RESUMEN
The largest light-harvesting antenna in nature, the chlorosome, is a heterogeneous helical BChl self-assembly that has evolved in green bacteria to harvest light for performing photosynthesis in low-light environments. Guided by NMR chemical shifts and distance constraints for Chlorobaculum tepidum wild-type chlorosomes, the two contrasting packing modes for syn-anti parallel stacks of BChl c to form polar 2D arrays, with dipole moments adding up, are explored. Layered assemblies were optimized using local orbital density functional and plane wave pseudopotential methods. The packing mode with the lowest energy contains syn-anti and anti-syn H-bonding between stacks. It can accommodate R and S epimers, and side chain variability. For this packing, a match with the available EM data on the subunit axial repeat and optical data is obtained with multiple concentric cylinders for a rolling vector with the stacks running at an angle of 21° to the cylinder axis and with the BChl dipole moments running at an angle ß â¼ 55° to the tube axis, in accordance with optical data. A packing mode involving alternating syn and anti parallel stacks that is at variance with EM appears higher in energy. A weak cross-peak at -6 ppm in the MAS NMR with 50 kHz spinning, assigned to C-181, matches the shift of antiparallel dimers, which possibly reflects a minor impurity-type fraction in the self-assembled BChl c.
Asunto(s)
Chlorobi , Chlorobi/metabolismo , Complejos de Proteína Captadores de Luz/química , Complejos de Proteína Captadores de Luz/metabolismo , Espectroscopía de Resonancia Magnética , Modelos MolecularesRESUMEN
Chlorophyll fluorescence is a ubiquitous tool in basic and applied plant science research. Various standard commercial instruments are available for characterization of photosynthetic material like leaves or microalgae, most of which integrate the overall fluorescence signals above a certain cut-off wavelength. However, wavelength-resolved (fluorescence signals appearing at different wavelengths having different time dependent decay) signals contain vast information required to decompose complex signals and processes into their underlying components that can untangle the photo-physiological process of photosynthesis. Hence, to address this we describe an advanced chlorophyll fluorescence spectrometer - ChloroSpec - allowing three-dimensional simultaneous detection of fluorescence intensities at different wavelengths in a time-resolved manner. We demonstrate for a variety of typical examples that most of the generally used fluorescence parameters are strongly wavelength dependent. This indicates a pronounced heterogeneity and a highly dynamic nature of the thylakoid and the photosynthetic apparatus under actinic illumination. Furthermore, we provide examples of advanced global analysis procedures integrating this three-dimensional signal and relevant information extracted from them that relate to the physiological properties of the organism. This conveniently obtained broad range of data can make ChloroSpec a new standard tool in photosynthesis research.
Asunto(s)
Clorofila , Fotosíntesis , Espectrometría de Fluorescencia , Clorofila/metabolismo , Espectrometría de Fluorescencia/métodos , Espectrometría de Fluorescencia/instrumentación , Fotosíntesis/fisiología , Hojas de la Planta/metabolismo , Fluorescencia , Tilacoides/metabolismoRESUMEN
Chlorosomes, the photosynthetic antenna complexes of green sulfur bacteria, are paradigms for light-harvesting elements in artificial designs, owing to their efficient energy transfer without protein participation. We combined magic angle spinning (MAS) NMR, optical spectroscopy and cryogenic electron microscopy (cryo-EM) to characterize the structure of chlorosomes from a bchQ mutant of Chlorobaculum tepidum. The chlorosomes of this mutant have a more uniform composition of bacteriochlorophyll (BChl) with a predominant homolog, [8Ethyl, 12Ethyl] BChl c, compared to the wild type (WT). Nearly complete 13C chemical shift assignments were obtained from well-resolved homonuclear 13C-13C RFDR data. For proton assignments heteronuclear 13C-1H (hCH) data sets were collected at 1.2 GHz spinning at 60 kHz. The CHHC experiments revealed intermolecular correlations between 132/31, 132/32, and 121/31, with distance constraints of less than 5 Å. These constraints indicate the syn-anti parallel stacking motif for the aggregates. Fourier transform cryo-EM data reveal an axial repeat of 1.49 nm for the helical tubular aggregates, perpendicular to the inter-tube separation of 2.1 nm. This axial repeat is different from WT and is in line with BChl syn-anti stacks running essentially parallel to the tube axis. Such a packing mode is in agreement with the signature of the Qy band in circular dichroism (CD). Combining the experimental data with computational insight suggests that the packing for the light-harvesting function is similar between WT and bchQ, while the chirality within the chlorosomes is modestly but detectably affected by the reduced compositional heterogeneity in bchQ.
