Long-term light adaptation of light-harvesting and energy-transfer processes in the glaucophyte Cyanophora paradoxa under different light conditions.

In response to fluctuation in light intensity and quality, oxygenic photosynthetic organisms modify their light-harvesting and excitation energy-transfer processes to maintain optimal photosynthetic activity. Glaucophytes, which are a group of primary symbiotic algae, possess light-harvesting antennas called phycobilisomes (PBSs) consistent with cyanobacteria and red algae. However, compared with cyanobacteria and red algae, glaucophytes are poorly studied and there are few reports on the regulation of photosynthesis in the group. In this study, we examined the long-term light adaptation of light-harvesting functions in a glaucophyte, Cyanophora paradoxa, grown under different light conditions. Compared with cells grown under white light, the relative number of PBSs to photosystems (PSs) increased in blue-light-grown cells and decreased in green-, yellow-, and red-light-grown cells. Moreover, the PBS number increased with increment in the monochromatic light intensity. More energy was transferred from PBSs to PSII than to PSI under blue light, whereas energy transfer from PBSs to PSII was reduced under green and yellow lights, and energy transfer from the PBSs to both PSs decreased under red light. Decoupling of PBSs was induced by intense green, yellow, and red lights. Energy transfer from PSII to PSI (spillover) was observed, but the contribution of the spillover did not distinctly change depending on the culture light intensity and quality. These results suggest that the glaucophyte C. paradoxa modifies the light-harvesting abilities of both PSs and excitation energy-transfer processes between the light-harvesting antennas and both PSs during long-term light adaption.

Structure of a monomeric photosystem I core associated with iron-stress-induced-A proteins from Anabaena sp. PCC 7120.

Iron-stress-induced-A proteins (IsiAs) are expressed in cyanobacteria under iron-deficient conditions. The cyanobacterium Anabaena sp. PCC 7120 has four isiA genes; however, their binding property and functional roles in PSI are still missing. We analyzed a cryo-electron microscopy structure of a PSI-IsiA supercomplex isolated from Anabaena grown under an iron-deficient condition. The PSI-IsiA structure contains six IsiA subunits associated with the PsaA side of a PSI core monomer. Three of the six IsiA subunits were identified as IsiA1 and IsiA2. The PSI-IsiA structure lacks a PsaL subunit; instead, a C-terminal domain of IsiA2 occupies the position of PsaL, which inhibits the oligomerization of PSI, leading to the formation of a PSI monomer. Furthermore, excitation-energy transfer from IsiAs to PSI appeared with a time constant of 55 ps. These findings provide insights into both the molecular assembly of the Anabaena IsiA family and the functional roles of IsiAs.

Biochemical and spectroscopic characterization of PSI-LHCI from the red alga Cyanidium caldarium.

Light-harvesting complexes (LHCs) have been diversified in oxygenic photosynthetic organisms, and play an essential role in capturing light energy which is transferred to two types of photosystem cores to promote charge-separation reactions. Red algae are one of the groups of photosynthetic eukaryotes, and their chlorophyll (Chl) a-binding LHCs are specifically associated with photosystem I (PSI). In this study, we purified three types of preparations, PSI-LHCI supercomplexes, PSI cores, and isolated LHCIs, from the red alga Cyanidium caldarium, and examined their properties. The polypeptide bands of PSI-LHCI showed characteristic PSI and LHCI components without contamination by other proteins. The carotenoid composition of LHCI displayed zeaxanthins, ß-cryptoxanthins, and ß-carotenes. Among the carotenoids, zeaxanthins were enriched in LHCI. On the contrary, both zeaxanthins and ß-cryptoxanthins could not be detected from PSI, suggesting that zeaxanthins and ß-cryptoxanthins are bound to LHCI but not PSI. A Qy peak of Chl a in the absorption spectrum of LHCI was shifted to a shorter wavelength than those in PSI and PSI-LHCI. This tendency is in line with the result of fluorescence-emission spectra, in which the emission maxima of PSI-LHCI, PSI, and LHCI appeared at 727, 719, and 677 nm, respectively. Time-resolved fluorescence spectra of LHCI represented no 719 and 727-nm fluorescence bands from picoseconds to nanoseconds. These results indicate that energy levels of Chls around/within LHCIs and within PSI are changed by binding LHCIs to PSI. Based on these findings, we discuss the expression, function, and structure of red algal PSI-LHCI supercomplexes.

