chlorophyll-a and violaxanthin

The distinctive lateral organization of the protein complexes in the thylakoid membrane investigated by Jan Anderson and co-workers is dependent on the balance of various attractive and repulsive forces. Modulation of these forces allows critical physiological regulation of photosynthesis that provides efficient light-harvesting in limiting light but dissipation of excess potentially damaging radiation in saturating light. The light-harvesting complexes (LHCII) are central to this regulation, which is achieved by phosphorylation of stromal residues, protonation on the lumen surface and de-epoxidation of bound violaxanthin. The functional flexibility of LHCII derives from a remarkable pigment composition and configuration that not only allow efficient absorption of light and efficient energy transfer either to photosystem II or photosystem I core complexes, but through subtle configurational changes can also exhibit highly efficient dissipative reactions involving chlorophyll-xanthophyll and/or chlorophyll-chlorophyll interactions. These changes in function are determined at a macroscopic level by alterations in protein-protein interactions in the thylakoid membrane. The capacity and dynamics of this regulation are tuned to different physiological scenarios by the exact protein and pigment content of the light-harvesting system. Here, the molecular mechanisms involved will be reviewed, and the optimization of the light-harvesting system in different environmental conditions described.

Topics: Arabidopsis; Chlorophyll; Cold Temperature; Electron Transport; Light-Harvesting Protein Complexes; Phosphorylation; Photochemical Processes; Photosynthesis; Photosystem I Protein Complex; Photosystem II Protein Complex; Protein Interaction Mapping; Sunlight; Thylakoids; Xanthophylls

Carotenoids represent the first line of defence of photosystems against singlet oxygen (

Topics: Arabidopsis; Carotenoids; Chlorophyll; Energy Transfer; Light-Harvesting Protein Complexes; Lutein; Photosystem II Protein Complex; Xanthophylls; Zeaxanthins

Plants use energy from the sun yet also require protection against the generation of deleterious photoproducts from excess energy. Photoprotection in green plants, known as nonphotochemical quenching (NPQ), involves thermal dissipation of energy and is activated by a series of interrelated factors: a pH drop in the lumen, accumulation of the carotenoid zeaxanthin (Zea), and formation of arrays of pigment-containing antenna complexes. However, understanding their individual contributions and their interactions has been challenging, particularly for the antenna arrays, which are difficult to manipulate in vitro. Here, we achieved systematic and discrete control over the array size for the principal antenna complex, light-harvesting complex II, using near-native in vitro membranes called nanodiscs. Each of the factors had a distinct influence on the level of dissipation, which was characterized by measurements of fluorescence quenching and ultrafast chlorophyll-to-carotenoid energy transfer. First, an increase in array size led to a corresponding increase in dissipation; the dramatic changes in the chlorophyll dynamics suggested that this is due to an allosteric conformational change of the protein. Second, a pH drop increased dissipation but exclusively in the presence of protein-protein interactions. Third, no Zea dependence was identified which suggested that Zea regulates a distinct aspect of NPQ. Collectively, these results indicate that each factor provides a separate type of control knob for photoprotection, which likely enables a flexible and tunable response to solar fluctuations.

Topics: Carotenoids; Chlorophyll; Energy Transfer; Hydrogen-Ion Concentration; Light; Light-Harvesting Protein Complexes; Nanostructures; Protein Binding; Protein Multimerization; Spinacia oleracea; Thylakoids; Xanthophylls; Zeaxanthins

Life on Earth depends on photosynthesis, the conversion of light energy into chemical energy. Plants collect photons by light harvesting complexes (LHC)-abundant membrane proteins containing chlorophyll and xanthophyll molecules. LHC-like proteins are similar in their amino acid sequence to true LHC antennae, however, they rather serve a photoprotective function. Whether the LHC-like proteins bind pigments has remained unclear. Here, we characterize plant LHC-like proteins (LIL3 and ELIP2) produced in the cyanobacterium Synechocystis sp. PCC 6803 (hereafter Synechocystis). Both proteins were associated with chlorophyll a (Chl) and zeaxanthin and LIL3 was shown to be capable of quenching Chl fluorescence via direct energy transfer from the Chl Q

Topics: Arabidopsis Proteins; Carotenoids; Chlorophyll; Chloroplast Proteins; Energy Transfer; Mutation; Protein Binding; Protein Folding; Protein Multimerization; Synechocystis; Xanthophylls; Zeaxanthins

In the present study, the effects of blue LED light on the regreening of citrus fruit were investigated in an in vitro system of Valencia orange flavedos. The results showed that blue LED light irradiation induced regreening in the flavedos. After four-week culture in vitro, the flavedos exhibited obviously green color in the blue LED light treatment, while the flavedos in the control were still in orange color. During the regreening process, the blue LED light treatment induced chlorophyll accumulation, and substantially altered the carotenoid composition in the flavedos. Compared with the control, the content of 9-cis-violaxanthin was decreased, while the contents of lutein, β-carotene, and all-trans-violaxanthin were increased by blue LED light. In addition, gene expression results showed that the up-regulation of CitLCYe and down-regulation of CitLCYb2 by blue LED light led to a shift from β,β-branch to β,ε-branch of the carotenoid biosynthetic pathway.

Topics: beta Carotene; Carotenoids; Chlorophyll; Citrus; Citrus sinensis; Fruit; Gene Expression Regulation, Plant; Light; Pigmentation; Xanthophylls

Antenna complexes in photosystems of plants and green algae are able to switch between a light-harvesting unquenched conformation and a quenched conformation so to avoid photodamage. When the switch is activated, nonphotochemical quenching (NPQ) mechanisms take place for an efficient deactivation of excess excitation energy. The molecular details of these mechanisms have not been fully clarified but different hypotheses have been proposed. Among them, a popular one involves excitation energy transfer (EET) from the singlet excited Chls to the lowest singlet state (S

Topics: Chlorophyll; Energy Transfer; Light-Harvesting Protein Complexes; Photosynthesis; Photosystem II Protein Complex; Plant Proteins; Plants; Xanthophylls; Zeaxanthins

Light, or visible radiation, serves as a source of energy for photosynthesis of plants and most algae. In addition, light and ultraviolet radiation (UV-A and UV-B) act as a biological signal, triggering several cellular processes that are mediated by photoreceptors. The aim of this study was to evaluate the physiological and biochemical responses of Osmundea pinnatifida driven by different radiations through putative photoreceptors. For this, O. pinnatifida was grown under different radiation treatments composed by high intensity of light emitted by a low pressure sodium lamp (SOX), aiming to saturate photosynthesis, which was supplemented by low intensities of visible (red, green and blue) and ultraviolet radiation (UV-A and UV-B), in order to activate photoreceptors. Growth rates, photosynthesis, antioxidant activity, polyphenols, soluble proteins, phycobiliproteins, mycosporine-like amino acids (MAAs) and carotenoids were evaluated during the experiment. Complementary UV-A radiation positively influenced growth rates after 15 days of experiment, although the presence of a peak of blue light in this treatment can also have contributed. UV-B radiation increased the concentration of zeaxanthin and chlorophyll a. The blue light caused the accumulation of chlorophyll a, violaxanthin, phycoerythrin and polyphenols on different days of the experiment. Phycoerythrin also increased under green and red light conditions. Our results showed that some compounds can be modulated by different radiation, and the involvement of photoreceptors is suggested. In red algae, photoreceptors sensitive to red, green and blue light have been identified, however little is known about UV photoreceptors. The presence of photoreceptors sensitive to UV radiation in O. pinnatifida is discussed.

Topics: Antioxidants; Carotenoids; Chlorophyll; Photosynthesis; Phycoerythrin; Plant Proteins; Polyphenols; Rhodophyta; Ultraviolet Rays; Xanthophylls

The influence of six different light regimes throughout the photosynthetically active radiation range (from 400 to 700 nm, including blue, green, yellow, red-orange, red, and white) at two intensities (100 and 300 µmol photons m

Topics: beta Carotene; Chlorophyll; Diatoms; Light; Photosynthesis; Pigments, Biological; Xanthophylls; Zeaxanthins

The aim of this study was to evaluate the impact of selected types of LED (light emitting diodes) lighting on the quality of alfalfa sprouts. In the experiment, cold white, warm white and multicolour: (red, green, blue-RGB) LEDs were applied, and dispersed sunlight was used as a control. The product was examined for the yield and the contents of dry matter, total polyphenols, ascorbic acid, chlorophylls, β-carotene, lutein, neoxanthin and violaxanthin. Cotyledons' mass in the whole plant increased under LED illumination and was up to 50% greater for sprouts grown in RGB light compared to those cultivated in dispersed sunlight. The highest chlorophyll and carotenoid pigment contents in cotyledons were observed under RGB LED and cold white treatments. Similarly, RGB LEDs allows one to obtain the product with the highest level of total phenolic compounds. The highest ascorbic acid content was observed in sprouts growing under sunlight, followed by RGB.

