PARTICIPATION OF THE TWO PHOTOSYSTEMS IN LIGHT DEPENDENT HYDROGEN EVOLUTION IN Scenedesmus obliquus

Abstract— The H₂-photoproduction in the presence of dithionite measured in wild type and mutant cells of Scenedesmus obliquus demonstrates two sequential phases. In mutants showing only PS I activity phase 1 of H₂-photoproduction is visible with its core activity. When PS II is developed during greening, considerable activity is added to the core of phase I and phase II activity appears. Addition of DCMU reduces H₂-photoproduction by about 90%. The residual activity is completely attributed to the core of phase I. It was concluded that the core of phase I is dependent upon PS I only and can use sources different from water as electron donors. Phase II is dependent upon the capacity of PS II, a functioning photosynthetic apparatus and water as electron donor. The results are supported by studies of wavelength dependent activity of the two separate phases of H₂-photoproduction.     

Thylakoid-associated polyamines adjust the UV-B sensitivity of the photosynthetic apparatus by means of light-harvesting complex II changes

The sensitivity of the photosynthetic apparatus to ultraviolet-B (UV-B) irradiation was studied in cultures of unicellular green alga Scenedesmus obliquus incubated in low light (low photosynthetically active radiation intensity [LL]) and high light (high photosynthetically active radiation intensity [HL]) conditions, treated or not with exogenous polyamines. Biochemical and physicochemical measurements showed that UV-B radiation induces a decrease in the thylakoid-associated putrescine (Put) and an increase in spermine (Spm), so that the reduction of Put/Spm ratio leads to the increase of light-harvesting complex II (LHCII) size per active reaction center and, consequently, the amplification of UV-B effects on the photosynthetic apparatus. The separation of oligomeric and monomeric forms of LHCII from isolated thylakoids showed that UV-B induces an increase in the oligomeric forms of LHCII, which was more intense in LL than in HL. By manipulating the LHCII size with exogenous polyamines, the sensitivity degree of the photosynthetic apparatus to UV-B changed significantly. Specifically, the addition of Put decreased highly the sensitivity of LL culture to UV-B because of the inhibitory effect of Put on the LHCII size increasing, whereas the addition of Spm enhanced the UV-B injury induced in HL culture because of the increasing of LHCII size. The ability of the photosynthetic apparatus to recover the UV-B induced changes was also investigated.     

KINETICS OF PHOTOSYNTHETIC APPARATUS ADAPTATION IN Scenedesmus obliquus TO CHANGE IN IRRADIANCE AND LIGHT QUALITY

Abstract— Cells of the unicellular green alga Scenedesmus obliquus grown under high (20 W m^-2) or low (5 W m^-2) irradiancies of white light show all characteristics of sun or shade plants, respectively. When transferred to alternate light conditions, the cells adapt within 6 h. When cells grown under high irradiance of white light are transferred to red (683 nm) or blue (424 nm) light, they show characteristics similar to cells adapted to low or high irradiancies of white light, respectively. This adaptation to different wavelengths takes about 12 h. The underlying changes in the photosynthetic apparatus are discussed.     

Photosystem I/molecular wire/metal nanoparticle bioconjugates for the photocatalytic production of H2

Photosystem I (PS I) is a robust photosynthetic complex that adeptly captures photons to create a charge-separated state with a quantum efficiency that approaches 1.0. This charge-separated state is stable for approximately 100 ms, and the low-potential reductant that is produced is poised at a redox potential favorable for H2 evolution. PS I has been covalently linked to Pt and Au nanoparticle surfaces by 1,6-hexanedithiol which serves as a molecular wire to both connect PS I to the particles and transfer electrons from the terminal electron transfer cofactor of PS I, FB, to the nanoparticle. Illumination of these Photosystem I/molecular wire/nanoparticle bioconjugates is able to catalyze the reaction: 2H+ + 2e(-)--> H2. Transfer of the electrons from PS I to the nanoparticle through the molecular wire is not rate-limiting for H2 evolution. Supplying the system with more efficient donor-side electron donating species results in a 5-fold increase in the rate of H2 evolution.     