Reversible down-regulation of photosystems I and II leads to fast photosynthesis recovery after long-term drought in Jatropha curcas.

Jatropha curcas is a drought-tolerant plant that maintains its photosynthetic pigments under prolonged drought, and quickly regains its photosynthetic capacity when water is available. It has been reported that drought stress leads to increased thermal dissipation in PSII, but that of PSI has been barely investigated, perhaps due to technical limitations in measuring the PSI absolute quantum yield. In this study, we combined biochemical analysis and spectroscopic measurements using an integrating sphere, and verified that the quantum yields of both photosystems are temporarily down-regulated under drought. We found that the decrease in the quantum yield of PSII was accompanied by a decrease in the core complexes of PSII while light-harvesting complexes are maintained under drought. In addition, in drought-treated plants, we observed a decrease in the absolute quantum yield of PSI as compared with the well-watered control, while the amount of PSI did not change, indicating that non-photochemical quenching occurs in PSI. The down-regulation of both photosystems was quickly lifted in a few days upon re-watering. Our results indicate, that in J. curcas under drought, the down-regulation of both PSII and PSI quantum yield protects the photosynthetic machinery from uncontrolled photodamage.

Excitation relaxation dynamics of carotenoids constituting the diadinoxanthin cycle.

Carotenoids (Cars) exhibit two functions in photosynthesis, light-harvesting and photoprotective functions, which are performed through the excited states of Cars. Therefore, increasing our knowledge on excitation relaxation dynamics of Cars is important for understanding of the functions of Cars. In light-harvesting complexes, there exist Cars functioning by converting the π-conjugation number in response to light conditions. It is well known that some microalgae have a mechanism controlling the conjugation number of Cars, called as the diadinoxanthin cycle; diadinoxanthin (10 conjugations) is accumulated under low light, whereas diatoxanthin (11 conjugations) appears under high light. However, the excitation relaxation dynamics of these two Cars have not been clarified. In the present study, we investigated excitation relaxation dynamics of diadinoxanthin and diatoxanthin in relation to their functions, by the ultrafast fluorescence spectroscopy. After an excitation to the S2 state, the intramolecular vibrational redistribution occurs, followed by the internal conversion to the S1 state. The S2 lifetimes were analyzed to be 175 fs, 155 fs, and 140 fs in diethyl ether, ethanol, and acetone, respectively, for diadinoxanthin, and 155 fs, 135 fs, and 125 fs in diethyl ether, ethanol, and acetone, respectively for diatoxanthin. By converting diadinoxanthin to diatoxanthin, the absorption spectra shift to longer wavelengths by 5-7 nm, and lifetimes of S2 and S1 states decrease by 11-13% and 52%, respectively. Differences in levels and lifetimes of excited states between diadinoxanthin and diatoxanthin are small; therefore, it is suggested that changes in the energy level of chlorophyll a are necessary to efficiently control the functions of the diadinoxanthin cycle.

Similar fish species composition despite larger environmental heterogeneity during severe hypoxia in a coastal ecosystem.

Environmental heterogeneity is one of the most influential factors that create compositional variation among local communities. Greater compositional variation is expected when an environmental gradient encompasses the most severe conditions where species sorting is more likely to operate. However, evidence for stronger species sorting at severer environment has typically been obtained for less mobile organisms and tests are scarce for those with higher dispersal ability that allows individuals to sensitively respond to environmental stress. Here, with the dynamics of fish communities in a Japanese bay revealed by environmental DNA metabarcoding analyses as a model case, we tested the hypothesis that larger environmental heterogeneity caused by severe seasonal hypoxia (lower concentration of oxygen in bottom waters in summer) leads to larger variation of species composition among communities. During summer, fish species richness was lower in the bottom layer, suggesting the severity of the hypoxic bottom water. In contrast to the prediction, we found that although the environmental parameters of bottom and surface water was clearly distinct in summer, fish species composition was more similar between the two layers. Our null model analysis suggested that the higher compositional similarity during hypoxia season was not a result of the sampling effect reflecting differences in the alpha or gamma diversity. Furthermore, a shift in the species occurrence from bottom to surface layers was observed during hypoxia season, which was consistent across species, suggesting that the severe condition in the bottom adversely affected fish species irrespective of their identity. These results suggest that larger environmental heterogeneity does not necessarily lead to higher compositional variation once the environmental gradient encompasses extremely severe conditions. This is most likely because individual organisms actively avoided the severity quasi-neutrally, which induced mass effect-like dispersal and lead to the mixing of species composition across habitats. By showing counter evidence against the prevailing view, we provide novel insights into how species sorting by environment acts in heterogeneous and severe conditions.