Topics: beta Carotene; Chlorophyll; Chromatography, High Pressure Liquid; Germination; Light; Lutein; Medicago sativa; Polyphenols; Seedlings; Xanthophylls

Dandelion (Taraxacum officinale) var. Garnet Stem was harvested from Texas and New Jersey for identification, quantification of phytochemicals, measurement of free radical scavenging activity, and bile acid binding capacity. The red midrib and petioles were extracted with methanol or ethanol and with or without water in combination with four different acids such as formic, hydrochloric, acetic, and citric acid. LC-ESI-HR-QTOF-MS was used to identify four anthocyanins including cyanidin-3-glucoside, cyanidin-3-(6-malonyl)-glucoside (A-1), cyanidin-3-(6-malonyl)-glucoside (A-2), and peonidin-3-(malonyl)-glucoside for the 1st time. In New Jersey samples, vitamin C and β-carotene were highest in the leaf blades versus whole leaf and petioles. Samples from Texas had highest amount of lutein, violaxanthin, and chlorophyll a and b in leaf blades versus whole leaf and petioles. Maximum DPPH free scavenging activity was found in MeOH: water: acid (80:19:1) and the combination of FA with EtOH: water: acid (80:19:1) demonstrated the higher level of total phenolic. Among six bile acids, sodium chenodeoxycholate was bound maximum in both Texas and New Jersey samples. This is the first report of anthocyanin identification from the midvein and petiole of Garnet Stem dandelion and results suggested that the phytochemicals and nutrients are highest in the leaf but may vary the amount depending on harvest location.. Four anthocyanins in the red midrib and petioles of Garnet Stem could be a potential source for antioxidants and can be used as a source of natural food color.

Topics: Anthocyanins; Antioxidants; Ascorbic Acid; beta Carotene; Bile Acids and Salts; Chenodeoxycholic Acid; Chlorophyll; Chlorophyll A; Glucosides; Lutein; New Jersey; Phytochemicals; Plant Leaves; Plant Stems; Taraxacum; Texas; Xanthophylls

The light-harvesting chlorophyll

Topics: Arabidopsis; Arabidopsis Proteins; Chlorophyll; Light; Light-Harvesting Protein Complexes; Mutation; Photosynthesis; Photosystem II Protein Complex; Plant Leaves; Protein Binding; Spectrophotometry; Xanthophylls

Resonance Raman spectroscopy was used to evaluate pigment-binding site properties in the violaxanthin-chlorophyll-a-binding protein (VCP) from Nannochloropsis oceanica. The pigments bound to this antenna protein are chlorophyll-a, violaxanthin, and vaucheriaxanthin. The molecular structures of bound Chl-a molecules are discussed with respect to those of the plant antenna proteins LHCII and CP29, the crystal structures of which are known. We show that three populations of carotenoid molecules are bound by VCP, each of which is in an all-trans configuration. We assign the lower-energy absorption transition of each of these as follows. One violaxanthin population absorbs at 485 nm, while the second population is red-shifted and absorbs at 503 nm. The vaucheriaxanthin population absorbs at 525 nm, a position red-shifted by 2138 cm

Topics: Carotenoids; Carrier Proteins; Chlorophyll; Light-Harvesting Protein Complexes; Spectrum Analysis, Raman; Xanthophylls

Up to 40% of incident light was screened in red Berberis leaves in vivo by anthocyanins, resulting also in up to 40% reduction of light-limited photosynthesis. The biological function of anthocyanins in leaves has been strongly discussed, but the hypothesis of a screening function is favored by most authors. For an evaluation of the function as photoprotective pigments, a quantification of their screening of the mesophyll is important. Here, chlorophyll fluorescence excitation of leaves of a red and a green variety of Berberis thunbergii was used to estimate the extent of screening by anthocyanins at 545 nm and over the whole photosynthetically active wavelength range. Growth at high light (430 µmol m

Topics: Anthocyanins; Berberis; Chlorophyll; Fluorescence; Light; Photosynthesis; Plant Leaves; Xanthophylls

Violaxanthin-chlorophyll a protein (VCP) from Nannochloropsis oceanica is a Chl a-only member of the LHC family of light-harvesting proteins. VCP binds carotenoids violaxanthin (Vio), vaucheriaxanthin (Vau), and vaucheriaxanthin-ester (Vau-ester). Here we report on energy transfer pathways in the VCP complex. The overall carotenoid-to-Chla energy transfer has efficiency over 90%. Based on their energy transfer properties, the carotenoids in VCP can be divided into two groups; blue carotenoids with the lowest energy absorption band around 480nm and red carotenoids with absorption extended up to 530nm. Both carotenoid groups transfer energy efficiently from their S2 states, reaching efficiencies of ~70% (blue) and ~60% (red). The S1 pathway, however, is efficient only for the red carotenoid pool for which two S1 routes characterized by 0.33 and 2.4ps time constants were identified. For the blue carotenoids the S1-mediated pathway is represented only by a minor route likely involving a hot S1 state. The relaxed S1 state of blue carotenoids decays to the ground state within 21ps. Presence of a fraction of non-transferring red carotenoids with the S1 lifetime of 13ps indicates some specific carotenoid-protein interaction that must shorten the intrinsic S1 lifetime of Vio and/or Vau whose S1 lifetimes in methanol are 26 and 29ps, respectively. The VCP complex from N. oceanica is the first example of a light-harvesting complex binding only non-carbonyl carotenoids with carotenoid-to-chlorophyll energy transfer efficiency over 90%.

Topics: Carotenoids; Chlorophyll; Chlorophyll A; Energy Transfer; Light-Harvesting Protein Complexes; Microalgae; Xanthophylls

The photosynthetic apparatus of higher plants acclimates to irradiance. Among the features which are changing is the pool size of the pigments belonging to the violaxanthin cycle, in which zeaxanthin is formed. In high light grown leaves, the violaxanthin cycle pool size is up to five times larger than in low light. The changes are reversible on a time scale of several days. Since it has been published that violaxanthin cycle pigments do not transfer absorbed energy to chlorophyll, we hypothesized that excitation of chlorophyll fluorescence in the blue spectral region may be reduced in high light-acclimated leaves. Fluorescence excitation spectra of leaves of the Arabidopsis thaliana tt3 mutant showed strong differences between high and low light-acclimated plants from 430 to 520 nm. The resulting difference spectrum was similar to carotenoids but shifted by about 20 nm to higher wavelengths. A good correlation was observed between the fluorescence excitation ratio F 470/F 660 and the violaxanthin cycle pool size when leaves were acclimated to a range of irradiances. In parallel to the decline of F 470/F 660 with high light acclimation also the quantum yield of photosynthetic oxygen evolution in blue light decreased. The data confirm that violaxanthin cycle carotenoids do not transfer absorbed light to chlorophyll. It is proposed to use the ratio F 470/F 660 as an indicator for the light acclimation status of the chloroplasts in a leaf.

Topics: Acclimatization; Arabidopsis; Carotenoids; Chlorophyll; Energy Transfer; Fluorescence; Light; Mutation; Photosynthesis; Pigmentation; Plant Leaves; Xanthophylls; Zeaxanthins

Non-photochemical quenching (NPQ) is a photoprotective mechanism in light-harvesting antennae. NPQ is triggered by chloroplast thylakoid lumen acidification and is accompanied by violaxanthin de-epoxidation to zeaxanthin, which further stimulates NPQ. In the present study, we show that violaxanthin can act in the opposite direction to zeaxanthin because an increase in the concentration of violaxanthin reduced NPQ in the light-harvesting antennae of Chromera velia. The correlation overlapped with a similar relationship between violaxanthin and NPQ as observed in isolated higher plant light-harvesting complex II. The data suggest that violaxanthin in C. velia can act as an inhibitor of NPQ, indicating that violaxanthin has to be removed from the vicinity of the protein to reach maximal NPQ.