Asunto(s)
Bacterioclorofilas , Chlorobi , Chlorobi/genética , Chlorobi/metabolismo , Bacterioclorofilas/química , Mutación , Complejos de Proteína Captadores de Luz/química , Complejos de Proteína Captadores de Luz/metabolismo , Complejos de Proteína Captadores de Luz/genética , Microscopía por Crioelectrón , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismoRESUMEN
We establish a general kinetic scheme for the energy transfer and radical-pair dynamics in photosystem I (PSI) of Chlamydomonas reinhardtii, Synechocystis PCC6803, Thermosynechococcus elongatus and Spirulina platensis grown under white-light conditions. With the help of simultaneous target analysis of transient-absorption data sets measured with two selective excitations, we resolved the spectral and kinetic properties of the different species present in PSI. WL-PSI can be described as a Bulk Chl a in equilibrium with a higher-energy Chl a, one or two Red Chl a and a reaction-center compartment (WL-RC). Three radical pairs (RPs) have been resolved with very similar properties in the four model organisms. The charge separation is virtually irreversible with a rate of ≈900 ns-1. The second rate, of RP1 â RP2, ranges from 70-90 ns-1 and the third rate, of RP2 â RP3, is ≈30 ns-1. Since RP1 and the Red Chl a are simultaneously present, resolving the RP1 properties is challenging. In Chlamydomonas reinhardtii, the excited WL-RC and Bulk Chl a compartments equilibrate with a lifetime of ≈0.28 ps, whereas the Red and the Bulk Chl a compartments equilibrate with a lifetime of ≈2.65 ps. We present a description of the thermodynamic properties of the model organisms at room temperature.
Asunto(s)
Chlamydomonas reinhardtii , Complejo de Proteína del Fotosistema I , Clorofila A , Transferencia de Energía , CinéticaRESUMEN
In natural environments, plants are exposed to rapidly changing light. Maintaining photosynthetic efficiency while avoiding photodamage requires equally rapid regulation of photoprotective mechanisms. We asked what the operation frequency range of regulation is in which plants can efficiently respond to varying light. Chlorophyll fluorescence, P700, plastocyanin, and ferredoxin responses of wild-types Arabidopsis thaliana were measured in oscillating light of various frequencies. We also investigated the npq1 mutant lacking violaxanthin de-epoxidase, the npq4 mutant lacking PsbS protein, and the mutants crr2-2, and pgrl1ab impaired in different pathways of the cyclic electron transport. The fastest was the PsbS-regulation responding to oscillation periods longer than 10 s. Processes involving violaxanthin de-epoxidase dampened changes in chlorophyll fluorescence in oscillation periods of 2 min or longer. Knocking out the PGR5/PGRL1 pathway strongly reduced variations of all monitored parameters, probably due to congestion in the electron transport. Incapacitating the NDH-like pathway only slightly changed the photosynthetic dynamics. Our observations are consistent with the hypothesis that nonphotochemical quenching in slow light oscillations involves violaxanthin de-epoxidase to produce, presumably, a largely stationary level of zeaxanthin. We interpret the observed dynamics of photosystem I components as being formed in slow light oscillations partially by thylakoid remodeling that modulates the redox rates.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Proteínas del Complejo del Centro de Reacción Fotosintética , Transporte de Electrón , Complejo de Proteína del Fotosistema II/metabolismo , Luz , Fotosíntesis/fisiología , Arabidopsis/metabolismo , Clorofila/metabolismo , Complejos de Proteína Captadores de Luz/metabolismo , Mutación/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Proteínas de la Membrana/metabolismoRESUMEN