Structural basis for the absence of low-energy chlorophylls in a photosystem I trimer from Gloeobacter violaceus.

Photosystem I (PSI) is a multi-subunit pigment-protein complex that functions in light-harvesting and photochemical charge-separation reactions, followed by reduction of NADP to NADPH required for CO2 fixation in photosynthetic organisms. PSI from different photosynthetic organisms has a variety of chlorophylls (Chls), some of which are at lower-energy levels than its reaction center P700, a special pair of Chls, and are called low-energy Chls. However, the sites of low-energy Chls are still under debate. Here, we solved a 2.04-Å resolution structure of a PSI trimer by cryo-electron microscopy from a primordial cyanobacterium Gloeobacter violaceus PCC 7421, which has no low-energy Chls. The structure shows the absence of some subunits commonly found in other cyanobacteria, confirming the primordial nature of this cyanobacterium. Comparison with the known structures of PSI from other cyanobacteria and eukaryotic organisms reveals that one dimeric and one trimeric Chls are lacking in the Gloeobacter PSI. The dimeric and trimeric Chls are named Low1 and Low2, respectively. Low2 is missing in some cyanobacterial and eukaryotic PSIs, whereas Low1 is absent only in Gloeobacter. These findings provide insights into not only the identity of low-energy Chls in PSI, but also the evolutionary changes of low-energy Chls in oxyphototrophs.

Structural basis for different types of hetero-tetrameric light-harvesting complexes in a diatom PSII-FCPII supercomplex.

Fucoxanthin chlorophyll (Chl) a/c-binding proteins (FCPs) function as light harvesters in diatoms. The structure of a diatom photosystem II-FCPII (PSII-FCPII) supercomplex have been solved by cryo-electron microscopy (cryo-EM) previously; however, the FCPII subunits that constitute the FCPII tetramers and monomers are not identified individually due to their low resolutions. Here, we report a 2.5 Å resolution structure of the PSII-FCPII supercomplex using cryo-EM. Two types of tetrameric FCPs, S-tetramer, and M-tetramer, are identified as different types of hetero-tetrameric complexes. In addition, three FCP monomers, m1, m2, and m3, are assigned to different gene products of FCP. The present structure also identifies the positions of most Chls c and diadinoxanthins, which form a complicated pigment network. Excitation-energy transfer from FCPII to PSII is revealed by time-resolved fluorescence spectroscopy. These structural and spectroscopic findings provide insights into an assembly model of FCPII and its excitation-energy transfer and quenching processes.

Structure of a tetrameric photosystem I from a glaucophyte alga Cyanophora paradoxa.

Photosystem I (PSI) is one of the two photosystems functioning in light-energy harvesting, transfer, and electron transfer in photosynthesis. However, the oligomerization state of PSI is variable among photosynthetic organisms. We present a 3.8-Å resolution cryo-electron microscopic structure of tetrameric PSI isolated from the glaucophyte alga Cyanophora paradoxa, which reveals differences with PSI from other organisms in subunit composition and organization. The PSI tetramer is organized in a dimer of dimers with a C2 symmetry. Unlike cyanobacterial PSI tetramers, two of the four monomers are rotated around 90°, resulting in a completely different pattern of monomer-monomer interactions. Excitation-energy transfer among chlorophylls differs significantly between Cyanophora and cyanobacterial PSI tetramers. These structural and spectroscopic features reveal characteristic interactions and excitation-energy transfer in the Cyanophora PSI tetramer, suggesting that the Cyanophora PSI could represent a turning point in the evolution of PSI from prokaryotes to eukaryotes.

