Antenna Chlorophyll Research Articles

Differential accumulation ofLhcbgene products in thylakoid membranes ofZea maysplants grown under contrasting light and temperature conditions

In higher plants many different genes encode Lhcb proteins that belong to a highly conserved protein family. Evolutionary conservation of this genetic redundancy suggests that individual gene products play different roles in light harvesting and photoprotection depending on environmental conditions. We have tested the hypothesis that expression/accumulation of individual light harvesting complex (Lhc) proteins depends on plant growth conditions. Zea mays plants were grown in different temperature (13 degrees C vs. 24 degrees C) and light (high vs. low) conditions. The thylakoid membranes were isolated and fractionated by sucrose gradient and the protein content of the different bands was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Significant differences were found in the accumulation of both the major light harvesting complex of photosystem II (LHCII) complexes and the minor antenna chlorophyll proteins CP29, CP26 and CP24. In particular, temperature seems to play a major role in driving the expression/accumulation of the different proteins: the LHCII/minor antenna ratio increases with decreasing temperature. The pigment composition and the spectroscopic properties of LHCII complexes isolated from low temperature grown plants are significantly different from those of LHCII purified from high temperature grown plants. Two-dimensional maps show that different LHCII proteins are accumulated at different levels depending on growth conditions. Moreover the low temperature/high light grown plants show an increased value of nonphotochemical quenching. These results suggest a specific role of different LHCII complexes in the organization of the potosystem II and photoprotection.

Non-photochemical quenching of chlorophyll a fluorescence by oxidised plastoquinone: new evidences based on modulation of the redox state of the endogenous plastoquinone pool in broken spinach chloroplasts

Twenty-five years ago, non-photochemical quenching of chlorophyll fluorescence by oxidised plastoquinone (PQ) was proposed to be responsible for the lowering of the maximum fluorescence yield reported to occur when leaves or chloroplasts were treated in the dark with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of electron flow beyond the primary quinone electron acceptor (Q A) of photosystem (PS) II [C. Vernotte, A.L. Etienne, J.-M. Briantais, Quenching of the system II chlorophyll fluorescence by the plastoquinone pool, Biochim. Biophys. Acta 545 (1979) 519-527]. Since then, the notion of PQ-quenching has received support but has also been put in doubt, due to inconsistent experimental findings. In the present study, the possible role of the native PQ-pool as a non-photochemical quencher was reinvestigated, employing measurements of the fast chlorophyll a fluorescence kinetics (from 50 μs to 5 s). The about 20% lowering of the maximum fluorescence yield F M, observed in osmotically broken spinach chloroplasts treated with DCMU, was eliminated when the oxidised PQ-pool was non-photochemically reduced to PQH 2 by dark incubation of the samples in the presence of NAD(P)H, both under anaerobic and aerobic conditions. Incubation under anaerobic conditions in the absence of NAD(P)H had comparatively minor effects. In DCMU-treated samples incubated in the presence of NAD(P)H fluorescence quenching started to develop again after 20–30 ms of illumination, i.e., the time when PQH 2 starts getting reoxidised by PS I activity. NAD(P)H-dependent restoration of F M was largely, if not completely, eliminated when the samples were briefly (5 s) pre-illuminated with red or far-red light. Addition to the incubation medium of HgCl 2 that inhibits dark reduction of PQ by NAD(P)H also abolished NAD(P)H-dependent restoration of F M. Collectively, our results provide strong new evidence for the occurrence of PQ-quenching. The finding that DCMU alone did not affect the minimum fluorescence yield F 0 allowed us to calculate, for different redox states of the native PQ-pool, the fractional quenching at the F 0 level ( Q 0) and to compare it with the fractional quenching at the F M level ( Q M). The experimentally determined Q 0/ Q M ratios were found to be equal to the corresponding F 0/ F M ratios, demonstrating that PQ-quenching is solely exerted on the excited state of antenna chlorophylls.