Topics: Alveolata; Chlorophyll; Fluorescence; Light-Harvesting Protein Complexes; Photochemical Processes; Time Factors; Xanthophylls

Plants harvest photons for photosynthesis using light-harvesting complexes (LHCs)-an array of chlorophyll proteins that can reversibly switch from harvesting to energy-dissipation mode to prevent over-excitation and damage of the photosynthetic apparatus. In unicellular algae and lower plants this process requires the LHCSR proteins which senses over-acidification of the lumen trough protonatable residues exposed to the thylakoid lumen to activate quenching reactions. Further activation is provided by replacement of the violaxanthin ligand with its de-epoxidized product, zeaxanthin, also induced by excess light. We have produced the ppLHCSR1 protein from Physcomitrella patens by over-expression in tobacco and purified it in either its violaxanthin- or the zeaxanthin-binding form with the aim of analyzing their spectroscopic properties at either neutral or acidic pH. Using femtosecond spectroscopy, we demonstrated that the energy dissipation is achieved by two distinct quenching mechanism which are both activated by low pH. The first is present in both ppLHCSR1-Vio and ppLHCSR1-Zea and is characterized by 30-40ps time constant. The spectrum of the quenching product is reminiscent of a carotenoid radical cation, suggesting that the pH-induced quenching mechanism is likely electron transfer from the carotenoid to the excited Chl a. In addition, a second quenching channel populating the S

Topics: Bryopsida; Chlorophyll; Electron Transport; Energy Transfer; Hydrogen-Ion Concentration; Kinetics; Light-Harvesting Protein Complexes; Models, Biological; Nicotiana; Photosynthesis; Plants, Genetically Modified; Protein Binding; Spectrum Analysis; Xanthophylls; Zeaxanthins

Aggregation induced conformational change of light harvesting antenna complexes is believed to constitute one of the pathways through which photosynthetic organisms can safely dissipate the surplus of energy while exposed to saturating light. In this study, Stark fluorescence (SF) spectroscopy is applied to minor antenna complexes (CP24, CP26 and CP29) both in their light-harvesting and energy-dissipating states to trace and characterize different species generated upon energy dissipation through aggregation (in-vitro) induced conformational change. SF spectroscopy could identify three spectral species in the dissipative state of CP24, two in CP26 and only one in CP29. The comprehensive analysis of the SF spectra yielded different sets of molecular parameters for the multiple spectral species identified in CP24 or CP26, indicating the involvement of different pigments in their formation. Interestingly, a species giving emission around the 730nm spectral region is found to form in both CP24 and CP26 following transition to the energy dissipative state, but not in CP29. The SF analyses revealed that the far red species has exceptionally large charge transfer (CT) character in the excited state. Moreover, the far red species was found to be formed invariably in both Zeaxanthin (Z)- and Violaxathin (V)-enriched CP24 and CP26 antennas with identical CT character but with larger emission yield in Z-enriched ones. This suggests that the carotenoid Z is not directly involved but only confers an allosteric effect on the formation of the far red species. Similar far red species with remarkably large CT character were also observed in the dissipative state of the major light harvesting antenna (LHCII) of plants [Wahadoszamen et al. PCCP, 2012], the fucoxanthin-chlorophyll protein (FCP) of brown algae [Wahadoszamen et al. BBA, 2014] and cyanobacterial IsiA [Wahadoszamen et al. BBA, 2015], thus pointing to identical sites and pigments active in the formation of the far red quenching species in different organisms.

Topics: Chlorophyll; Energy Transfer; Light; Light-Harvesting Protein Complexes; Photosynthesis; Protein Conformation; Species Specificity; Spectrometry, Fluorescence; Spinacia oleracea; Structure-Activity Relationship; Xanthophylls; Zeaxanthins

Assessing photosynthesis rates at the ecosystem scale and over large regions is important for tracking the global carbon cycle and remote sensing has provided new and useful approaches for performing this assessment. The photochemical reflectance index (PRI) is a good estimator of short-term light-use efficiency (LUE) at the leaf scale; however, confounding factors appear at larger temporal and spatial scales. In this study, canopy-scale PRI variability was investigated for three species (Fagus sylvatica L., Quercus robur L. and Pinus sylvestris L.) growing under contrasting soil moisture conditions. Throughout the growing season, no significant differences in chlorophyll content and in violaxanthin, antheraxanthin and zeaxanthin were found between species or treatments. The daily PRI vs PAR (photosynthetically active radiation) relationships were determined using continuous measurements obtained at high frequency throughout the entire growing season, from early spring budburst to later autumn senescence, and were used to deconvolute the physiological PRI variability related to LUE variations due to phenological variability and related to temporal changes in the biochemical and structural canopy attributes. The PRI vs PAR relationship is used to show that the canopy-scale PRI measured at low radiation depends on the chlorophyll content of the canopy. The range of PRI variations at an intra-daily scale and the dynamics of the xanthophyll pool do not vary between days, which suggests that the PRI responds to a xanthophyll ratio. The PAR values at PRI saturation are mainly related to the canopy chlorophyll content during budburst and senescence and to the soil moisture content when the chlorophyll content is no longer a limiting factor. This parameter is significantly lower in the oak species that experience less stress from variations in soil moisture and is species dependant. These results provide new insights regarding the analysis and the meaning of PRI variability as a proxy for LUE at the canopy scale.

Topics: Chlorophyll; Fagus; Photochemical Processes; Photosynthesis; Pinus; Plant Leaves; Quercus; Seasons; Soil; Xanthophylls; Zeaxanthins

The antioxidant activity and individual bioactive compounds of lettuce, cultivated with 2.5-30% (v/v) of fresh or composted espresso spent coffee grounds, were assessed. A progressive enhancement of lettuce's antioxidant capacity, evaluated by radical scavenging effect and reducing power, was exhibited with the increment of fresh spent coffee amounts, while this pattern was not so clear with composted treatments. Total reducing capacity also improved, particularly for low spent coffee concentrations. Additionally, very significant positive correlations were observed for all carotenoids in plants from fresh spent coffee treatments, particularly for violaxanthin, evaluated by HPLC. Furthermore, chlorophyll a was a good discriminating factor between control group and all spent coffee treated samples, while vitamin E was not significantly affected. Espresso spent coffee grounds are a recognised and valuable source of bioactive compounds, proving herein, for the first time, to potentiate the antioxidant pool and quality of the vegetables produced.

Topics: Antioxidants; Carotenoids; Chlorophyll; Chlorophyll A; Chromatography, High Pressure Liquid; Coffee; Crops, Agricultural; Fertilizers; Lactuca; Lutein; Plant Extracts; Soil; Tocopherols; Xanthophylls

Violaxanthin-chlorophyll a binding protein (VCP) is the major light harvesting complex (LHC) of the Heterokonta Nannochloropsis gaditana. It binds chlorophyll a, violaxanthin and vaucheriaxanthin, the last in the form of 19' deca/octanoate esters. Photosynthetic apparatus of algae belonging to this group have been poorly characterized in the past, but they are now receiving an increasing interest also because of their possible biotechnological application in biofuel production. In this work, isolated VCP proteins have been studied by means of advanced EPR techniques in order to prove the presence of the photoprotective mechanism based on the triplet-triplet energy transfer (TTET), occurring between chlorophyll and carotenoid molecules. This process has been observed before in several light harvesting complexes belonging to various photosynthetic organisms. We used Optically Detected Magnetic Resonance (ODMR) to identify the triplet states populated by photo-excitation, and describe the optical properties of the chromophores carrying the triplet states. In parallel, time-resolved EPR (TR-EPR) and pulse EPR have been employed to get insight into the TTET mechanism and reveal the structural features of the pigment sites involved in photoprotection. The analysis of the spectroscopic data shows a strong similarity among VCP, FCP from diatoms and LHC-II from higher plants. Although these antenna proteins have differentiated sequences and bind different pigments, results suggest that in all members of the LHC superfamily there is a protein core with a conserved structural organization, represented by two central carotenoids surrounded by five chlorophyll a molecules, which plays a fundamental photoprotective role in Chl triplet quenching through carotenoid triplet formation.