Characterization of photosystem II assembly complexes containing ONE-HELIX PROTEIN1 in Arabidopsis thaliana.

The assembly process of photosystem II (PSII) requires several auxiliary proteins to form assembly intermediates. In plants, early assembly intermediates comprise D1 and D2 subunits of PSII together with a few auxiliary proteins including at least ONE-HELIX PROTEIN1 (OHP1), OHP2, and HIGH-CHLOROPHYLL FLUORESCENCE 244 (HCF244) proteins. Herein, we report the basic characterization of the assembling intermediates, which we purified from Arabidopsis transgenic plants overexpressing a tagged OHP1 protein and named the OHP1 complexes. We analyzed two major forms of OHP1 complexes by mass spectrometry, which revealed that the complexes consist of OHP1, OHP2, and HCF244 in addition to the PSII subunits D1, D2, and cytochrome b559. Analysis of chlorophyll fluorescence showed that a major form of the complex binds chlorophyll a and carotenoids and performs quenching with a time constant of 420 ps. To identify the localization of the auxiliary proteins, we solubilized thylakoid membranes using a digitonin derivative, glycodiosgenin, and separated them into three fractions by ultracentrifugation, and detected these proteins in the loose pellet containing the stroma lamellae and the grana margins together with two chlorophyll biosynthesis enzymes. The results indicated that chlorophyll biosynthesis and assembly may take place in the same compartments of thylakoid membranes. Inducible suppression of the OHP2 mRNA substantially decreased the OHP2 protein in mature Arabidopsis leaves without a significant reduction in the maximum quantum yield of PSII under low-light conditions, but it compromised the yields under high-light conditions. This implies that the auxiliary protein is required for acclimation to high-light conditions.

Excitation-energy transfer in heterocysts isolated from the cyanobacterium Anabaena sp. PCC 7120 as studied by time-resolved fluorescence spectroscopy.

Heterocysts are formed in filamentous heterocystous cyanobacteria under nitrogen-starvation conditions, and possess a very low amount of photosystem II (PSII) complexes than vegetative cells. Molecular, morphological, and biochemical characterizations of heterocysts have been investigated; however, excitation-energy dynamics in heterocysts are still unknown. In this study, we examined excitation-energy-relaxation processes of pigment-protein complexes in heterocysts isolated from the cyanobacterium Anabaena sp. PCC 7120. Thylakoid membranes from the heterocysts showed no oxygen-evolving activity under our experimental conditions and no thermoluminescence-glow curve originating from charge recombination of S2QA-. Two dimensional blue-native/SDS-PAGE analysis exhibits tetrameric, dimeric, and monomeric photosystem I (PSI) complexes but almost no dimeric and monomeric PSII complexes in the heterocyst thylakoids. The steady-state fluorescence spectrum of the heterocyst thylakoids at 77 K displays both characteristic PSI fluorescence and unusual PSII fluorescence different from the fluorescence of PSII dimer and monomer complexes. Time-resolved fluorescence spectra at 77 K, followed by fluorescence decay-associated spectra, showed different PSII and PSI fluorescence bands between heterocysts and vegetative thylakoids. Based on these findings, we discuss excitation-energy-transfer mechanisms in the heterocysts.

Electrostatic charge controls the lowest LH1 Qy transition energy in the triply extremophilic purple phototrophic bacterium, Halorhodospira halochloris.