Remodeling of Light-Harvesting Protein Complexes in Chlamydomonas in Response to Environmental Changes

Photosynthesis is dependent on light. Photosynthetic organisms struggle their entire lives to optimize photosynthetic function and minimize photooxidative damage in response to light quantity and quality. When the absorbed light energy exceeds the capacity of photosynthetic energy consumption, overreduction of electron transport carriers and accumulation of excitation energy in the light-harvesting-antennae may occur. The latter favors the production of excited triplet-state chlorophyll molecules ( 3 Chl) that can interact with O2 to cause the formation of reactive singlet oxygen ( 1 O2). The former favors the direct reduction of O2 by photosystem I (PSI) and the subsequent generation of reactive oxygen species, such as superoxide (O2 ), hydrogen peroxide (H2O2), and the hydroxyl radical (OH) (5, 6, 75). These reactive oxygen species are able to cause photo-oxidative damage to photosystem II (PSII), a primary target for photoinhibition (2, 4, 9), but also to PSI (59, 60), in particular under weak light at chilled temperatures (111, 121). Thus, adaptation mechanisms that balance the energy input, through photochemistry via the photosynthetic machinery, with the energy output through CO2 assimilation and other metabolic pathways, are essential for plant survival. The function of the light-harvesting complexes (LHCs) is, as the name implies, to increase the effective absorption crosssection of the photosystems and to supply them with excitation energy. The LHCs bind chlorophyll a and b but lack any photochemical activity of their own. Both PSI and PSII have their own core antenna pigments, but the addition of the LHCs increases the number of antenna pigments connected to each reaction center by a factor of 2 to 4, depending upon the conditions (47). Energy transfer operates in a time scale of femtoseconds to picoseconds. The transfer time of excitation energy between neighboring Chl molecules in the antenna has been estimated to be 100 to 300 fs (45). It had long been known that the pigment molecules must be closely packed to allow such fast and efficient transfer, a supposition beautifully confirmed by the emergence of the X-ray crystal structures of PSI (65), PSII (42, 66, 135), and LHCI-PSI (12) in the last several years (see discussion below). On average, excitation energy makes 100 to 1,000 such hops between antenna chlorophylls before it is trapped at a reaction center. Depending upon the arrangement of pigment molecules within LHCs and of LHCs vis-a `-vis the photosystems, excitation energy can be directed to specific photosystems, temporarily trapped on low-energy chlorophylls, or even converted to heat by “nonphotochemical quenching.” It does not require a great stretch of imagination to see how evolution might have taken advantage of such possibilities for the development of regulatory mechanisms to deal with variable illumination. The regulation of light energy input and distribution, by the dynamic regulation of the light-harvesting system, probably plays the most important role in balancing the light and dark reactions of photosynthesis. Light-harvesting systems and their functioning in algae and vascular plants have been addressed

Genomics of green algal hydrogen research

This article summarizes knowledge on genes and their respective proteins in the field of green algal hydrogen research. Emphasis is placed on recently cloned genes from the unicellular green alga Chlamydomonas reinhardtii, including HydA1 and HydA2, which encode homologous [Fe]-hydrogenases, Tla1, which encodes a chlorophyll antenna size regulatory gene, SulP, which encodes a chloroplast sulfate permease, and Sta7, which encodes an isoamylase. Analysis of the structure and function of these genes and of their respective proteins in C. reinhardtii, and related unicellular green algae, is presented in light of the role they play in the hydrogen metabolism in these organisms. A discussion is offered as to the potential application of these genes in the field of hydrogen photoproduction.

Optimization and evolution of light harvesting in photosynthesis: the role of antenna chlorophyll conserved between photosystem II and photosystem I.