Non-photochemical quenching (NPQ) in photosynthetic organisms provides the necessary photoprotection that allows them to cope with largely and quickly varying light intensities. It involves deactivation of excited states mainly at the level of the antenna complexes of photosystem II using still largely unknown molecular mechanisms. In higher plants the main contribution to NPQ is the so-called qE-quenching, which can be switched on and off in a few seconds. This quenching mechanism is affected by the low pH-induced activation of the small membrane protein PsbS which interacts with the major light-harvesting complex of photosystem II (LHCII). We are reporting here on a mechanistic study of the PsbS-induced LHCII quenching using ultrafast time-resolved chlorophyll (Chl) fluorescence. It is shown that the PsbS/LHCII interaction in reconstituted proteoliposomes induces highly effective and specific quenching of the LHCII excitation by a factor ≥ 20 via Chl-Chl charge-transfer (CT) state intermediates which are weakly fluorescent. Their characteristics are very broad fluorescence bands pronouncedly red-shifted from the typical unquenched LHCII fluorescence maximum. The observation of PsbS-induced Chl-Chl CT-state emission from LHCII in the reconstituted proteoliposomes is highly reminiscent of the in vivo quenching situation and also of LHCII quenching in vitro in aggregated LHCII, indicating a similar quenching mechanism in all those situations. The PsbS mutant lacking the two proton sensing Glu residues induced significant, but much smaller, quenching than wild type. Added zeaxanthin had only minor effects on the yield of quenching in the proteoliposomes. Overall our study shows that PsbS co-reconstituted with LHCII in liposomes represents an excellent in vitro model system with characteristics that are reflecting closely the in vivo qE-quenching situation.
Asunto(s)
Proteínas de Arabidopsis/química , Arabidopsis/química , Complejos de Proteína Captadores de Luz/química , Complejo de Proteína del Fotosistema II/química , Proteolípidos/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Clorofila/química , Clorofila/metabolismo , Fluorescencia , Concentración de Iones de Hidrógeno , Complejos de Proteína Captadores de Luz/genética , Complejos de Proteína Captadores de Luz/metabolismo , Mutación , Complejo de Proteína del Fotosistema II/genética , Complejo de Proteína del Fotosistema II/metabolismo , Espectrometría de Fluorescencia , Tilacoides/química , Zeaxantinas/químicaRESUMEN
Light-harvesting complex II (LHCII) is the major antenna complex in higher plants and green algae. It has been suggested that a major part of the excited state energy dissipation in the so-called "non-photochemical quenching" (NPQ) is located in this antenna complex. We have performed an ultrafast kinetics study of the low-energy fluorescent states related to quenching in LHCII in both aggregated and the crystalline form. In both sample types the chlorophyll (Chl) excited states of LHCII are strongly quenched in a similar fashion. Quenching is accompanied by the appearance of new far-red (FR) fluorescence bands from energetically low-lying Chl excited states. The kinetics of quenching, its temperature dependence down to 4 K, and the properties of the FR-emitting states are very similar both in LHCII aggregates and in the crystal. No such FR-emitting states are found in unquenched trimeric LHCII. We conclude that these states represent weakly emitting Chl-Chl charge-transfer (CT) states, whose formation is part of the quenching process. Quantum chemical calculations of the lowest energy exciton and CT states, explicitly including the coupling to the specific protein environment, provide detailed insight into the chemical nature of the CT states and the mechanism of CT quenching. The experimental data combined with the results of the calculations strongly suggest that the quenching mechanism consists of a sequence of two proton-coupled electron transfer steps involving the three quenching center Chls 610/611/612. The FR-emitting CT states are reaction intermediates in this sequence. The polarity-controlled internal reprotonation of the E175/K179 aa pair is suggested as the switch controlling quenching. A unified model is proposed that is able to explain all known conditions of quenching or non-quenching of LHCII, depending on the environment without invoking any major conformational changes of the protein.