Halorhodospira (Hlr.) halochloris is a unique phototrophic purple bacterium because it is a triple extremophile-the organism is thermophilic, alkalophilic, and halophilic. The most striking photosynthetic feature of Hlr. halochloris is that the bacteriochlorophyll (BChl) b-containing core light-harvesting (LH1) complex surrounding its reaction center (RC) exhibits its LH1 Qy absorption maximum at 1016 nm, which is the lowest transition energy among phototrophic organisms. Here we report that this extraordinarily red-shifted LH1 Qy band of Hlr. halochloris exhibits interconvertible spectral shifts depending on the electrostatic charge distribution around the BChl b molecules. The 1016 nm band of the Hlr. halochloris LH1-RC complex was blue-shifted to 958 nm upon desalting or pH decrease but returned to its original position when supplemented with salts or pH increase. Resonance Raman analysis demonstrated that these interconvertible spectral shifts are not associated with the strength of hydrogen-bonding interactions between BChl b and LH1 polypeptides. Furthermore, circular dichroism signals for the LH1 Qy transition of Hlr. halochloris appeared with a positive sign (as in BChl b-containing Blastochloris species) and opposite those of BChl a-containing purple bacteria, possibly due to a combined effect of slight differences in the transition dipole moments between BChl a and BChl b and in the interactions between adjacent BChls in their assembled state. Based on these findings and LH1 amino acid sequences, it is proposed that Hlr. halochloris evolved its unique and tunable light-harvesting system with electrostatic charges in order to carry out photosynthesis and thrive in its punishing hypersaline and alkaline habitat.

Proteome analysis of response to different spectral light irradiation in Synechocystis sp. PCC 6803.

In cyanobacteria, it is known that the excitation ratios of photosystem (PS) I and PSII changes with the wavelength of irradiated light due to mobile phycobilisome (PBS) and spillover, affecting the photosynthetic ATP/NADPH synthesis ratio and metabolic flux state. However, the mechanisms by which these changes are controlled have not been well studied. In this study, we performed a targeted proteomic analysis of Synechocystis sp. PCC 6803 under different spectral light conditions to clarify the regulation mechanisms of mobile PBS, spillover and metabolisms under different light qualities at the protein level. The results showed an increase in the amount of proteins mainly involved in CO2 fixation under Red1 light conditions with a high specific growth rate, suggesting that the rate of intracellular metabolism is controlled by the rate of carbon uptake, not by changes in the amount of each enzyme. Correlation analysis between protein levels and PSI/PSII excitation ratios revealed that PsbQUY showed high correlations and significantly increased under Blue and Red2 light conditions, where the PSI/PSII excitation ratio was higher due to spillover. In the strains lacking the genes encoding these proteins, a decrease in the PSI/PSII excitation ratio was observed, suggesting that PsbQUY contribute to spillover occurrence. SIGNIFICANCE: In cyanobacteria, the photosynthetic apparatus's responses, such as state transition [mobile PBS and spillover], occur due to the intensity and wavelength of irradiated light, resulting in changes in photosynthetic electron transport and metabolic flux states. Previous studies have analyzed the response of Synechocystis sp. PCC 6803 to light intensity from various directions, but only spectroscopic analysis of the photosynthetic apparatus has been done on the response to changes in the wavelength of irradiated light. This study analyzed the response mechanisms of mobile PBS, spillover, photosynthetic, and metabolic systems in Synechocystis sp. PCC 6803 under six different spectral light conditions by a targeted proteomic analysis. As a result, many proteins were successfully quantified, and the metabolic enzymes and photosynthetic apparatus were analyzed using an integrated approach. Principal component and correlation analyses and volcano plots revealed that the PSII subunits PsbQ, PsbU, and PsbY have a strong correlation with the PSI/PSII excitation ratio and contribute to spillover occurrence. Thus, statistical analysis based on proteome data revealed that PsbQ, PsbU, and PsbY are involved in spillover, as revealed by spectroscopic analysis.

High-light modification of excitation-energy-relaxation processes in the green flagellate Euglena gracilis.

Photosynthetic organisms finely tune their photosynthetic machinery including pigment compositions and antenna systems to adapt to various light environments. However, it is poorly understood how the photosynthetic machinery in the green flagellate Euglena gracilis is modified under high-light conditions. In this study, we examined high-light modification of excitation-energy-relaxation processes in Euglena cells. Oxygen-evolving activity in the cells incubated at 300 µmol photons m-2 s-1 (HL cells) cannot be detected, reflecting severe photodamage to photosystem II (PSII) in vivo. Pigment compositions in the HL cells showed relative increases in 9'-cis-neoxanthin, diadinoxanthin, and chlorophyll b compared with the cells incubated at 30 µmol photons m-2 s-1 (LL cells). Absolute fluorescence spectra at 77 K exhibit smaller intensities of the PSII and photosystem I (PSI) fluorescence in the HL cells than in the LL cells. Absolute fluorescence decay-associated spectra at 77 K of the HL cells indicate suppression of excitation-energy transfer from light-harvesting complexes (LHCs) to both PSI and PSII with the time constant of 40 ps. Rapid energy quenching in LHCs and PSII in the HL cells is distinctly observed by averaged Chl-fluorescence lifetimes. These findings suggest that Euglena modifies excitation-energy-relaxation processes in addition to pigment compositions to deal with excess energy. These results provide insights into the photoprotection strategies of this alga under high-light conditions.