The efficiency of oxygenic photosynthesis depends on the presence of core antenna chlorophyll closely associated with the photochemical reaction centers of both photosystem II (PSII) and photosystem I (PSI). Although the number and overall arrangement of these chlorophylls in PSII and PSI differ, structural comparison reveals a cluster of 26 conserved chlorophylls in nearly identical positions and orientations. To explore the role of these conserved chlorophylls within PSII and PSI we studied the influence of their orientation on the efficiency of photochemistry in computer simulations. We found that the native orientations of the conserved chlorophylls were not optimal for light harvesting in either photosystem. However, PSII and PSI each contain two highly orientationally optimized antenna chlorophylls, located close to their respective reaction centers, in positions unique to each photosystem. In both photosystems the orientation of these optimized bridging chlorophylls had a much larger impact on photochemical efficiency than the orientation of any of the conserved chlorophylls. The differential optimization of antenna chlorophyll is discussed in the context of competing selection pressures for the evolution of light harvesting in photosynthesis.

The Long-Wavelength Chlorophyll States of Plant LHCI at Room Temperature: A Comparison with PSI-LHCI

The red antenna states of the external antenna complexes of higher plant photosystem I, known as LHCI, have been analyzed by measurement of their preequilibrium fluorescence upon direct excitation at 280 K. In addition to the previously detected F735 state, a hitherto undetected low-energy state with emission maximum around 713 nm was observed. The 280 K bandwidths (FWHM) are 55 nm for the F735 state and ∼27 nm for the F713-nm state, much greater than for non-red-shifted antenna chlorophylls. The origin absorption band for the F735-nm state was directly detected by determination of its excitation (action) spectrum and lies at 709–710 nm. The absorption spectrum for F735, calculated using the Stepanov expression, closely overlaps the excitation spectrum, indicating that the very large Stokes shift (25 nm) is due to vibrational relaxation within the excited-state manifold and solvent effects can be excluded. Fluorescence anisotropy measurements, with direct excitation of F735, indicate that the transition dipoles of the two red states are parallel. Similar experiments performed in the long-wavelength absorbing tail of PSI-LHCI indicate the presence of emission state(s) that are red-shifted with respect to F735 of isolated LHCI. It is suggested that these are brought about by interactions between the complexes in PSI-LHCI, which occur in some yet undefined way, and which are broken upon solubilization of the component parts.

Energy transfer followed by electron transfer in a supramolecular triad composed of boron dipyrrin, zinc porphyrin, and fullerene: a model for the photosynthetic antenna-reaction center complex.

The first example of a working model of the photosynthetic antenna-reaction center complex, constructed via self-assembled supramolecular methodology, is reported. For this, a supramolecular triad is assembled by axially coordinating imidazole-appended fulleropyrrolidine to the zinc center of a covalently linked zinc porphyrin-boron dipyrrin dyad. Selective excitation of the boron dipyrrin moiety in the boron dipyrrin-zinc porphyrin dyad resulted in efficient energy transfer (k(ENT)(singlet) = 9.2 x 10(9) s(-)(1); Phi(ENT)(singlet) = 0.83) creating singlet excited zinc porphyrin. Upon forming the supramolecular triad, the excited zinc porphyrin resulted in efficient electron transfer to the coordinated fullerenes, resulting in a charge-separated state (k(cs)(singlet) = 4.7 x 10(9) s(-)(1); Phi(CS)(singlet) = 0.9). The observed energy transfer followed by electron transfer in the present supramolecular triad mimics the events of natural photosynthesis. Here, the boron dipyrrin acts as antenna chlorophyll that absorbs light energy and transports spatially to the photosynthetic reaction center, while the electron transfer from the excited zinc porphyrin to fullerene mimics the primary events of the reaction center where conversion of the electronic excitation energy to chemical energy in the form of charge separation takes place. The important feature of the present model system is its relative "simplicity" because of the utilized supramolecular approach to mimic rather complex "combined antenna-reaction center" events of photosynthesis.