Asunto(s)
Clorofila/química , Complejos de Proteína Captadores de Luz/química , Clorofila/metabolismo , Cristalización , Transporte de Electrón , Fluorescencia , Cinética , Complejos de Proteína Captadores de Luz/metabolismo , Modelos Químicos , Modelos Moleculares , Teoría Cuántica , Relación Señal-Ruido , Espectrometría de Fluorescencia/métodos , Spinacia oleracea/química , TemperaturaRESUMEN
Nonphotochemical quenching (NPQ) is the process that protects the photosynthetic apparatus of plants and algae from photodamage by dissipating as heat the energy absorbed in excess. Studies on NPQ have almost exclusively focused on photosystem II (PSII), as it was believed that NPQ does not occur in photosystem I (PSI). Recently, Ballottari et al. [Ballottari M, et al. (2014) Proc Natl Acad Sci USA 111:E2431-E2438], analyzing PSI particles isolated from an Arabidopsis thaliana mutant that accumulates zeaxanthin constitutively, have reported that this xanthophyll can efficiently induce chlorophyll fluorescence quenching in PSI. In this work, we have checked the biological relevance of this finding by analyzing WT plants under high-light stress conditions. By performing time-resolved fluorescence measurements on PSI isolated from Arabidopsis thaliana WT in dark-adapted and high-light-stressed (NPQ) states, we find that the fluorescence kinetics of both PSI are nearly identical. To validate this result in vivo, we have measured the kinetics of PSI directly on leaves in unquenched and NPQ states; again, no differences were observed. It is concluded that PSI does not undergo NPQ in biologically relevant conditions in Arabidopsis thaliana The possible role of zeaxanthin in PSI photoprotection is discussed.
Asunto(s)
Arabidopsis/metabolismo , Complejo de Proteína del Fotosistema I/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Zeaxantinas/metabolismoRESUMEN
Photosynthesis starts when a pigment in the photosynthetic antennae absorbs a photon. The electronic excitation energy is then transferred through the network of light-harvesting pigments to special chlorophyll (Chl) molecules in the reaction centres, where electron transfer is initiated. Energy transfer and primary electron transfer processes take place on timescales ranging from femtoseconds to nanoseconds, and can be monitored in real time via time-resolved fluorescence spectroscopy. This method is widely used for measurements on unicellular photosynthetic organisms, isolated photosynthetic membranes, and individual complexes. Measurements on intact leaves remain a challenge due to their high structural heterogeneity, high scattering, and high optical density, which can lead to optical artefacts. However, detailed information on the dynamics of these early steps, and the underlying structure-function relationships, is highly informative and urgently required in order to get deeper insights into the physiological regulation mechanisms of primary photosynthesis. Here, we describe a current methodology of time-resolved fluorescence measurements on intact leaves in the picosecond to nanosecond time range. Principles of fluorescence measurements on intact leaves, possible sources of alterations of fluorescence kinetics and the ways to overcome them are addressed. We also describe how our understanding of the organisation and function of photosynthetic proteins and energy flow dynamics in intact leaves can be enriched through the application of time-resolved fluorescence spectroscopy on leaves. For that, an example of a measurement on Zea mays leaves is presented.
Asunto(s)
Transporte de Electrón , Transferencia de Energía , Espectrometría de Fluorescencia , Zea mays/química , Clorofila/metabolismo , Fluorescencia , Fotones , Fotosíntesis , Hojas de la Planta/química , Hojas de la Planta/fisiología , Zea mays/fisiologíaRESUMEN
The global rise in sea surface temperatures causes regular exposure of corals to high temperature and high light stress, leading to worldwide disastrous coral bleaching events (loss of symbiotic dinoflagellates (Symbiodinium) from reef-building corals). Our picosecond chlorophyll fluorescence experiments on cultured Symbiodinium clade C cells exposed to coral bleaching conditions uncovered the transformations of the alga's photosynthetic apparatus (PSA) that activate an extremely efficient non-photochemical "super-quenching" mechanism. The mechanism is associated with a transition from an initially heterogeneous photosystem II (PSII) pool to a homogeneous "spillover" pool, where nearly all excitation energy is transferred to photosystem I (PSI). There, the inherently higher stability of PSI and high quenching efficiency of P(700)(+) allow dumping of PSII excess excitation energy into heat, resulting in almost complete cessation of photosynthetic electron transport (PET). This potentially reversible "super-quenching" mechanism protects the PSA against destruction at the cost of a loss of photosynthetic activity. We suggest that the inhibition of PET and the consequent inhibition of organic carbon production (e.g. sugars) in the symbiotic Symbiodinium provide a trigger for the symbiont expulsion, i.e. bleaching.