Mutations in hik26 and slr1916 lead to high-light stress tolerance in Synechocystis sp. PCC6803.

Increased tolerance to light stress in cyanobacteria is a desirable feature for their applications. Here, we obtained a high light tolerant (Tol) strain of Synechocystis sp. PCC6803 through an adaptive laboratory evolution, in which the cells were repeatedly sub-cultured for 52 days under high light stress conditions (7000 to 9000 µmol m-2 s-1). Although the growth of the parental strain almost stopped when exposed to 9000 µmol m-2 s-1, no growth inhibition was observed in the Tol strain. Excitation-energy flow was affected because of photosystem II damage in the parental strain under high light conditions, whereas the damage was alleviated and normal energy flow was maintained in the Tol strain. The transcriptome data indicated an increase in isiA expression in the Tol strain under high light conditions. Whole genome sequence analysis and reverse engineering revealed two mutations in hik26 and slr1916 involved in high light stress tolerance in the Tol strain.

Role of type I NADH dehydrogenase in Synechocystis sp. PCC 6803 under phycobilisome excited red light.

Cyanobacterial type I NADH dehydrogenase (NDH-1) is involved in various bioenergetic reactions including respiration, cyclic electron transport (CET), and CO2 uptake. The role of NDH-1 is usually minor under normal growth conditions and becomes important under stress conditions. However, in our previous study, flux balance analysis (FBA) simulation predicted that the drive of NDH-1 as CET pathway with a photosystem (PS) I/PSII excitation ratio around 1.0 contributes to achieving an optimal specific growth rate. In this study, to experimentally elucidate the predicted functions of NDH-1, first, we measured the PSI/PSII excitation ratios of Synechocystis sp. PCC 6803 grown under four types of spectral light conditions. The specific growth rate was the highest and PSI/PSII excitation ratio was with 0.88 under the single-peak light at 630 nm (Red1). Considering this measured excitation ratios, FBA simulation predicted that NDH-1-dependent electron transport was the major pathway under Red1. Moreover, in actual culture, an NDH-1 deletion strain had slower growth rate than that of wild type only under Red1 light condition. Therefore, we experimentally demonstrated that NDH-1 plays an important role under optimal light conditions such as Red1 light, where Synechocystis exhibits the highest specific growth rate and PSI/PSII excitation ratio of around 1.0.

Molecular organizations and function of iron-stress-induced-A protein family in Anabaena sp. PCC 7120.

Iron-stress-induced-A proteins (IsiAs) are expressed in cyanobacteria under iron-deficient conditions, and surround photosystem I (PSI) trimer with a ring formation. A cyanobacterium Anabaena sp. PCC 7120 has four isiA genes; however, it is unknown how the IsiAs are associated with PSI. Here we report on molecular organizations and function of the IsiAs in this cyanobacterium. A deletion mutant of the isiA1 gene was constructed, and the four types of thylakoids were prepared from the wild-type (WT) and ΔisiA1 cells under iron-replete (+Fe) and iron-deficient (-Fe) conditions. Immunoblotting analysis exhibits a clear expression of the IsiA1 in the WT-Fe. The PSI-IsiA1 supercomplex is found in the WT-Fe, and excitation-energy transfer from IsiA1 to PSI is verified by time-resolved fluorescence analyses. Instead of the IsiA1, both IsiA2 and IsiA3 are bound to PSI monomer in the ΔisiA1-Fe. These findings provide insights into multiple-expression system of the IsiA family in this cyanobacterium.

Enhancement of excitation-energy quenching in fucoxanthin chlorophyll a/c-binding proteins isolated from a diatom Phaeodactylum tricornutum upon excess-light illumination.