Antagonistic blue- and red-light regulation of cab-gene expression during photosynthetic adaptation in Scenedesmus obliquus.

During adaptation of the photosynthetic apparatus of the green alga Scenedesmus obliquus to various light qualities, the accumulation of chlorophylls and pigment-protein complexes (with specific consideration of chlorophyll a/b-binding (Cab) proteins) and cab-gene expression were determined. The fluence rate dependences for chlorophyll accumulation and cab-gene expression were very different. Very low fluence rates of violet (404 nm), blue (461 nm) and red (650 nm) light below the photosynthetic threshold, i.e. between 10(-3) and 10(-1) mumol m-2 s-1, inhibited all of these reactions in cells grown under heterotrophic conditions. At elevated fluence rates (above 1 mumol m-2 s-1), red light retained its negative regulation, whereas blue light stimulated pigment accumulation. Under autotrophic conditions the pattern was more complex, because chlorophyll accumulation was unaffected by light below the photosynthetic threshold. However, the expression of cab-genes was inhibited by red light but stimulated by blue light. Cells adapted to fluence rates, which ensured photosynthetic energy supply (above 1 mumol m-2 s-1), showed an increase in chlorophyll accumulation, blue light being more effective than red light. The results confirm and extend our previous discovery of two antagonistically acting photoreceptors in Scenedesmus which mediate and coordinate the complex functional and structural changes associated with photosynthetic adaptation. One of these receptor pigments is a blue-light receptor with positive action; the other is a violet-red-light receptor which can operate far below the photosynthetic threshold and exerts a negative regulation.     

THE ROLE OF CYCLIC ELECTRON FLOW AROUND PHOTOSYSTEM I and EXCITATION ENERGY DISTRIBUTION BETWEEN THE PHOTOSYSTEMS UPON ACCLIMATION TO HIGH IONIC STRESS IN Dunaliella salina

Abstract— The distribution of excitation energy between the two photosystems in the halophylic alga Dunaliella salina has been analyzed under ionic stress. In the transition from state 1 to state 2, it was found that a, the absorption cross-section of photosystem (PS) I increased from 42 to 49% until an equal distribution between PS I and PS II was obtained in state 2. Acclimation of the algae to different salt concentrations did not change the fractions of light absorbed in PS II and PS I, but slowed down the transition time from state 1 to state 2. A large increase in ΔpH induced fluorescence quenching was observed which was abolished by the uncoupler nigericin. Photoacoustic quantum yield spectra of energy storage indicated a larger energy storage at 700 nm induced upon stress. The additional ΔpH quenching of fluorescence and the additional quantum yield of energy storage at 700 nm, in the stressed algae, are consistent with the operation of a cyclic, energy-storing pathway in PS I which is uncoupler sensitive.     

Detergent-protein interactions in aqueous buffer suspensions of Photosystem I (PS I)

Systematic and uniform monolayer formation of Photosystem I (PS I) onto self-assembled monolayer (SAM) substrates to enable unidirectional electron transfer is crucial for its successful use in the fabrication of bio-hybrid solid-state electronic or photovoltaic devices. Yet, our recent studies (Mukherjee et al., 2010) indicate that surface self-assembly of PS I from aqueous buffer suspensions onto alkanethiolate SAM/Au substrates frequently leads to complex columnar structures due to solution phase protein aggregations. We investigate the effect of two prototypical non-ionic detergents, n-Dodecyl-β-D-Maltoside (DM) and Triton X-100 (TX-100), on protein-protein interactions via the protein-detergent interfacial chemistry. Dynamic light scattering (DLS) experiments are used to demonstrate the impact of relative protein/detergent concentrations on aggregation dynamics of PS I suspensions. In turn, the surface attachment characteristics of PS I adsorbed from the aforementioned suspensions onto SAM/Au substrate is examined by atomic force (AFM) microscopy. Our results indicate that relative concentration of PS I and detergents (DM or, TX-100) with respect to their critical micelle concentrations (CMC) determines the extent of self-association between PS I complexes driven by the screening induced by detergent micelles and/or, inter-protein distances. Such interfacial phenomena during the PS I-detergent complexation process drives the colloidal system through various regimes of phase separations, suspension and/or, de-aggregation, wherein individual PS I complexes can exist in a frustrated state that prevents favorable orientations for PS I-PS I interactions. The present study presents a novel strategy, heretofore not considered, for tailoring inter-protein distances and protein-protein interactions in solution phase, thereby allowing a superior control on the surface attachment of PS I onto SAM/Au substrates.     