Electrochromic Shift of Chlorophyll Absorption in Photosystem I from Synechocystis sp. PCC 6803: A Probe of Optical and Dielectric Properties around the Secondary Electron Acceptor

Nanosecond absorption dynamics at ∼685 nm after excitation of photosystem I (PS I) from Synechocystis sp. PCC 6803 is consistent with electrochromic shift of absorption bands of the Chl a pigments in the vicinity of the secondary electron acceptor A 1. Based on experimental optical data and structure-based simulations, the effective local dielectric constant has been estimated to be between 3 and 20, which suggests that electron transfer in PS I is accompanied by considerable protein relaxation. Similar effective dielectric constant values have been previously observed for the bacterial photosynthetic reaction center and indicate that protein reorganization leading to effective charge screening may be a necessary structural property of proteins that facilitate the charge transfer function. The data presented here also argue against attributing redmost absorption in PS I to closely spaced antenna chlorophylls (Chls) A38 and A39, and suggest that optical transitions of these Chls, along with that of connecting chlorophyll (A40) lie in the range 680–695 nm.

The Effect of Zeaxanthin as the Only Xanthophyll on the Structure and Function of the Photosynthetic Apparatus in Arabidopsis thaliana

In green plants, the xanthophyll carotenoid zeaxanthin is synthesized transiently under conditions of excess light energy and participates in photoprotection. In the Arabidopsis lut2 npq2 double mutant, all xanthophylls were replaced constitutively by zeaxanthin, the only xanthophyll whose synthesis was not impaired. The relative proportions of the different chlorophyll antenna proteins were strongly affected with respect to the wild-type strain. The major antenna, LHCII, did not form trimers, and its abundance was strongly reduced as was CP26, albeit to a lesser extent. In contrast, CP29, CP24, LHCI proteins, and the PSI and PSII core complexes did not undergo major changes. PSII-LHCII supercomplexes were not detectable while the PSI-LHCI supercomplex remained unaffected. The effect of zeaxanthin accumulation on the stability of the different Lhc proteins was uneven: the LHCII proteins from lut2 npq2 had a lower melting temperature as compared with the wild-type complex while LHCI showed increased resistance to heat denaturation. Consistent with the loss of LHCII, light-state 1 to state 2 transitions were suppressed, the photochemical efficiency in limiting light was reduced and photosynthesis was saturated at higher light intensities in lut2 npq2 leaves, resulting in a photosynthetic phenotype resembling that of high light-acclimated leaves. Zeaxanthin functioned in vivo as a light-harvesting accessory pigment in lut2 npq2 chlorophyll antennae. As a whole, the in vivo data are consistent with the results obtained by using recombinant Lhc proteins reconstituted in vitro with purified zeaxanthin. While PSII photoinhibition was similar in wild type and lut2 npq2 exposed to high light at low temperature, the double mutant was much more resistant to photooxidative stress and lipid peroxidation than the wild type. The latter observation is consistent with an antioxidant and lipid protective role of zeaxanthin in vivo.

Energy transfer and trapping in the Photosystem I complex of Synechococcus PCC 7942 and in its supercomplex with IsiA

The cyanobacterium Synechococcus PCC 7942 grown under iron starvation assembles a supercomplex consisting of a trimeric Photosystem I (PSI) complex encircled by a ring of 18 CP43′ or IsiA complexes. It has previously been shown that PSI of Synechococcus PCC 7942 contains less special long-wavelength (‘red’) chlorophylls than PSI of most other cyanobacteria. Here we present a comparative analysis by time-resolved absorption difference and fluorescence spectroscopy of the processes of energy transfer and trapping in trimeric PSI and PSI–IsiA supercomplexes from Synechococcus PCC 7942. All experiments were performed with the primary electron donor of PSI (P700) in the oxidized state. Our data suggest that in the PSI complex the excitation energy is equilibrated with a lifetime of 0.6 ps among the so-called bulk chlorophylls, is distributed in 3–4 ps between the bulk and red chlorophylls, and is trapped in the reaction center in 19 ps. This trapping time is shorter than that observed for other cyanobacteria, which we attribute to the lower content of red chlorophylls in PSI of this organism. In the PSI–IsiA supercomplexes, the distribution of excited states is blue-shifted compared to that in PSI, leading to a lengthening of the equilibration processes. We attributed a phase of about 1 ps to initial energy equilibration steps among the IsiA and PSI core bulk chlorophylls, a 5–7 ps phase to equilibration between bulk and red chlorophylls within the PSI core, and a 38 ps phase to trapping in the reaction center. The data suggest that the excitation energy is equilibrated among the IsiA and PSI core antenna chlorophylls before trapping occurs. Data analysis based on a simple kinetic model revealed an intrinsic rate constant for energy transfer from IsiA to PSI in the range of 2±1 ps. Based on this value we suggest the presence of one or more linker chlorophylls between the IsiA and PSI core complexes. These results confirm that IsiA acts as an effective light-harvesting antenna for PSI.

