaction centers were closed. (3) Theintermediate pool in cells of low PS I/PS II became extremelyelectron-rich when shifted to PS II light, and most of the PSII reaction centers were closed. The electron transport stateis released from such biased states by regulation of PS I/PSII. Results supported our previously proposed hypothesis thatthe stoichiometry between PS I and PS II is regulated so asto keep the two photosystems in the open state. The relationshipbetween the steady state of electron transport and the regulationof PS I/PS II is discussed. (Received August 2, 1990; Accepted December 10, 1990)     

Effect of dehydration on light-induced reactions in photosystem II: photoreactions of cytochrome b559

The effect of dehydration on the reaction pattern of photosystem II (PS II) has been studied by measuring and analyzing spectral changes induced by continuous wavelength illumination in films of untreated and hydroxylamine-washed PS II membrane fragments dehydrated to different levels. The obtained data revealed (i) the extent of light-induced formation of about one Q(A)(-*)per 230 chlorophylls (Chl) remains virtually invariant to dehydration down to the lowest values of relative humidity (6-8% RH); (ii) a decrease of the RH to 30% leads to severe blockage of the electron transfer from Q(A)(-*) to Q(B) and the progressive replacement of water oxidation by photooxidation of high potential (HP) cytochrome (Cyt) b559 in untreated PS II samples or accessory Chl and carotenoid (Car) molecules in samples with preoxidized Cyt b559; (iii) photooxidation of Cyt b559 is followed by its photoreduction, concomitant with photooxidation of Chl and Car; (iv) in dry samples with preoxidized Cyt b559, not more than a half of total Cyt b559 can be photochemically reduced, independent of the extent of Cyt b559 in the HP form; (v) at low RH values, Cyt b559 photoreduction in samples with preoxidized heme groups and photoaccumulation of Q(A)(-*) take place with biphasic kinetics with similar rate constants for both processes; (vi) Cyt b559 photoreduction in dry samples is DCMU insensitive, while the dark rereduction of photooxidized Cyt b559 is inhibited by DCMU; (vii) fast and slow kinetic phases of Cyt b559 photoreduction dramatically differ in their dependencies on the intensity of CW illumination and are associated with electron donation to Cyt b559 from Q(A)(-*) and pheophytin(-*), respectively. The pathways of light-induced electron transfer in PS II involving Cyt b559 are discussed.     

Effects of natural shade on soybean thylakoid membrane composition

The effect of natural shade on chloroplast thylakoid membrane activity and composition was examined for soybean (Glycine Max. cv. Young) grown under field conditions. Plots with high (10 plants m^–1 row) or low (1 plant m^–1 row) plant density were established. Expanding leaves were tagged at 50, 58 and 65 days after planting (DAP). At 92 DAP, tagged leaves were used as reference points to characterize canopy light environments and isolate thylakoid membranes. Light environments ranged from a photosynthetic photon flux density (PPFD) of 87% of full sun to a PPFD of 10% of full sun. The decline in PPFD was accompanied by an increase in the far-red/red (735 nm/645 nm) ratio from 0.9 to approximately six. The major effects of shade on chloroplast thylakoid membranes were a reduction in chloroplast coupling factor and a shift in light-harvesting capacity from Photosystem I to Photosystem II. Photosynthetic electron transport capacity was not affected by differences in PPFD, but was 20 to 30% higher in the 1 plant m^–1 row treatment. The plant density effect on electron transport was associated with differences in plastocyanin concentration, suggesting that plastocyanin is a limiting factor in soybean. Shade did not have a significant effect on the concentration of Photosystem II, Cyt b₆f, or Photosystem I complexes.Abbreviations CF₁ chloroplast coupling factor - DAP days after planting - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCIP 2,6-dichlorophenolindophenol - FR/R far-red/red - PBS 10 mM sodium phosphate (pH 7.0), 150 mM NaCl - PPFD photosynthetic photon flux density - PS I Photosystem I - PS II Photosystem II - P700 reaction center of Photosystem I - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - TBS 20 mM Tris-HCl (pH 7.5), 500 mM NaCl - TTBS 20 mM Tris-HCl (pH 7.5), 500 mM NaCl, 0.05% (w/v) polyoxyethylenesorbitan monolaurate (Tween-20) The US Government right to retain a non-exclusive, royalty-free licence in and to any copyright is acknowledged.The US Government right to retain a non-exclusive, royalty-free licence in and to any copyright is acknowledged.