Cytochrome b6/f complex as an indigenous photodynamic generator of singlet oxygen in thylakoid membranes

Possible association of photodynamic sensitization by cytochrome b6/f complex (cyt b6/f) via singlet oxygen (1O2) mechanism with photoinhibition damage to photosystem II (PS II) was studied using such subthylakoid preparations as photosystem I (PS I) particles, PS II core complex and cyt b6/f from spinach leaves. Upon exposure to bright light, PS II core complex lost photosynthetic electron transport activity to a certain extent, whose-spectral dependence implied that pheophytin a is likely involved in photoinactivation of PS II core complex in itself. The presence of PS I particles exerted virtually no effect on PS II core photoinactivation. However, the inclusion of cyt b6/f in samples resulted in a marked exacerbation of the photoinactivation, particularly in UV-A and blue light. Such effect of cyt b6/f was suppressed by azide and enhanced by the medium deuteration. Photogeneration of 1O2 from cyt b6/f was confirmed by ESR and spectrophotometry, chemically trapping 1O2. Action spectra for both 1O2 photoproduction and PS II core photoinactivation by cyt b6/f bore a close resemblance to each other, seemingly carrying the absorption characteristics of the Rieske Fe-S protein. A complex deficient in the Rieske protein prepared from intact cyt b6/f showed virtually no generation of 1O2 in light, whereas an efficient photoformation of 1O2 was seen in the Rieske protein preparation. The results suggest that cyt b6/f, rather specifically the Rieske center, may play a prominent role in photoinhibition processes through type II photosensitization in thylakoids.     

KINETIC STUDIES ON THE STABILIZATION OF THE PRIMARY RADICAL PAIR P680+ Pheo- IN DIFFERENT PHOTOSYSTEM II PREPARATIONS FROM HIGHER PLANTS*

Abstract— The stabilization of the primary radical pair P680+ pheophytin (Pheo)^- through rapid electron transfer from Pheo to the special plastoquinone of photosystem II (PS II), Q_A, was analyzed on the basis of time-resolved (40 ps) UV-absorption changes detected in different PS II preparations from higher plants. Lifetime measurements of¹Chi* fluorescence by single photon counting and a numerical analysis of the redox reactions revealed (1) at exciton densities required for light saturation of the stable charge separation, annihilation processes dominate the excited state decay leading to very similar lifetimes of ¹Chi* in systems with open and closed reaction centers and (2) the difference of absorption changes induced by actinic flashes of comparatively high photon density in samples with open and photochemically closed reaction centers, respectively, provides a suitable measure of the rate constant of QA formation. Conclusion 2 was confirmed in PS II membrane fragments by measurements at three wavelengths (280 nm, 292 nm and 325 nm) where the difference spectrum of Q^-_A formation exhibits characteristic features. The numerical evaluation of the experimental data led to the following results: (1) the rate constant of Q^-_A formation was found to be (300 ± 100 ps)^-1 in PS II membrane fragments and PS II core complexes deprived of the distal and proximal antenna and (2) an iron depletion treatment of membrane fragments does not affect these kinetics. The implications of these results are briefly discussed in terms of the PS II reaction pattern.     