The orientations of core antenna chlorophylls in photosystem II are optimized to maximize the quantum yield of photosynthesis

In photosystem II (PSII) the probability that energy absorbed by core antenna chlorophyll (Chl) is transferred to the reaction center (RC) is extremely high. Although close proximity between antenna Chl ensures a high transfer efficiency, relative pigment orientation can fractionally modify it. This level of refinement has often been assumed to be superfluous as so many subsequent processes limit the overall efficiency of photosynthesis. Nevertheless, did natural selection act on the most efficient step of energy conversion in PSII by optimizing the orientation of antenna Chl? Our Monte Carlo simulations sampled the orientation space of Chls in kinetic models for excitation energy transfer based on the X-ray structures of PSII from Thermosynechococcus vulcanus and Synechocystis elongatus. Our results revealed that the orientations of key antenna Chls are optimized to maximize photosynthesis while the orientations of the two peripheral RC Chls (Chl Z) are not.

Photoacclimation in Spathiphyllum

We studied photoacclimation in Spathiphyllum grown at an irradiance of 40 or 420 μmol/m 2 s (LL or HL, respectively). All parameters studied responded to acclimation. Leaves at LL, in contrast to HL, were thinner and oriented perpendicular to the incident light, had more chlorophyll per g f w, fewer stomata on the upper leaf surface and a reduced layer of mesophyll cells. Their chloroplasts at HL had wider grana with less thylakoids per granum, and better organized photosystems than at LL. PSI and PSII activities per mg chlorophyll ( V max), and PSI and PSII content (total activity per g f w), were lower at LL than at HL and so was the light requirement for saturation of the PSI or PSII partial photoreactions, suggesting that fewer photosystems with larger antenna size prevail at LL, but many more with smaller antenna size at HL. Analysis of chlorophyll distribution among the thylakoid pigment–protein complexes showed less antenna chlorophyll serving PSII (CPa + LHCP 1 + LHCP 3) than that serving PSI (CPIa + CPI + LHCP 2) at LL as compared to HL, and thus a lower PSII/PSI ratio at LL, in agreement with the general finding that LL plants, with larger PSII antenna size, have lower PSII/PSI ratio. The increase in PSI antenna size at LL was correlated with the increase in the distribution of chlorophyll in pigment–protein complexes serving PSI, and a very large chlorophyll/protein molar ratio in the isolated CPI complex. On the other hand, the PSII antenna chlorophyll (CPa + LHCP 1 + LHCP 3) on a g f w basis, and the chlorophyll a/b ratio remained more or less constant at LL or HL. This may reflect our finding that Spathiphyllum contains mainly the 27 kDa inner LHCII antenna protein, the size of which remains unaffected by photoacclimation. The increase in the distribution of chlorophyll in pigment–protein complexes serving PSII at HL, therefore, reflects the higher population of PSII at HL. Very high PSI activity was found at HL, which we attribute to the highly organized small in size PSI.