Topics: Amino Acid Sequence; Carotenoids; Chlorophyll; Chlorophyll A; Chlorophyll Binding Proteins; Energy Transfer; Light-Harvesting Protein Complexes; Photosynthesis; Protein Conformation; Stramenopiles; Xanthophylls

It is proposed that xanthophylls, and carotenoids in general, may assist in energy transfer from the chlorophyll Soret band to the Q band. Ground-state (1Ag ) and excited-state (1Bu ) optimizations of violaxanthin (Vx) and zeaxanthin (Zx) are performed in an environment mimicking the light-harvesting complex II (LHCII), including the closest chlorophyll b molecule (Chl). Time-dependent density functional theory (TD-DFT, CAM-B3LYP functional) is used in combination with a semi-empirical description to obtain the excited-state geometries, supported by additional DFT/multireference configuration interaction calculations, with and without point charges representing LHCII. In the ground state, Vx and Zx show similar properties. At the 1Bu minimum, the energy of the Zx 1Bu state is below the Chl Q band, in contrast to Vx. Both Vx and Zx may act as acceptors of Soret-state energy; transfer to the Q band seems to be favored for Vx. These findings suggest that carotenoids may generally mediate Soret-to-Q energy flow in LHCII.

Topics: Carotenoids; Chlorophyll; Energy Transfer; Light-Harvesting Protein Complexes; Models, Molecular; Quantum Theory; Thermodynamics; Xanthophylls; Zeaxanthins

In highlight stress conditions, the mechanism of non-photochemical quenching (NPQ) of chlorophyll fluorescence is triggered at the chloroplast level. This process allows thermal quenching of the excessive excitation energy and it is strictly related to the efficiency of the xanthophyll cycle. Nowadays, the utilization of the nuclear magnetic resonance (NMR) spectroscopy provides a powerful complementary way for the identification and quantitative analysis of plant metabolites either in vivo or in tissue extracts. Seeing that the oxidative damage caused by light stress in plants and the consequent involvement of pigments are widely studied, NMR spectroscopy can be utilized to compare crude leaf extract at different levels of light stress, allowing an analysis of these compounds. In this paper, the identification of possible relationships between light stress and ¹H NMR signal variations is discussed. The analysis of the ¹H NMR (1D) spectra of two agronomic species (Spinacia oleracea and Beta vulgaris) exposed to different light intensities is presented. In particular, change in carotenoids and xanthophylls signals are analyzed.

Topics: Beta vulgaris; Carotenoids; Chlorophyll; Complex Mixtures; Crops, Agricultural; Light; Magnetic Resonance Spectroscopy; Plant Extracts; Plant Leaves; Spinacia oleracea; Stress, Physiological; Xanthophylls

Light-harvesting by the xanthophylls in the antenna of photosystem II (PSII) is a very efficient process (with 80% of the absorbed energy being transfer to chlorophyll). However, the efficiencies of the individual xanthophylls vary considerably, with violaxanthin in LHCII contributing very little to light-harvesting. To investigate the origin of the variation we used Time Dependent Density Functional Theory to model the Coulombic interactions between the xanthophyll 1(1)B(u)(+) states and the chlorophyll Soret band states in the LHCII and CP29 antenna complexes. The results show that the central L1 and L2 binding sites in both complexes favored close cofacial associations between the bound xanthophylls and chlorophyll a, implying efficient energy transfer, consistent with previously reported experimental evidence. Additionally, we found that the peripheral V1 binding site in LHCII did not favor close xanthophyll-chlorophyll associations, confirming observations that violaxanthin in LHCII is not an effective light-harvester. Finally, violaxanthin bound into the L2 site of the CP29 complex was found to be very strongly coupled to its neighboring chlorophylls.

Topics: Binding Sites; Chlorophyll; Energy Transfer; Light-Harvesting Protein Complexes; Photosystem II Protein Complex; Quantum Theory; Xanthophylls

It has long been suspected that photoprotective mechanisms in green algae are similar to those in seed plants. However, exceptions have recently surfaced among aquatic and marine green algae in several taxonomic classes. Green algae are highly diverse genetically, falling into 13 named classes, and they are diverse ecologically, with many lineages including members from freshwater, marine, and terrestrial habitats. Genetically similar species living in dramatically different environments are potentially a rich source of information about variations in photoprotective function. Using aquatic and desert-derived species from three classes of green algae, we examined the induction of photoprotection under high light, exploring the relationship between nonphotochemical quenching and the xanthophyll cycle. In liquid culture, behavior of aquatic Entransia fimbriata (Klebsormidiophyceae) generally matched patterns observed in seed plants. Nonphotochemical quenching was lowest after overnight dark adaptation, increased with light intensity, and the extent of nonphotochemical quenching correlated with the extent of deepoxidation of xanthophyll cycle pigments. In contrast, overnight dark adaptation did not minimize nonphotochemical quenching in the other species studied: desert Klebsormidium sp. (Klebsormidiophyceae), desert and aquatic Cylindrocystis sp. (Zygnematophyceae), and desert Stichococcus sp. (Trebouxiophyceae). Instead, exposure to low light reduced nonphotochemical quenching below dark-adapted levels. De-epoxidation of xanthophyll cycle pigments paralleled light-induced changes in nonphotochemical quenching for species within Klebsormidiophyceae and Trebouxiophyceae, but not Zygnematophyceae. Inhibition of violaxanthin-zeaxanthin conversion by dithiothreitol reduced high-light-associated nonphotochemical quenching in all species (Zygnematophyceae the least), indicating that zeaxanthin can contribute to photoprotection as in seed plants but to different extents depending on taxon or lineage.

Topics: Adaptation, Physiological; Aquatic Organisms; Chlorophyll; Chlorophyta; Desert Climate; Fluorescence; Light; Molecular Sequence Data; Phylogeny; Xanthophylls; Zeaxanthins

Nonphotochemical quenching (NPQ) dissipates excess energy to protect the photosynthetic apparatus from excess light. The moss Physcomitrella patens exhibits strong NPQ by both algal-type light-harvesting complex stress-related (LHCSR)-dependent and plant-type S subunit of Photosystem II (PSBS)-dependent mechanisms. In this work, we studied the dependence of NPQ reactions on zeaxanthin, which is synthesized under light stress by violaxanthin deepoxidase (VDE) from preexisting violaxanthin. We produced vde knockout (KO) plants and showed they underwent a dramatic reduction in thermal dissipation ability and enhanced photoinhibition in excess light conditions. Multiple mutants (vde lhcsr KO and vde psbs KO) showed that zeaxanthin had a major influence on LHCSR-dependent NPQ, in contrast with previous reports in Chlamydomonas reinhardtii. The PSBS-dependent component of quenching was less dependent on zeaxanthin, despite the near-complete violaxanthin to zeaxanthin exchange in LHC proteins. Consistent with this, we provide biochemical evidence that native LHCSR protein binds zeaxanthin upon excess light stress. These findings suggest that zeaxanthin played an important role in the adaptation of modern plants to the enhanced levels of oxygen and excess light intensity of land environments.

Topics: Adaptation, Physiological; Biosynthetic Pathways; Bryopsida; Chlorophyll; Gene Knockout Techniques; Light; Light-Harvesting Protein Complexes; Photosystem II Protein Complex; Thylakoids; Xanthophylls; Zeaxanthins

The xanthophyll cycle (Xc), which involves violaxanthin de-epoxidase (VDE) and the zeaxanthin epoxidase (ZEP), is one of the most rapid and efficient responses of plant and algae to high irradiance. High light intensity can activate VDE to convert violaxanthin (Vx) to zeaxanthin (Zx) via antheraxanthin (Ax). However, it remains unclear whether VDE remains active under low light or dark conditions when there is no significant accumulation of Ax and Zx, and if so, how the ΔpH required for activation of VDE is built. In this study, we used salicylaldoxime (SA) to inhibit ZEP activity in the intertidal macro-algae Ulva sp. (Ulvales, Chlorophyta) and then characterized VDE under low light and dark conditions with various metabolic inhibitors. With inhibition of ZEP by SA, VDE remained active under low light and dark conditions, as indicated by large accumulations of Ax and Zx at the expense of Vx. When PSII-mediated linear electron transport systems were completely inhibited by SA and DCMU, alternative electron transport systems (i.e., cyclic electron transport and chlororespiration) could maintain VDE activity. Furthermore, accumulations of Ax and Zx decreased significantly when SA, DCMU, or DBMIB together with an inhibitor of chlororespiration (i.e., propyl gallate (PG)) were applied to Ulva sp. This result suggests that chlororespiration not only participates in the build-up of the necessary ΔpH, but that it also possibly influences VDE activity indirectly by diminishing the oxygen level in the chloroplast.