A STUDY OF CHLOROPHYLL FLUORESCENCE IN VIVO BY A FLOW METHOD

Abstract— A flow method has been devised for the study of fluorescence transients during the induction period of photosynthetic systems. The principle of transformation of time-transients into space-transients is quantitatively worked out and the characteristics and mode of operation of the apparatus is described and illustrated with experimental results.
With this method spectroscopic and kinetic investigation of in vivo chlorophyll fluorescence have been undertaken. The fluorescence emission can be split into two componmts—a variable part and a constant part—with different spectral distributions. The excitation spectrum of the light induced fluorescence changes shows that the variable part is sensitized by system 11, whereas the constant part mainly sensitized by system II has a small component originating from system I. The light-induced fluorescence change decays in darkness with half-time of about 10 ^-2 s; this decay is highly sensitive to the action of inhibitors of oxygen evolution.
The flow method can be extended to measurements of oxygen, luminescence, absorption differences. It permits using high-intensity analytic beams with no actinic action; it is well suited for the integration of low level signals.     

Changes in the energy distribution in mutant thylakoid membranes of pea with modified pigment content. II. Changes due to magnesium ions concentration

Low-temperature (77K) steady-state chlorophyll fluorescence emission spectra, room temperature fluorescence and light scattering of thylakoid membranes isolated from pea mutants were studied as a function of Mg2+ concentration. The mutants have modified pigment content and altered structural organization of the pigment-protein complexes, distinct surface electric properties and functions. The analysis of the 77K emission spectra revealed that Mg2+-depletion of the medium caused not only an increased energy flow toward photosystem I in all investigated membranes but also changes in the quenching of the fluorescence, most probably by internal conversion. The results indicated that the macroorganization of the photosynthetic apparatus of mutants at supramolecular level (distribution and segregation of two photosystems in thylakoid membranes) and at supermolecular level (stacking of photosystem II supercomplexes) required different Mg ion concentrations. The data confirmed that the segregation of photosystems and the stacking of thylakoid membranes are two distinct phenomena and elucidated some features of their mechanisms. The segregation is initiated by changes in the lateral microorganization of light harvesting complexes II, their migration (repulsion from photosystem I) and subsequent separation of the two photosystems. Most likely 3D aggregation and formation of macrodomains, containing only photosystem II antenna complexes, play a certain precursory role for the increasing degree of the membrane stacking and the energy coupling between the light harvesting complexes II and the core complexes of photosystem II in the frame of photosystem II supercomplexes.     

PHOTOTHERMAL DEFLECTION SPECTROSCOPY OF PHOTOSYNTHETIC PIGMENTS in vivo and in vitro

Abstract— This Technical Note describes the design of a photothermal beam deflection apparatus which allows the easy and rapid measurement of thermal dissipation of absorbed light energy in various photosynthetic materials including whole plant leaves. This system is based on the "mirage effect" in which the refractive index gradient induced in a fluid in contact with the sample, irradiated with an intensity-modulated light, causes the periodic deflection of a laser beam parallel to the sample surface. The deflection of the probe laser beam is detected by a position sensor, the output of which is processed by a lock-in amplifier. Photothermal deflection signals can be monitored in vivo in intact leaves placed in various (liquid or gaseous) environments with a satisfactory signal-to-noise ratio between 100 (in water) and 50 (in air) at low modulation frequencies (ca 30 Hz). It is shown that this new and simple photothermal technique is a very sensitive tool for the measurement of absorption spectra of photosynthetic pigments both in vivo (leaves, algae or chloroplasts) and in model systems (Langmuir-Blodgett and solid films of chlorophyll).     

光合放氧复合物结构及其放氧机理的研究

光合水氧化是地球上最重要的生化过程之一.光合放氧生物包括光系统Ⅰ(PSⅠ)和光系统Ⅱ(PSⅡ)两种类型反应中心,光系统Ⅱ反应中心能以水作为电子给体,利用光能氧化水产生质子和氧气.对于水如何被氧化这个难题前人已做了大量的工作,但到目前为止放氧复合物(OEC)的结构及水氧化的机理仍不清楚.本文结合当前研究结果,就光合放氧复合物的结构及光合放氧机理进行了综述,希望能有助于推进这方面的工作.     