Photosynthetic organisms regulate pigment composition and molecular oligomerization of light-harvesting complexes in response to solar light intensities, in order to improve light-harvesting efficiency. Here we report excitation-energy dynamics and relaxation of fucoxanthin chlorophyll a/c-binding protein (FCP) complexes isolated from a diatom Phaeodactylum tricornutum grown under high-light (HL) illumination. Two types of FCP complexes were prepared from this diatom under the HL condition, whereas one FCP complex was isolated from the cells grown under a low-light (LL) condition. The subunit composition and oligomeric states of FCP complexes under the HL condition are different from those under the LL condition. Absorption and fluorescence spectra at 77 K of the FCP complexes also vary between the two conditions, indicating modifications of the pigment composition and arrangement upon the HL illumination. Time-resolved fluorescence curves at 77 K of the FCP complexes under the HL condition showed shorter lifetime components compared with the LL condition. Fluorescence decay-associated spectra at 77 K showed distinct excitation-energy-quenching components and alterations of energy-transfer pathways in the FCP complexes under the HL condition. These findings provide insights into molecular and functional mechanisms of the dynamic regulation of FCPs in this diatom under excess-light conditions.

Basic pH-induced modification of excitation-energy dynamics in fucoxanthin chlorophyll a/c-binding proteins isolated from a pinguiophyte, Glossomastix chrysoplasta.

Photosynthetic organisms have diversified light-harvesting complexes (LHCs) to collect solar energy efficiently, leading to an acquisition of their ecological niches. Herein we report on biochemical and spectroscopic characterizations of fucoxanthin chlorophyll a/c-binding protein (FCP) complexes isolated from a marine pinguiophyte Glossomastix chrysoplasta. The pinguiophyte FCP showed one subunit band in SDS-PAGE and one protein-complex band with a molecular weight at around 66 kDa in clear-native PAGE. By HPLC analysis, the FCP possesses chlorophylls a and c, fucoxanthin, and violaxanthin. To clarify excitation-energy-relaxation processes in the FCP, we measured time-resolved fluorescence spectra at 77 K of the FCP adapted to pH 5.0, 6.5, and 8.0. Fluorescence curves measured at pH 5.0 and 8.0 showed shorter lifetime components compared with those at pH 6.5. The rapid decay components at pH 5.0 and 8.0 are unveiled by fluorescence decay-associated (FDA) spectra; fluorescence decays occur in the 270 and 160-ps FDA spectra only at pH 5.0 and 8.0, respectively. In addition, energy-transfer pathways with time constants of tens of picoseconds are altered under the basic pH condition but not the acidic pH condition. These findings provide novel insights into pH-dependent energy-transfer and energy-quenching machinery in not only FCP family but also photosynthetic LHCs.

Estimation of linear and cyclic electron flows in photosynthesis based on 13C-metabolic flux analysis.

Photosynthetic organisms produce ATP and NADPH using light as an energy source and further utilize these cofactors during metabolism. Photosynthesis involves linear and cyclic electron flows; as the cyclic electron flow produces ATP more effectively than the linear electron flow without NADPH, the cell efficiently adjusts ATP and NADPH production using the two different pathways. Nevertheless, direct measurement of ATP and NADPH production during photosynthesis has been difficult. In the present study, the photosynthetic ATP and NADPH production rates of Synechocystis sp. PCC 6803 under three different single peak wavelength lights (blue: 470 nm, R630: 630 nm, and R680: 680 nm) were evaluated based on 13C-metabolic flux analysis (13C-MFA) by considering the mass balance of ATP and NADPH between photosynthesis and metabolism. The ratios of ATP/NADPH production via photosynthesis were estimated as 3.13, 1.70, and 2.10 under blue, R630, and R680 light conditions, respectively. Moreover, the linear and cyclic electron flow ratios were estimated to be 1.1-2.2, 0.2-0.5, and 0.5-1.0 under blue, R630, and R680 light conditions, respectively. The predicted linear and cyclic electron flow ratios were consistent with the excitation ratio between photosystems I and II, as observed in the steady-state fluorescence spectra.