Asunto(s)
Antozoos/parasitología , Dinoflagelados/fisiología , Estrés Fisiológico/fisiología , Simbiosis/fisiología , Temperatura , Animales , Clorofila/metabolismo , Dinoflagelados/metabolismo , Dinoflagelados/ultraestructura , Transporte de Electrón/efectos de la radiación , Cinética , Luz , Mediciones Luminiscentes/métodos , Microscopía Electrónica de Transmisión , Modelos Biológicos , Oxidación-Reducción/efectos de la radiación , Fotosíntesis/efectos de la radiación , Complejo de Proteína del Fotosistema I/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Tilacoides/metabolismo , Tilacoides/efectos de la radiación , Factores de TiempoRESUMEN
The thylakoid membrane protein PsbS is critical for quenching excessive excitation energy in mechanisms that involve the light-harvesting complexes of photosystem II. Liposomes of thylakoid lipids have been shown to be a very good platform to study photosynthetic membrane proteins and their interactions. In this study, we simultaneously refolded and reconstituted functional pea PsbS into liposomes of thylakoid lipids starting from denatured expressed protein. Intrinsic fluorescence spectroscopy, trypsin digestion, and circular dichroism spectroscopy were used to characterize the native state of PsbS in the proteoliposomes. The functionality of refolded PsbS was further demonstrated by its effect on the fluorescence quenching of the major antenna system of photosystem II (LHCII) co-inserted into the liposomes. The fluorescence yield of native trimeric LHCII was lowered by PsbS by 50% at neutral pH and by a further 25% upon lowering the pH to 4.5. Furthermore, the acid-induced fluorescence reduction was completely reversed by addition of N,N'-dicyclohexylcarbodiimide, an inhibitor of protein protonation. These results indicate that reconstituted PsbS induces strong quenching of LHCII sensing changes in local pH via its protonation sites.
Asunto(s)
Proteínas de Arabidopsis/química , Complejos de Proteína Captadores de Luz/química , Liposomas/química , Complejo de Proteína del Fotosistema II/química , Tilacoides/química , Secuencia de Aminoácidos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Dicroismo Circular , Diciclohexilcarbodiimida/química , Fluorescencia , Concentración de Iones de Hidrógeno , Complejos de Proteína Captadores de Luz/genética , Complejos de Proteína Captadores de Luz/metabolismo , Datos de Secuencia Molecular , Pisum sativum/química , Complejo de Proteína del Fotosistema II/genética , Complejo de Proteína del Fotosistema II/metabolismo , Desnaturalización Proteica , Replegamiento Proteico , Espectrometría de Fluorescencia , Tirosina/químicaRESUMEN
Non-photochemical quenching (NPQ) protects photosynthetic organisms against photodamage by high light. One of the key measuring parameters for characterizing NPQ is the high-light induced decrease in chlorophyll fluorescence. The originally measured data are maximal fluorescence (Fm') signals as a function of actinic illumination time (Fm'(t)). Usually these original data are converted into the so-called Stern-Volmer quenching function, NPQSV(t), which is then analyzed and interpreted in terms of various NPQ mechanisms and kinetics. However, the interpretation of this analysis essentially depends on the assumption that NPQ follows indeed a Stern-Volmer relationship. Here, we question this commonly assumed relationship, which surprisingly has never been proven. We demonstrate by simulation of quenching data that particularly the conversion of time-dependent quenching curves like Fm'(t) into NPQSV(t) is (mathematically) not "innocent" in terms of its effects. It distorts the kinetic quenching information contained in the originally measured function Fm'(t), leading to a severe (often sigmoidal) distortion of the time-dependence of quenching and has negative impact on the ability to uncover the underlying quenching mechanisms and their contribution to the quenching kinetics. We conclude that the commonly applied analysis of time-dependent NPQ in NPQSV(t) space should be reconsidered. First, there exists no sound theoretical basis for this common practice. Second, there occurs no loss of information whatsoever when analyzing and interpreting the originally measured Fm'(t) data directly. Consequently, the analysis of Fm'(t) data has a much higher potential to provide correct mechanistic answers when trying to correlate quenching data with other biochemical information related to quenching.