Crystal structure of cyanobacterial photosystem II at 3.2 Å resolution: a closer look at the Mn-cluster

In the crystal structure of photosystem II (PSII) from the cyanobacterium Thermosynechococcus elongatus at 3.2 A resolution, several loop regions of the principal protein subunits are now defined that were not interpretable previously at 3.8 A resolution. The head groups and side chains of the organic cofactors of the electron transfer chain and of antenna chlorophyll a (Chl a) have been modeled, coordinating and hydrogen bonding amino acids identified and the nature of the binding pockets derived. The orientations of these cofactors resemble those of the reaction center from anoxygenic purple bacteria, but differences in hydrogen bonding and protein environment modulate their properties and provide the unique high redox potential (1.17 V) of the primary donor. Coordinating amino acids of manganese cluster, redox-active TyrZ and non-haem Fe2+ have been determined, and an all-trans β-carotene connects cytochrome b-559, ChlZ and primary electron donor (coordinates are available under PDB-code 1W5C).

Species-specific Differences of the Spectroscopic Properties of P700: ANALYSIS OF THE INFLUENCE OF NON-CONSERVED AMINO ACID RESIDUES BY SITE-DIRECTED MUTAGENESIS OF PHOTOSYSTEM I FROM CHLAMYDOMONAS REINHARDTII

We applied optical spectroscopy, magnetic resonance techniques, and redox titrations to investigate the properties of the primary electron donor P700 in photosystem I (PS I) core complexes from cyanobacteria (Thermosynechococcus elongatus, Spirulina platensis, and Synechocystis sp. PCC 6803), algae (Chlamydomonas reinhardtii CC2696), and higher plants (Spinacia oleracea). Remarkable species-specific differences of the optical properties of P700 were revealed monitoring the (3P700-P700) and (P700+.-P700) absorbance and CD difference spectra. The main bleaching band in the Qy region differs in peak position and line width for the various species. In cyanobacteria the absorbance of P700 extends more to the red compared with algae and higher plants which is favorable for energy transfer from red core antenna chlorophylls to P700 in cyanobacteria. The amino acids in the environment of P700 are highly conserved with two distinct deviations. In C. reinhardtii a Tyr is found at position PsaB659 instead of a Trp present in all other organisms, whereas in Synechocystis a Phe is found instead of a Trp at the homologous position PsaA679. We constructed several mutants in C. reinhardtii CC2696. Strikingly, no PS I could be detected in the mutant YW B659 indicating steric constraints unique to this organism. In the mutants WA A679 and YA B659 significant changes of the spectral features in the (3P700-P700), the (P700+.-P700) absorbance difference and in the (P700+.-P700) CD difference spectra are induced. The results indicate structural differences among PS I from higher plants, algae, and cyanobacteria and give further insight into specific protein-cofactor interactions contributing to the optical spectra.

Structural changes upon excitation of D1-D2-Cyt b559 photosystem II reaction centers depend on the beta-carotene content.

Different preparations of D1-D2-Cyt b559 complexes from spinach with different beta-carotene (Car) content [on average from <0.5 to 2 per reaction center (RC)] were studied by means of laser-induced optoacoustic spectroscopy. phiP680(+)Pheo(-) does not depend on the preparation (or on the Car content) inasmuch as the magnitude of the prompt heat (produced within 20 ns) does not vary for the different samples upon excitation at 675 and 620 nm. The energy level of the primary charge-separated state, P680(+)Pheo(-), was determined as EP680(+)Pheo(-) = 1.55 eV. Thus, an enthalpy change accompanying charge separation from excited P680 of deltaH*P680Pheo-->P680(+)Pheo(-) = -0.27 eV is obtained. Calculations using the heat evolved during the time-resolved decay of P680(+)Pheo(-) (< or = 100 ns) affords a triplet (3[P680Pheo]) quantum yield phi3[P680Pheo] = 0.5 +/- 0.14. The structural volume change, deltaV1, corresponding to the formation of P680(+)Pheo(-), strongly depends on the Car content; it is ca. -2.5 A3 molecule(-1) for samples with <0.5 Car on average, decreases (in absolute value) to -0.5 +/- 0.2 A3 for samples with an average of 1 Car, and remains the same for samples with two Cars per RC. This suggests that the Car molecules induce changes in the ground-state RC conformation, an idea which was confirmed by preferential excitation of Car with blue light, which produced different carotene triplet lifetimes in samples with 2 Car compared to those containing less carotene. We conclude that the two beta-carotenes are not structurally equivalent. Upon blue-light excitation (480 nm, preferential carotene absorption) the fraction of energy stored is ca. 60% for the 9Chl-2Car sample, whereas it is 40% for the preparations with one or less Cars on average, indicating different paths of energy distribution after Car excitation in these RCs with remaining chlorophyll antennae.