Effects of changes of irradiance on the pigment composition of Gracilaria tenuistipitata var. liui Zhang et Xia

In plants, excess irradiation can damage the photosynthetic apparatus, although some protective mechanisms exist. The excess energy can be dissipated as thermal energy, and pigments (i.e., carotenoids) also play an important role in protecting the photosynthetic apparatus by epoxidating reactions. Chromatographic analysis of pigment extracts of Gracilaria tenuistipitata shows that zeaxanthin is the major carotenoid in this alga, accounting for up to 82% of total carotenoids. Short-term (55 h) and long-term (10 days) response of the pigments shows that Chl a, β-carotene and zeaxanthin degradation after light increase follows negative exponential trends, while the response of biliproteins is almost linear. Decreasing the irradiance results in a clear saturating response of the synthesis of Chl a and β-carotene after one to two days. Biliprotein synthesis displays a double linear trend, the first one lasting for four days in the cases of both R-phycoerythrin (RPE) and R-phycocyanin (RPC). The response of zeaxanthin is always faster than that of Chl a or biliproteins to changes of irradiance. Our results might indicate the presence of two pools of zeaxanthin in this alga, with different acclimation responses to the changes in the photon flux density.     

Reversed-phase HPLC determination of chlorophyll a' and naphthoquinones in photosystem I of red algae: existence of two menaquinone-4 molecules in photosystem I of Cyanidium caldarium.

Chlorophyll (Chl) a', the C13(2)-epimer of Chl a, is one of the two Chl molecules constituting the primary electron donor (P700) of photosystem (PS) I of a thermophilic cyanobacterium Synechococcus elongatus. To examine whether PS I of other oxygenic photosynthetic organisms in general contain one Chl a' molecule in P700, the pigment composition of thylakoid membranes and PS I preparations isolated from red algae Porphyridium purpureum and Cyanidium caldarium was examined by reversed-phase HPLC with particular attention to Chl a' and phylloquinone (PhQ), the secondary electron acceptor of PS I. The two red algae contained one Chl a' molecule at the core part of PS I. In PS I of C. caldarium, two menaquinone-4 (MQ-4) molecules were detected in place of PhQ used by higher plants and cyanobacteria. The 1:2:1 stoichiometry among Chl a', PhQ (MQ-4) and P700 in PS I of the red algae indicates that one Chl a' molecule universally exists in PS I of oxygenic photosynthetic organisms, and two MQ-4 molecules are associated with PS I of C. caldarium.     

生物光化学

生物光化学研究光在动植物体内所引起的生化现象。例如:经过各种不同波长的光辐照后的生命现象,生长规律,某些生理和病理过程,疾病的产生和治疗机理,细胞的辐射损伤和自然防御,以及光合色素在生物进化中的作用等。本文就光引起的现象:视觉、生物钟(光周期性)、植物的光合作用、辐射损伤及其修复、牛皮癣的治疗、新生儿黄疸病的治疗机理,以及光合色素——藻胆蛋白等七种现象,做了综述性的介绍。     

Polymer-Modified Single-Walled Carbon Nanotubes Affect Photosystem II Photochemistry,Intersystem Electron Transport Carriers and Photosystem I End Acceptors in Pea Plants

Single-walled carbon nanotubes (SWCNT) have recently been attracting the attention of plant biologists as a prospective tool for modulation of photosynthesis in higher plants. However, the exact mode of action of SWCNT on the photosynthetic electron transport chain remains unknown. In this work, we examined the effect of foliar application of polymer-grafted SWCNT on the donor side of photosystem II, the intersystem electron transfer chain and the acceptor side of photosystem I. Analysis of the induction curves of chlorophyll fluorescence via JIP test and construction of differential curves revealed that SWCNT concentrations up to 100 mg/L did not affect the photosynthetic electron transport chain. SWCNT concentration of 300 mg/L had no effect on the photosystem II donor side but provoked inactivation of photosystem II reaction centres and slowed down the reduction of the plastoquinone pool and the photosystem I end acceptors. Changes in the modulated reflection at 820 nm, too, indicated slower re-reduction of photosystem I reaction centres in SWCNT-treated leaves. We conclude that SWCNT are likely to be able to divert electrons from the photosynthetic electron transport chain at the level of photosystem I end acceptors and plastoquinone pool in vivo. Further research is needed to unequivocally prove if the observed effects are due to specific interaction between SWCNT and the photosynthetic apparatus.     