Asunto(s)
Clorofila/química , FluorescenciaRESUMEN
To protect the photosynthetic apparatus against photo-damage in high sunlight, the photosynthetic antenna of oxygenic organisms can switch from a light-harvesting to a photoprotective mode through the process of non-photochemical quenching (NPQ). There is growing evidence that light-harvesting proteins of photosystem II participate in photoprotection by a built-in capacity to switch their conformation between light-harvesting and energy-dissipating states. Here we applied high-resolution Magic-Angle Spinning Nuclear Magnetic Resonance on uniformly (13)C-enriched major light-harvesting complex II (LHCII) of Chlamydomonas reinhardtii in active or quenched states. Our results reveal that the switch into a dissipative state is accompanied by subtle changes in the chlorophyll (Chl) a ground-state electronic structures that affect their NMR responses, particularly for the macrocycle (13)C4, (13)C5 and (13)C6 carbon atoms. Inspection of the LHCII X-ray structures shows that of the Chl molecules in the terminal emitter domain, where excited-state energy accumulates prior to further transfer or dissipation, the C4, 5 and 6 atoms are in closest proximity to lutein; supporting quenching mechanisms that involve altered Chl-lutein interactions in the dissipative state. In addition the observed changes could represent altered interactions between Chla and neoxanthin, which alters its configuration under NPQ conditions. The Chls appear to have increased dynamics in unquenched, detergent-solubilized LHCII. Our work demonstrates that solid-state Nuclear Magnetic Resonance is applicable to investigate high-resolution structural details of light-harvesting proteins in varied functional conditions, and represents a valuable tool to address their molecular plasticity associated with photoprotection.
Asunto(s)
Clorofila/química , Complejos de Proteína Captadores de Luz/química , Espectroscopía de Resonancia Magnética/métodos , Clorofila ARESUMEN
Chlorophyll a fluorescence is a non-invasive tool widely used in photosynthesis research. According to the dominant interpretation, based on the model proposed by Duysens and Sweers (1963, Special Issue of Plant and Cell Physiology, pp 353-372), the fluorescence changes reflect primarily changes in the redox state of Q(A), the primary quinone electron acceptor of photosystem II (PSII). While it is clearly successful in monitoring the photochemical activity of PSII, a number of important observations cannot be explained within the framework of this simple model. Alternative interpretations have been proposed but were not supported satisfactorily by experimental data. In this review we concentrate on the processes determining the fluorescence rise on a dark-to-light transition and critically analyze the experimental data and the existing models. Recent experiments have provided additional evidence for the involvement of a second process influencing the fluorescence rise once Q(A) is reduced. These observations are best explained by a light-induced conformational change, the focal point of our review. We also want to emphasize that-based on the presently available experimental findings-conclusions on α/ß-centers, PSII connectivity, and the assignment of FV/FM to the maximum PSII quantum yield may require critical re-evaluations. At the same time, it has to be emphasized that for a deeper understanding of the underlying physical mechanism(s) systematic studies on light-induced changes in the structure and reaction kinetics of the PSII reaction center are required.