Topics: Chlorophyll; Chloroplasts; Electron Transport; Light; Oxidation-Reduction; Oxidoreductases; Photoperiod; Protons; Reactive Oxygen Species; Ulva; Xanthophylls; Zeaxanthins

Under excess light, the efficient PSII light-harvesting antenna is switched into a photoprotected state in which potentially harmful absorbed energy is thermally dissipated. Changes occur rapidly and reversibly, enhanced by de-epoxidation of violaxanthin (V) to zeaxanthin (Z). This process is usually measured as non-photochemical quenching (NPQ) of chlorophyll (Chl) fluorescence. Using instrumentation for instantaneous leaf freezing, NPQ, spectral reflectance, and interconversions within the xanthophyll cycle with time resolution of seconds were recorded from Quercus coccifera leaves during low light (LL) to high light (HL) transitions, followed by relaxation at LL. During the first 30 s of both the LL to HL and HL to LL transitions, no activity of the xanthophyll cycle was detected, whereas 70-75% of the NPQ was formed and relaxed, respectively, by that time, the latter being traits of a rapidly reversible photoprotective energy dissipation. Three different Z pools were identified, which play different roles in energy dissipation and photoprotection. In conclusion, ΔpH was crucial to NPQ formation and relaxation in Q. coccifera during light transitions. Only a minor fraction of Z was associated to quenching, whereas the largest Z pool was not related to thermal dissipation. The latter is proposed to participate in photoprotection acting as antioxidant.

Topics: Antioxidants; Chlorophyll; Energy Transfer; Hydrogen-Ion Concentration; Light; Oxidation-Reduction; Photochemical Processes; Photosystem II Protein Complex; Plant Leaves; Quercus; Time Factors; Xanthophylls; Zeaxanthins

Chromerida are photoautotrophic alveolates so far only isolated from corals in Australia. It has been shown that these secondary plastid-containing algae are closely related to apicomplexan parasites and share various morphological and molecular characters with both Apicomplexa and Dinophyta. So far, the only known representative of the phylum was Chromera velia. Here we provide a formal description of another chromerid, Vitrella brassicaformis gen. et sp. nov., complemented with a detailed study on its ultrastructure, allowing insight into its life cycle. The novel alga differs significantly from the related chromerid C. velia in life cycle, morphology as well as the plastid genome. Analysis of photosynthetic pigments on the other hand demonstrate that both chromerids lack chlorophyll c, the hallmark of phototrophic chromalveolates. Based on the relatively high divergence between C. velia and V. brassicaformis, we propose their classification into distinct families Chromeraceae and Vitrellaceae. Moreover, we predict a hidden and unexplored diversity of the chromerid algae.

Topics: Alveolata; beta Carotene; Cell Membrane; Cell Wall; Chlorophyll; Chlorophyll A; Coral Reefs; Flagella; Genome, Plastid; Microscopy, Electron; Phylogeny; Pigments, Biological; Plastids; Spores, Protozoan; Xanthophylls

Collecting energy for photosystem II is facilitated by several pigments, xanthophylls and chlorophylls, embedded in the light harvesting complex II (LHCII). One xanthophyll, violaxanthin (Vio), is loosely bound at a site close to a chlorophyll b (Chl). No final answer has yet been found for the role of this specific xanthophyll. We study the electronic structure of Vio in the presence of Chl and under the influence of the LHCII environment, represented by a point charge field (PCF). We compare the capability of the long range corrected density functional theory (DFT) functional CAM-B3LYP to B3LYP for the modeling of the UV/vis spectrum of the Vio+Chl pair. CAM-B3LYP was reported to allow for a very realistic reproduction of bond length alternation of linear polyenes, which has considerable impact on the carotenoid structure and spectrum. To account for the influence of the LHCII environment, the chromophore geometries are optimized using an ONIOM(DFT/6-31G(d):PM6) scheme. Our calculations show that the energies of the locally excited states are almost unaffected by the presence of the partner chromophore or the PCF. There are, however, indications for excitonic coupling of the Chl Soret band and Vio. We propose that Vio may accept energy from blue-light excited Chl.

Topics: Chlorophyll; Electrons; Light-Harvesting Protein Complexes; Models, Molecular; Pisum sativum; Protein Conformation; Quantum Theory; Solutions; Xanthophylls

In photosynthetic organisms, feedback dissipation of excess absorbed light energy balances harvesting of light with metabolic energy consumption. This mechanism prevents photodamage caused by reactive oxygen species produced by the reaction of chlorophyll (Chl) triplet states with O₂. Plants have been found to perform the heat dissipation in specific proteins, binding Chls and carotenoids (Cars), that belong to the Lhc family, while triggering of the process is performed by the PsbS subunit, needed for lumenal pH detection. PsbS is not found in algae, suggesting important differences in energy-dependent quenching (qE) machinery. Consistent with this suggestion, a different Lhc-like gene product, called LhcSR3 (formerly known as LI818) has been found to be essential for qE in Chlamydomonas reinhardtii. In this work, we report the production of two recombinant LhcSR isoforms from C. reinhardtii and their biochemical and spectroscopic characterization. We found the following: (i) LhcSR isoforms are Chl a/b- and xanthophyll-binding proteins, contrary to higher plant PsbS; (ii) the LhcSR3 isoform, accumulating in high light, is a strong quencher of Chl excited states, exhibiting a very fast fluorescence decay, with lifetimes below 100 ps, capable of dissipating excitation energy from neighbor antenna proteins; (iii) the LhcSR3 isoform is highly active in the transient formation of Car radical cation, a species proposed to act as a quencher in the heat dissipation process. Remarkably, the radical cation signal is detected at wavelengths corresponding to the Car lutein, rather than to zeaxanthin, implying that the latter, predominant in plants, is not essential; (iv) LhcSR3 is responsive to low pH, the trigger of non-photochemical quenching, since it binds the non-photochemical quenching inhibitor dicyclohexylcarbodiimide, and increases its energy dissipation properties upon acidification. This is the first report of an isolated Lhc protein constitutively active in energy dissipation in its purified form, opening the way to detailed molecular analysis. Owing to its protonatable residues and constitutive excitation energy dissipation, this protein appears to merge both pH-sensing and energy-quenching functions, accomplished respectively by PsbS and monomeric Lhcb proteins in plants.

Topics: Absorption; Algal Proteins; Amino Acid Sequence; Chlamydomonas reinhardtii; Chlorophyll; Feedback, Physiological; Fluorescence; Light; Light-Harvesting Protein Complexes; Lutein; Molecular Sequence Data; Protein Binding; Protein Isoforms; Protein Processing, Post-Translational; Protein Refolding; Protein Structure, Tertiary; Recombinant Proteins; Sequence Alignment; Thermodynamics; Thylakoids; Xanthophylls

Although the wheat hybrids have often shown higher grain yields, the physiological basis of the higher yields remains unknown. Previous studies suggest that tolerance to photoinhibition in the hybrid may be one of the physiological bases (Yang et al., 2006, Plant Sci 171:389-97). The objective of this study was to further investigate the possible mechanism responsible for tolerance to photoinhibition in the hybrid. Photosystem II (PSII) photochemistry, the xanthophyll cycle, and antioxidative defense system were compared between the hybrid and its parents subjected to high light stress (1500μmolm(-2)s(-1)). The analyses of oxygen-evolving activity, chlorophyll fluorescence, and protein blotting demonstrated that the higher tolerance in the hybrid than in its parents was associated with its higher tolerance of PSII to photoinhibition. High light induced an increase in non-photochemical quenching, and this increase was greater in the hybrid than in its parents. There were no differences in the pool size of the xanthophyll cycle between the hybrid and its parents. The content of violaxanthin decreased significantly, whereas the content of zeaxanthin+antherxanthin increased considerably during high light treatments. However, the decrease in violaxanthin content and the increase in zeaxanthin+antherxanthin content were greater in the hybrid than in its parents. High light resulted in a significant accumulation of H(2)O(2), O(2)(-) and catalytic Fe, and this accumulation was less in the hybrid than in its parents. High light induced a significant increase in the activities of superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase, dehydroascorbate reductase, and monodehydroascorbate reductase, and these increases were greater in the hybrid than its parents. These results suggest that the higher tolerance to photoinhibition in the hybrid may be associated with its higher capacity for antioxidative defense metabolism and the xanthophyll cycle.