THE PHOTOSYNTHETIC APPARATUS OF Acetabularia mediterranea GROWN UNDER RED OR BLUE LIGHT. BIOPHYSICAL QUANTIFICATION and CHARACTERIZATION OF PHOTOSYSTEM II and ITS CORE COMPONENTS

Abstract— The photosynthetic activity of white light-grown Acetabularia mediterranea Lamouroux (= A. acetabulum (L.) Silva) decreases under continuous red light to less than 20% within 3 weeks. Subsequent blue light reactivates photosynthesis within a relatively short period of 3 days. In a former publication (Wennicke and Schmid, Plant Physiol. 84 ,1252–1256, 1987) we have shown that the regulated rate limiting step, which is an immediate light driven reaction, is part of photosystem II (PS II). The following biophysical properties of PS II were analyzed in thylakoids isolated from algae grown 3 weeks under either blue or red light with or without subsequent 3 days of blue light illumination: (a) fluorescence induction in the short time domain dominated by Q_A reduction, (b) the slow fluorescence decline reflecting pheophytin photoaccumulation, (c) absorption changes at 320 and 830 nm under repetitive flash excitation as indicator for the turnover of Q_A and P₆₈₀, respectively, (d) oscillation pattern of the oxygen yield by a flash train in dark adapted samples and (e) the binding capacity for atrazine. None of these PS II functions were severely affected, but a minor impairment of20–30% was observed in the thylakoids from algae grown for 3 weeks in red irradiation. The changes do not fully account for the drastic reduction of the electron transport through PS II which was 80% after red light treatment. Therefore, the regulated rate-limiting step appears to not be mainly located in the PS II core complex itself. It seems likely that the regulation process predominantly comprises the antenna system.     

The mechanism of ultrasonic improvement of weld line strength of injection molded polystyrene and polystyrene/polyethylene blend parts

The effect of ultrasonic oscillations and ultrasonic oscillation‐induced modes on weld line strength of polystyrene(PS) and polystyrene/polyethylene(PS/HDPE) blend was investigated. And the mechanism of ultrasonic improvement of weld line strength of PS and PS/HDPE blend was also studied. The presence of ultrasonic oscillations can enhance the weld line strength of PS and PS/HDPE blend. Compared with mode I(ultrasonic oscillations were induced into mold at the whole process of injection molding), the induced ultrasonic oscillations as mode II(ultrasonic oscillations were induced into mold after injection mold filling) is more effective to increase weld line strength of PS and PS/HDPE blend. The mechanism for ultrasonic improvement of weld line strength of PS and PS/HDPE blend is that the ultrasonic oscillations can improve the molecular diffusion across weld line of the melt at the core, and make against the fusion of melt at the skin. © 2006 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 44: 1520–1530, 2006     

Photoinhibition of photosystem I is accelerated by dimethyldithiocarbamate, an inhibitor of superoxide dismutase, during light-chilling of spinach leaves

In vivo photoinhibition of photosystem I (PS I) was investigated at chilling temperature using the leaves of the chilling-resistant spinach plant treated with an inhibitor of superoxide dismutase, diethyldithiocarbamate (DDC). When spinach leaves were treated with DDC during chilling at 4 degrees C for 12 h with a light intensity of 120 micromol m(-2) s(-1), the activity of PS I and the content of iron-sulfur centers declined to about 50% and 25% of the non-DDC-treated controls, respectively. A native green gel analysis of thylakoid membranes isolated from the DDC-treated leaves resolved a novel chlorophyll-protein complex, which was identified as the light-harvesting complex I (LHC I)-deficient PS I complex when examined by 77 K fluorescence spectroscopy and two-dimensional sodium dodecyl sulfate gel electrophoresis. The possible dissociation of LHC I as an early structural change in the PS I complex after DDC-induced photoinhibition of PS I is discussed.