Asunto(s)
Clorofila/metabolismo , Fluorescencia , Clorofila A , Luz , Modelos Biológicos , Fotosíntesis/fisiología , Complejo de Proteína del Fotosistema II/metabolismoRESUMEN
A modified dye-sensitized solar cell consisting of a thin TiO2 barrier layer sensitized with natural trimeric light-harvesting complex II (LHCII) from spinach was used as a biomimetic model to study the effects of LHCII aggregation on the photovoltaic properties. The aggregation of individual trimers induced molecular reorganization, which dramatically increased the photocurrent. The morphology of small- and large-size LHCII aggregates deposited on a surface was confirmed by atomic force microscopy. Enhanced LHCII immobilization was accomplished via electrostatic interaction with amine-functionalized photoanodes. The photocurrent responses of the assembled solar cells under illumination at three characteristic wavelength bands in the UV-Vis absorption spectra of LHCII solutions confirmed that a significant photocurrent was generated by LHCII photosensitizers. The enhanced photocurrent by large aggregated LHCII is shown to correlate with the quenching in the far-red fluorescence deriving from chlorophyll-chlorophyll charge transfer states that are effectively coupled with the TiO2 surface and thus inject electrons into the TiO2 conduction band. The large aggregated LHCII with more chlorophyll-chlorophyll charge transfer states is a much better sensitizer since it injects electrons more efficiently into the conduction band of TiO2 than the small aggregated LHCII mostly consisting of unquenched chlorophyll excited state. The assembled solar cells demonstrated remarkable stability in both aqueous buffer and acetonitrile electrolytes over 30 days.
Asunto(s)
Fuentes de Energía Bioeléctrica , Complejos de Proteína Captadores de Luz/química , Complejos de Proteína Captadores de Luz/efectos de la radiación , Energía Solar , Titanio/química , Transferencia de Energía/efectos de la radiación , Diseño de Equipo , Análisis de Falla de Equipo , LuzRESUMEN
Observations of low-lying dark states in several photosynthetic complexes challenge our understanding of the mechanisms behind their efficient energy transfer processes. Computational models are necessary for providing novel insights into the nature and function of dark states, especially since these are not directly accessible in spectroscopy experiments. Here, we will focus on signatures of dark-type states in chlorosomes, a light-harvesting complex from green sulfur bacteria well-known for uniting a broad absorption band with very efficient energy transfer. In agreement with experiments, our simulations of two-dimensional electronic spectra capture the ultrafast exciton transfer occurring in 100s of femtoseconds within a single chlorosome cylinder. The sub-100 fs process corresponds to relaxation within the single-excitation manifold in a single chlorosome tube, where all initially created populations in the bright exciton states are quickly transferred to dark-type exciton states. Structural inhomogeneities on the local scale cause a redistribution of the oscillator strength, leading to the emergence of these dark-type exciton states, which dominate ultrafast energy transfer. The presence of the dark-type exciton states suppresses energy loss from an isolated chlorosome via fluorescence quenching, as observed experimentally. Our results further question whether relaxation to dark-exciton states is a leading process or merely competes with transfer to the baseplate within the photosynthetic apparatus of green sulfur bacteria.
RESUMEN
Photoprotection of photosystem II (PSII) is essential to avoid the light-induced damage of the photosynthetic apparatus due to the formation of reactive oxygen species (=photo-oxidative stress) under excess light. Carotenoids are known to play a crucial role in these processes based on their property to deactivate triplet chlorophyll (³Chl*) and singlet oxygen (¹O2*). Xanthophylls are further assumed to be involved either directly or indirectly in the non-photochemical quenching (NPQ) of excess light energy in the antenna of PSII. This review gives an overview on recent progress in the understanding of the photoprotective role of the xanthophylls zeaxanthin (which is formed in the light in the so-called xanthophyll cycle) and lutein with emphasis on the NPQ processes associated with PSII of higher plants. The current knowledge supports the view that the photoprotective role of Lut is predominantly restricted to its function in the deactivation of ³Chl*, while zeaxanthin is the major player in the deactivation of excited singlet Chl (¹Chl*) and thus in NPQ (non-photochemical quenching). Additionally, zeaxanthin serves important functions as an antioxidant in the lipid phase of the membrane and is likely to act as a key component in the memory of the chloroplast with respect to preceding photo-oxidative stress. This article is part of a Special Issue entitled: Photosystem II.