Role of the reversible xanthophyll cycle in the photosystem II damage and repair cycle in Dunaliella salina.

The Dunaliella salina photosynthetic apparatus organization and function was investigated in wild type (WT) and a mutant (zea1) lacking all beta,beta-epoxycarotenoids derived from zeaxanthin (Z). The zea1 mutant lacked antheraxanthin, violaxanthin, and neoxanthin from its thylakoid membranes but constitutively accumulated Z instead. It also lacked the so-called xanthophyll cycle, which, upon irradiance stress, reversibly converts violaxanthin to Z via a de-epoxidation reaction. Despite the pronounced difference observed in the composition of beta,beta-epoxycarotenoids between WT and zea1, no discernible difference could be observed between the two strains in terms of growth, photosynthesis, organization of the photosynthetic apparatus, photo-acclimation, sensitivity to photodamage, or recovery from photo-inhibition. WT and zea1 were probed for the above parameters over a broad range of growth irradiance and upon light shift experiments (low light to high light shift and vice versa). A constitutive accumulation of Z in the zea1 strain did not affect the acclimation of the photosynthetic apparatus to irradiance, as evidenced by indistinguishable irradiance-dependent adjustments in the chlorophyll antenna size and photosystem content of WT and zea1 strain. In addition, a constitutive accumulation of Z in the zea1 strain did not affect rates of photodamage or the recovery of the photosynthetic apparatus from photo-inhibition. However, Z in the WT accumulated in parallel with the accumulation of photodamaged PSII centers in the chloroplast thylakoids and decayed in tandem with a chloroplast recovery from photo-inhibition. These results suggest a role for Z in the protection of photodamaged and disassembled PSII reaction centers, apparently needed while PSII is in the process of degradation and replacement of the D1/32-kD reaction center protein.

Dynamic Interaction between the D1 protein, CP43 and OEC33 at the lumenal side of photosystem II in spinach chloroplasts: evidence from light-induced cross-Linking of the proteins in the donor-side photoinhibition.

During the donor-side photoinhibition of spinach photosystem II, the reaction center D1 protein cross-linked with the antenna chlorophyll binding protein CP43 of photosystem II lacking the oxygen-evolving complex (OEC) subunit proteins. The cross-linking did not occur upon illumination of photosystem II samples that retained the OEC33, nor when OEC33-depleted photosystem II samples were reconstituted with the OEC33 prior to illumination. These results suggest that the D1 protein, CP43 and the OEC33 are located in close proximity at the lumenal side of photosystem II, and that the OEC33 suppresses the unnecessary contact between the D1 protein and CP43. Previously we presented data showing the D1 protein located adjacent to CP43 on the stromal side of photosystem II [Ishikawa et al. (1999) BIOCHIM: Biophys. Acta 1413: 147]. The present data suggest that the spatial arrangement of the D1 protein and CP43 at the lumenal side of photosystem II in spinach chloroplasts is similar to that at the stromal side of photosystem II and is consistent with the assignment of these proteins recently proposed on the crystal structures of the photosystem II complexes from cyanobacteria [Zouni et al. (2001) Nature 409: 739, Kamiya and Shen 2003 PROC: Natl. Acad. Sci. USA, 100: 98]. Moreover, the data suggest that the binding condition and positioning of the OEC33 in the photosystem II complex from higher plants may be different from those in cyanobacteria.