Topics: Adaptation, Physiological; Antioxidants; Chimera; Chlorophyll; Light; Oxygen; Photochemistry; Photosystem II Protein Complex; Plant Leaves; Stress, Physiological; Time Factors; Triticum; Xanthophylls; Zeaxanthins

Carotenoids are naturally occurring pigments that absorb light in the spectral region in which the sun irradiates maximally. These molecules transfer this energy to chlorophylls, initiating the primary photochemical events of photosynthesis. Carotenoids also regulate the flow of energy within the photosynthetic apparatus and protect it from photoinduced damage caused by excess light absorption. To carry out these functions in nature, carotenoids are bound in discrete pigment-protein complexes in the proximity of chlorophylls. A few three-dimensional structures of these carotenoid complexes have been determined by X-ray crystallography. Thus, the stage is set for attempting to correlate the structural information with the spectroscopic properties of carotenoids to understand the molecular mechanism(s) of their function in photosynthetic systems. In this Account, we summarize current spectroscopic data describing the excited state energies and ultrafast dynamics of purified carotenoids in solution and bound in light-harvesting complexes from purple bacteria, marine algae, and green plants. Many of these complexes can be modified using mutagenesis or pigment exchange which facilitates the elucidation of correlations between structure and function. We describe the structural and electronic factors controlling the function of carotenoids as energy donors. We also discuss unresolved issues related to the nature of spectroscopically dark excited states, which could play a role in light harvesting. To illustrate the interplay between structural determinations and spectroscopic investigations that exemplifies work in the field, we describe the spectroscopic properties of four light-harvesting complexes whose structures have been determined to atomic resolution. The first, the LH2 complex from the purple bacterium Rhodopseudomonas acidophila, contains the carotenoid rhodopin glucoside. The second is the LHCII trimeric complex from higher plants which uses the carotenoids lutein, neoxanthin, and violaxanthin to transfer energy to chlorophyll. The third, the peridinin-chlorophyll-protein (PCP) from the dinoflagellate Amphidinium carterae, is the only known complex in which the bound carotenoid (peridinin) pigments outnumber the chlorophylls. The last is xanthorhodopsin from the eubacterium Salinibacter ruber. This complex contains the carotenoid salinixanthin, which transfers energy to a retinal chromophore. The carotenoids in these pigment-protein complexes transfer

Topics: Carotenoids; Chlorophyll; Dinoflagellida; Energy Transfer; Eukaryota; Glucosides; Glycosides; Light; Light-Harvesting Protein Complexes; Lutein; Photosynthesis; Rhodopseudomonas; Thylakoids; Xanthophylls

Leaves are the main photosynthetically active tissues in most plants. However, stems and fruits are also important for the overall carbon balance of the plant because of their contribution to fixation of the CO(2) released by respiration. Photosynthesis could not be possible without a complete set of photoprotection mechanisms, which include the ubiquitous violaxanthin (V) cycle and the taxonomically restricted lutein epoxide (Lx) cycle. In this work, we characterise carotenoid stoichiometry in photosynthetic stems and fruits of avocado in comparison with that of leaves and specifically whether Lx is present in these tissues and also whether it is involved in a light-driven cycle. Avocado was selected as model species to study whether both cycles were functional in non-foliar photosynthetic structures (stems and fruits). An unusual pigment composition was observed in avocado fruit, with a high content of cis-V and cis-Lx, suggesting a different photosynthetic function. In stems, both xanthophylls de-epoxidated upon illumination, but only V recovered in the dark, indicating the existence of a possible 'truncated' Lx cycle. Lx in fruits was de-epoxidated only when its pool was higher than a threshold of 30 mmol mol(-1) chlorophyll, indicating a high non-photoconvertible pool of Lx. We conclude that, at least in stems, the dynamic regulation of photosynthetic activity could also depend on the Lx cycle.

Topics: Chlorophyll; Fruit; Light; Lutein; Persea; Photosynthesis; Plant Stems; Xanthophylls

Lhcb6 (CP24) is a monomeric antenna protein of photosystem II, which has been shown to play special roles in photoprotective mechanisms, such as the Non-Photochemical Quenching and reorganization of grana membranes in excess light conditions. In this work we analyzed Lhcb6 in vivo and in vitro: we show this protein, upon activation of the xanthophyll cycle, accumulates zeaxanthin into inner binding sites faster and to a larger extent than any other pigment-protein complex. By comparative analysis of Lhcb6 complexes violaxanthin or zeaxanthin binding, we demonstrate that zeaxanthin not only down-regulates chlorophyll singlet excited states, but also increases the efficiency of chlorophyll triplet quenching, with consequent reduction of singlet oxygen production and significant enhancement of photo-stability. On these bases we propose that Lhcb6, the most recent addition to the Lhcb protein family which evolved concomitantly to the adaptation of photosynthesis to land environment, has a crucial role in zeaxanthin-dependent photoprotection.

Topics: Arabidopsis; Arabidopsis Proteins; Chlorophyll; Chlorophyll Binding Proteins; Epoxy Compounds; Kinetics; Light; Light-Harvesting Protein Complexes; Photobleaching; Photosystem II Protein Complex; Plant Leaves; Protein Binding; Singlet Oxygen; Spectrum Analysis; Thylakoids; Xanthophylls; Zeaxanthins

The photoprotective role of the universal violaxanthin cycle that interconverts violaxanthin (V), antheraxanthin (A), and zeaxanthin (Z) is well established, but functions of the analogous conversions of lutein-5,6-epoxide (Lx) and lutein (L) in the selectively occurring Lx cycle are still unclear. We investigated carotenoid pools in Lx-rich leaves of avocado (Persea americana) during sun or shade acclimation at different developmental stages. During sun exposure of mature shade leaves, an unusual decrease in L preceded the deepoxidation of Lx to L and of V to A+Z. In addition to deepoxidation, de novo synthesis increased the L and A+Z pools. Epoxidation of L was exceptionally slow, requiring about 40 d in the shade to restore the Lx pool, and residual A+Z usually persisted overnight. In young shade leaves, the Lx cycle was reversed initially, with Lx accumulating in the sun and declining in the shade. De novo synthesis of xanthophylls did not affect alpha- and beta-carotene pools on the first day, but during long-term acclimation alpha-carotene pools changed noticeably. Nonetheless, the total change in alpha- and beta-branch carotenoid pools was equal. We discuss the implications for regulation of metabolic flux through the alpha- and beta-branches of carotenoid biosynthesis and potential roles for L in photoprotection and Lx in energy transfer to photosystem II and explore physiological roles of both xanthophyll cycles as determinants of photosystem II efficiency.

Topics: Acclimatization; Chlorophyll; Kinetics; Lutein; Persea; Plant Leaves; Sunlight; Xanthophylls; Zeaxanthins

Nonphotochemical quenching (NPQ) is a fundamental mechanism in photosynthesis by which plants protect themselves against excess excitation energy and which is thus of crucial importance for plant survival and fitness. Recently, carotenoid radical cation (Car(*+)) formation has been discovered to be a key step in the feedback deexcitation quenching component (qE) of NPQ, whose molecular mechanism and location remains elusive. A recent model for qE suggests that the replacement of violaxanthin (Vio) by zeaxanthin (Zea) in photosynthetic pigment binding pockets can in principle result in qE via the so-called "gear-shift" or electron transfer quenching mechanisms. We performed pump-probe measurements on individual antenna complexes of photosystem II (CP24, CP26 and CP29) upon excitation of the chlorophylls (Chl) into their first excited Q(y) state at 660 nm when either Vio or Zea was bound to those complexes. The Chl lifetime was then probed by measuring the decay kinetics of the Chl excited state absorption (ESA) at 900 nm. The charge-transfer quenching mechanism, which is characterized by a spectral signature of the transiently formed Zea radical cation (Zea(*+)) in the near-IR, has also been addressed, both in solution and in light-harvesting complexes of photosystem II (LHC-II). Applying resonant two-photon two-color ionization (R2P2CI) spectroscopy we show here the formation of beta-Car(*+) in solution, which occurs on a femtosecond time-scale by direct electron transfer to the solvent. The beta-Car(*+) maxima strongly depend on the solvent polarity. Moreover, our two-color two-photon spectroscopy on CP29 reveals the spectral position of Zea(*+) in the near-IR region at 980 nm.