Asunto(s)
Luz , Luteína/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Xantófilas/metabolismo , Modelos MolecularesRESUMEN
Non-photochemical quenching (NPQ) of chlorophyll fluorescence is thought to be an indicator of an essential regulation and photoprotection mechanism against high-light stress in photosynthetic organisms. NPQ is typically characterized by modulated pulse fluorometry and it is often assumed implicitly to be a good proxy for the actual physiological photoprotection capacity of the organism. Using the results of previously published ultrafast fluorescence measurements on intact leaves of w.t. and mutants of Arabidopsis (Holzwarth et al. 2009) we have developed exact relationships for the fluorescence quenching and the corresponding Photosystem II acceptor side photoprotection effects under NPQ conditions. The approach based on the exciton-radical pair equilibrium model assumes that photodamage results from triplet states generated in the reaction center. The derived relationships allow one to distinguish and determine the individual and combined quenching as well as photoprotection contributions of each of the multiple NPQ mechanisms. Our analysis shows inter alia that quenching and photoprotection are not linearly related and that antenna detachment, which can be identified with the so-called qE mechanism, contributes largely to the measured fluorescence quenching but does not correspond to the most efficient photoprotective response. Conditions are formulated which allow simultaneously the maximal photosynthetic electron flow as well as maximal acceptor side photoprotection. It is shown that maximal photoprotection can be achieved if NPQ is regulated in such a way that PSII reaction centers are open under given light conditions. The results are of fundamental importance for a proper interpretation of the physiological relevance of fluorescence-based NPQ data.
Asunto(s)
Arabidopsis/metabolismo , Luz , Procesos Fotoquímicos , Complejo de Proteína del Fotosistema II/metabolismo , Arabidopsis/efectos de la radiación , Oscuridad , Fluorescencia , Cinética , Modelos Biológicos , Modelos Moleculares , Procesos Fotoquímicos/efectos de la radiación , Factores de TiempoRESUMEN
Photosystem I (PSI) is a large pigment-protein complex that unites a reaction center (RC) at the core with approximately 100 core antenna chlorophylls surrounding it. The RC is composed of two cofactor branches related by a pseudo-C2 symmetry axis. The ultimate electron donor, P(700) (a pair of chlorophylls), and the tertiary acceptor, F(X) (a Fe(4)S(4) cluster), are both located on this axis, while each of the two branches is made up of a pair of chlorophylls (ec2 and ec3) and a phylloquinone (PhQ). Based on the observed biphasic reduction of F(X), it has been suggested that both branches in PSI are competent for electron transfer (ET), but the nature and rate of the initial electron transfer steps have not been established. We report an ultrafast transient absorption study of Chlamydomonas reinhardtii mutants in which specific amino acids donating H-bonds to the 13(1)-keto oxygen of either ec3(A) (PsaA-Tyr696) or ec3(B) (PsaB-Tyr676) are converted to Phe, thus breaking the H-bond to a specific ec3 cofactor. We find that the rate of primary charge separation (CS) is lowered in both mutants, providing direct evidence that the primary ET event can be initiated independently in each branch. Furthermore, the data provide further support for the previously published model in which the initial CS event occurs within an ec2/ec3 pair, generating a primary ec2(+)ec3(-) radical pair, followed by rapid reduction by P(700) in the second ET step. A unique kinetic modeling approach allows estimation of the individual ET rates within the two cofactor branches.
Asunto(s)
Complejo de Proteína del Fotosistema I/química , Transporte de Electrón , Cinética , Modelos Moleculares , Análisis Espectral/métodosRESUMEN
We establish a general kinetic scheme for energy transfer and trapping in the photosystem I (PSI) of cyanobacteria grown under white light (WL) or far-red light (FRL) conditions. With the help of simultaneous target analysis of all emission and transient absorption datasets measured in five cyanobacterial strains, we resolved the spectral and kinetic properties of the different species present in PSI. WL-PSI can be described by Bulk Chl a, two Red Chl a, and a reaction center compartment (WL-RC). The FRL-PSI contains two additional Chl f compartments. The lowest excited state of the FRL-RC is downshifted by ≈ 29 nm. The rate of charge separation drops from ≈900 ns-1 in WL-RC to ≈300 ns-1 in FRL-RC. The delayed trapping in the FRL-PSI (≈130 ps) is explained by uphill energy transfer from the Chl f compartments with Gibbs free energies of ≈kBT below that of the FRL-RC.