The revision of the model of primary energy conversion in purple bacteria

A simulation method is suggested which enables one to check whether a model for excitation energy exchange in an ensemble of dye molecules fits available experimental data. In particular, this method may deal with photosynthetic units (PSUs) in which excitation migration in antenna chlorophylls and their substantial trapping in reaction centers (RCs) take place. Its application to the purple bacteria has proved that the model, which was generally accepted during the last 20–30 years, is in contradiction with recent experimental facts and thus requires modernization. Two physical mechanisms are discussed: femtosecond polarization of mobile hydrogen atoms near the reaction center special pair (“water latch”), and the presence of excitons delocalized over several core-bacteriochlorophylls (BChls). Our considerations give evidence that neither of these mechanisms alone can resolve the conflict, but their cumulative action appears to be sufficient. Unfortunately, these mechanisms were as yet only partially addressed experimentally.

Chlorophyll antenna size adjustments by irradiance in Dunaliella salina involve coordinate regulation of chlorophyll a oxygenase (CAO) and Lhcb gene expression.

To elucidate the mechanism of irradiance-dependent adjustments in the chlorophyll antenna size of photosynthesis, we addressed the regulation of expression of genes encoding a variety of chlorophyll biosynthesis enzymes and that of the Lhcb genes in the model organism Dunaliella salina. Among the chlorophyll biosynthesis enzymes tested, only the chlorophyll a oxygenase (CAO) gene responded to changes in the level of irradiance with substantial mRNA level and kinetics of change that were similar to those of the Lhcb genes. Evidence is presented for the operation of a cytosolic signal transduction pathway for the rapid (order of minutes) regulation of both CAO and Lhcb gene expression by irradiance. Inhibitor studies and transient activation of Ca2+-dependent kinase suggested phopholipase-C activation to Ca2+ release, and activation of a specific Ca2+/CaM-dependent protein kinase in this cytosolic signal transduction pathway. The redox state of the plastoquinone pool also serves to regulate CAO and Lhcb gene expression on a slower time scale (hours) and probably serves as a plastidic-origin signal that acts coordinately with the cytosolic signal transduction pathway. It is proposed that irradiance-dependent adjustments in the chlorophyll antenna size occur by coordinate regulation of CAO and Lhcb gene expression via two distinct signal transduction pathways in photosynthetic organisms.

Energy Equilibration among the Chlorophylls in the Electron-Transfer System of Photosystem I Reaction Center from Spinach

Excitation-wavelength dependence of the transient absorbance changes of photosystem I reaction center particles with a reduced number of antenna chlorophylls (16 chlorophylls/primary electron donor chlorophyll (P700)) hasbeen studied in an effort to understand the energy equilibration among the chlorophylls in the electron-transfer system. The photobleaching and stimulated-emission signals in the Q y band region of the chlorophylls upon the preferential excitation of chlorophyll spectral forms at the blue edge (662 nm) and red edge (697 nm) of the Q y band are analyzed. In the case of the red-edge excitation, spectral equilibration proceeds with a time constant of 0.34 (′0.07) ps, which is attributable to the energy equilibration between P700 and neighboring chlorophylls absorbing around 686 nm in the electron-transfer system. This equilibration seems to precede the fastest phase of the primary charge separation (apparent time constant of 0.8 ps) reported previously (Kumazaki et al. J. Phys. Chem. B 2001, 105, 1093). A slow decay of the excited states because of the slow phase of the primary charge separation proceeds with a time constant of 7.2 (′0.6) ps. In the case of the blue-edge excitation, vibrational relaxation and downhill energy transfer proceed with a time constant of 0.38 (′0.08) ps, which are followed by a slow downhill energy transfer from residual antenna chlorophylls to the electron-transfer system. Even with the slow energy transfer from the residual antenna chlorophylls to the electron-transfer system, the overall primary charge separation is completed with a time constant of 10 (′0.7) ps. These interpretations are in part supported by control experiments on chlorophyll a in ethanol under equivalent optical conditions. Implications of these results for understanding the primary processes in more intact photosystem I are discussed.