Topics: Arabidopsis Proteins; Cations, Monovalent; Chlorophyll; Chlorophyll Binding Proteins; Electron Transport; Free Radicals; Light-Harvesting Protein Complexes; Models, Biological; Photochemical Processes; Photosystem II Protein Complex; Plant Proteins; Recombinant Proteins; Spectrophotometry; Xanthophylls; Zeaxanthins

Zeaxanthin epoxidase (ZE, E.C. 1.14.13.90), an enzyme belonging to the lipocalin superfamily, catalyses the conversion of zeaxanthin to antheraxanthin and violaxanthin. These reactions are part of the xanthophyll biosynthetic pathway and the xanthophyll cycle. The role of carotenoids in the dissipation of excessive light energy has been widely studied using mutants with a disabled carotenoid biosynthetic pathway. In this paper, the transgenic line MaZEP7 with partially disabled ZE activity is described and compared with wild-type plants and npq2 mutant lacking active ZE. We examined the presence and the abundance of aba1 transcripts, measured pigment composition, xanthophyll cycle functioning and chlorophyll fluorescence in all three lines. The MaZEP7 line contains additional copies of the aba1 gene introduced by agroinfiltration, but no enhanced aba1 transcript level was observed. In addition, ZE activity in MaZEP7 was impaired, resulting in an altered xanthophyll profile. In dark-adapted plants, violaxanthin and neoxanthin levels were lower than in wild-type plants, whereas antheraxanthin and zeaxanthin levels were considerably higher. The presence of lutein epoxide was also observed. Violaxanthin levels changed only minimally during light exposition, whereas antheraxanthin was converted to zeaxanthin and there was no epoxidation during the course of the experiment indicating disturbed xanthophyll cycle functioning. The amounts of carotenoids and chlorophylls on a dry weight basis and chl a/chl b ratio were similar in all lines. The presence of epoxidated pigments in MaZEP7 plants indicates that epoxidation occurs, but it is likely very slow. Chlorophyll fluorescence measurements showed that the dependence of electron transport rates on light intensity for the MaZEP7 line resembled the npq2 mutant. Kinetic measurements showed that the MaZEP7 line exhibited very rapid induction and a high steady-state value of non-photochemical quenching.

Topics: Arabidopsis; Carotenoids; Chlorophyll; Gene Expression Regulation, Plant; Kinetics; Light; Oxidoreductases; Photochemistry; Plants, Genetically Modified; Xanthophylls; Zeaxanthins

The effects of high Zn concentration were investigated in sugar beet (Beta vulgaris L.) plants grown in a controlled environment in hydroponics. High concentrations of Zn sulphate in the nutrient solution (50, 100 and 300 microm) decreased root and shoot fresh and dry mass, and increased root/shoot ratios, when compared to control conditions (1.2 microm Zn). Plants grown with excess Zn had inward-rolled leaf edges and a damaged and brownish root system, with short lateral roots. High Zn decreased N, Mg, K and Mn concentrations in all plant parts, whereas P and Ca concentrations increased, but only in shoots. Leaves of plants treated with 50 and 100 microm Zn developed symptoms of Fe deficiency, including decreases in Fe, chlorophyll and carotenoid concentrations, increases in carotenoid/chlorophyll and chlorophyll a/b ratios and de-epoxidation of violaxanthin cycle pigments. Plants grown with 300 microm Zn had decreased photosystem II efficiency and further growth decreases but did not have leaf Fe deficiency symptoms. Leaf Zn concentrations of plants grown with excess Zn were high but fairly constant (230-260 microg.g(-1) dry weight), whereas total Zn uptake per plant decreased markedly with high Zn supply. These data indicate that sugar beet could be a good model to investigate Zn homeostasis mechanisms in plants, but is not an efficient species for Zn phytoremediation.

Topics: Beta vulgaris; Biological Transport; Carbon Dioxide; Chlorophyll; Chlorophyll A; FMN Reductase; Hydroponics; Minerals; Nitrogen; Oxygen; Photosystem II Protein Complex; Plant Structures; Trace Elements; Xanthophylls; Zinc; Zinc Sulfate

In the Crassulacean acid metabolism (CAM) plants Clusia alata Triana and Planch., decarboxylation of citrate during phase III of CAM took place later than malate decarboxylation. The interdependence of these two CO(2) and NADPH sources is discussed. High light accelerated malate decarboxylation during the day and lowered citrate levels. Strong light stress also activated mechanisms that can protect the plant against oxidative stress. Upon transfer from low light (200micromol m(-2)s(-1)) to high light (650-740micromol m(-2)s(-1)), after 2 days, there was a transient increase of non-photochemical quenching (NPQ) of fluorescence of chlorophyll a of photosystem II. This indicated acute photoinhibition, which declined again after 7 days of exposure. Conversely, after 1 week exposure to high light, the mechanisms of interconversion of violaxanthin (V), antheraxanthin (A), zeaxanthin (Z) (epoxydation/de-epoxydation) were activated. This was accompanied by an increase in pigment levels at dawn and dusk.

Topics: Adaptation, Physiological; Chlorophyll; Chlorophyll A; Citric Acid; Clusia; Decarboxylation; Fluorescence; Light; Malates; Photosynthesis; Photosystem II Protein Complex; Stress, Physiological; Xanthophylls; Zeaxanthins

The interactive effects of root-zone salinity and sunlight on leaf biochemistry, with special emphasis on antioxidant defences, were analysed in Olea europaea L. cv. Allora, during the summer period. Plants were grown outside under 15% (shade plants) or 100% sunlight (sun plants) and supplied with 0 or 125 mM NaCl. The following measurements were performed: (1) the contribution of ions and soluble carbohydrates to osmotic potentials; (2) the photosystem II (PSII) photochemistry and the photosynthetic pigment concentration; (3) the concentration and the tissue-specific distribution of leaf flavonoids; (4) the activity of antioxidant enzymes; and (5) the leaf oxidative damage. The concentrations of Na(+) and Cl(-) were significantly greater in sun than in shade leaves, as also observed for the concentration of the 'antioxidant' sugar-alcohol mannitol. The de-epoxidation state of violaxanthin-cycle pigments increased in response to salinity stress in sun leaves. This finding agrees with a greater maximal PSII photochemistry (F(v)/F(m)) at midday, detected in salt-treated than in control plants, growing in full sunshine. By contrast, salt-treated plants in the shade suffered from midday depression in F(v)/F(m) to a greater degree than that observed in control plants. The high concentration of violaxanthin-cycle pigments in sun leaves suggests that zeaxanthin may protect the chloroplast from photo-oxidative damage, rather than dissipating excess excitation energy via non-photochemical quenching mechanisms. Dihydroxy B-ring-substituted flavonoid glycosides accumulate greatly in the mesophyll, not only in the epidermal cells, in response to high sunlight. The activity of antioxidant enzymes varied little because of sunlight irradiance, but declined sharply in response to high salinity in shade leaves. Interestingly, control and particularly salt-treated plants in the shade underwent greater oxidative damage than their sunny counterparts. These findings, which conform to the evolution of O. europaea in sunny environments, suggest that under partial shading, the antioxidant defence system may be ineffective to counter salt-induced oxidative damage.

Topics: Antioxidants; Chlorophyll; Electron Spin Resonance Spectroscopy; Flavonoids; Free Radicals; Lipid Peroxidation; Olea; Oxidation-Reduction; Oxidative Stress; Phenols; Photosystem II Protein Complex; Plant Leaves; Plant Roots; Polyphenols; Seasons; Sodium Chloride; Sunlight; Water; Xanthophylls

The response of Norway spruce saplings (Picea abies [L.] Karst.) was monitored continuously during short-term exposure (10 days) to high irradiance (HI; 1000micromol m(-2)s(-1)). Compared with plants acclimated to low irradiance (100micromol m(-2)s(-1)), plants after HI exposure were characterized by a significantly reduced CO(2) assimilation rate throughout the light response curve. Pigment contents varied only slightly during HI exposure, but a rapid and strong response was observed in xanthophyll cycle activity, particularly within the first 3 days of the HI treatment. Both violaxanthin convertibility under HI and the amount of zeaxanthin pool sustained in darkness increased markedly under HI conditions. These changes were accompanied by an enhanced non-radiative dissipation of absorbed light energy (NRD) and the acceleration of induction of both NRD and de-epoxidation of the xanthophyll cycle pigments. We found a strong negative linear correlation between the amount of sustained de-epoxidized xanthophylls and the photosystem II (PSII) photochemical efficiency (F(V)/F(M)), indicating photoprotective down-regulation of the PSII function. Recovery of F(V)/F(M) at the end of the HI treatment revealed that Norway spruce was able to cope with a 10-fold elevated irradiance due particularly to an efficient NRD within the PSII antenna that was associated with enhanced violaxanthin convertibility and a light-induced accumulation of zeaxanthin that persisted in darkness.

Topics: Absorption; Carbon Dioxide; Chlorophyll; Chlorophyll A; Electron Transport; Epoxy Compounds; Fluorescence; Light; Lutein; Norway; Photosynthesis; Photosystem II Protein Complex; Picea; Time Factors; Xanthophylls; Zeaxanthins

Recently, a mechanism for the energy-dependent component (qE) of non-photochemical quenching (NPQ), the fundamental photo-protection mechanism in green plants, has been suggested. Replacement of violaxanthin by zeaxanthin in the binding pocket of the major light harvesting complex LHC-II may be sufficient to invoke efficient chlorophyll fluorescence quenching. Our quantum chemical calculations, however, show that the excited state energies of violaxanthin and zeaxanthin are practically identical when their geometry is constrained to the naturally observed structure of violaxanthin in LHC-II. Therefore, since violaxanthin does not quench LHC-II, zeaxanthin should not either. This theoretical finding is nicely in agreement with experimental results obtained by femtosecond spectroscopy on LHC-II complexes containing violaxanthin or zeaxanthin.

Topics: Chlorophyll; Computer Simulation; Fluorescence; Light-Harvesting Protein Complexes; Models, Molecular; Molecular Structure; Protein Structure, Secondary; Xanthophylls; Zeaxanthins

In the present study, xanthophyll composition of eight parasitic Cuscuta species under different light conditions was investigated. Neoxanthin was not detected in four of the eight species examined, while in others it occurred at the level of several percent of total xanthophylls. In C. gronovii and C. lupuliformis it was additionally found that the neoxanthin content was considerably stimulated by strong light. In dark-adapted plants, lutein epoxide level amounted to 10-22% of total xanthophylls in only three species, the highest being for C. lupuliformis, while in others it was below 3%, indicating that the lutein epoxide cycle is limited to only certain Cuscuta species. The obtained data also indicate that the presence of the lutein epoxide cycle and of neoxanthin is independent and variable among the Cuscuta species. The xanthophyll cycle carotenoids violaxanthin, antheraxanthin and zeaxanthin were identified in all the examined species and occurred at the level found in other higher plants. The xanthophyll and lutein epoxide cycle pigments showed typical response to high light stress. The obtained results also suggest that the ability of higher plants to synthesize lutein epoxide probably does not depend on the substrate specificity of zeaxanthin epoxidase but on the availability of lutein for the enzyme.

Topics: Animals; Carotenoids; Chlorophyll; Chromatography, High Pressure Liquid; Cuscuta; Epoxy Compounds; Light; Lutein; Pigmentation; Substrate Specificity; Time Factors; Xanthophylls

Energy-dependent quenching of excess absorbed light energy (qE) is a vital mechanism for regulating photosynthetic light harvesting in higher plants. All of the physiological characteristics of qE have been positively correlated with charge transfer between coupled chlorophyll and zeaxanthin molecules in the light-harvesting antenna of photosystem II (PSII). We found evidence for charge-transfer quenching in all three of the individual minor antenna complexes of PSII (CP29, CP26, and CP24), and we conclude that charge-transfer quenching in CP29 involves a delocalized state of an excitonically coupled chlorophyll dimer. We propose that reversible conformational changes in CP29 can "tune" the electronic coupling between the chlorophylls in this dimer, thereby modulating the energy of the chlorophyll-zeaxanthin charge-transfer state and switching on and off the charge-transfer quenching during qE.

Topics: Arabidopsis Proteins; Chlorophyll; Chlorophyll A; Chlorophyll Binding Proteins; Chloroplast Proteins; Electron Transport; Electrophysiology; Light; Light-Harvesting Protein Complexes; Lutein; Models, Molecular; Photosystem II Protein Complex; Protein Conformation; Recombinant Proteins; Ribonucleoproteins; Structure-Activity Relationship; Xanthophylls; Zeaxanthins

The aba4-1 mutant completely lacks neoxanthin but retains all other xanthophyll species. The missing neoxanthin in light-harvesting complex (Lhc) proteins is compensated for by higher levels of violaxanthin, albeit with lower capacity for photoprotection compared with proteins with wild-type levels of neoxanthin. Detached leaves of aba4-1 were more sensitive to oxidative stress than the wild type when exposed to high light and incubated in a solution of photosensitizer agents. Both treatments caused more rapid pigment bleaching and lipid oxidation in aba4-1 than wild-type plants, suggesting that neoxanthin acts as an antioxidant within the photosystem II (PSII) supercomplex in thylakoids. While neoxanthin-depleted Lhc proteins and leaves had similar sensitivity as the wild type to hydrogen peroxide and singlet oxygen, they were more sensitive to superoxide anions. aba4-1 intact plants were not more sensitive than the wild type to high-light stress, indicating the existence of compensatory mechanisms of photoprotection involving the accumulation of zeaxanthin. However, the aba4-1 npq1 double mutant, lacking zeaxanthin and neoxanthin, underwent stronger PSII photoinhibition and more extensive oxidation of pigments than the npq1 mutant, which still contains neoxanthin. We conclude that neoxanthin preserves PSII from photoinactivation and protects membrane lipids from photooxidation by reactive oxygen species. Neoxanthin appears particularly active against superoxide anions produced by the Mehler's reaction, whose rate is known to be enhanced in abiotic stress conditions.

Topics: Arabidopsis; Arabidopsis Proteins; Binding Sites; Centrifugation, Density Gradient; Chlorophyll; Energy Metabolism; Free Radical Scavengers; Light; Light-Harvesting Protein Complexes; Models, Molecular; Molecular Sequence Data; Mutation; Oxidants; Oxidative Stress; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Plant Leaves; Protein Binding; Reactive Oxygen Species; Temperature; Xanthophylls

Fluorescence lifetime-resolved images of chlorophyll fluorescence were acquired at the maximum P-level and during the slower transient (up to 250 s, including P-S-M-T) in the green photosynthetic alga Chlamydomonas reinhardtii. At the P-level, wild type and the violaxanthin-accumulating mutant npq1 show similar fluorescence intensity and fluorescence lifetime-resolved images. The zeaxanthin-accumulating mutant npq2 displays reduced fluorescence intensity at the P-level (about 25-35% less) and corresponding lifetime-resolved frequency domain phase and modulation values compared to wild type/npq1. A two-component analysis of possible lifetime compositions shows that the reduction of the fluorescence intensity can be interpreted as an increase in the fraction of a short lifetime component. This supports the important photoprotection function of zeaxanthin in photosynthetic samples, and is consistent with the notion of a 'dimmer switch'. Similar, but quantitatively different, behaviour was observed in the intensity and fluorescence lifetime-resolved imaging measurements for cells that were treated with the electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethyl urea, the efficient PSI electron acceptor methyl viologen and the protonophore nigericin and. Lower fluorescence intensities and lifetimes were observed for all npq2 mutant samples at the P-level and during the slow fluorescence transient, compared to wild type and the npq1 mutant. The fluorescence lifetime-resolved measurements during the slow fluorescence changes after the P level up to 250 s for the wild type and the two mutants, in the presence and absence of the above inhibitors, were analyzed with a graphical procedure (polar plots) to determine lifetime compositions. At higher illumination intensity, wild type and npq1 cells show a rise in fluorescence intensity and corresponding rise in the species concentration of the slow lifetime component after the initial decrease following the P level. This reversal is absent in the npq2 mutant, and for all samples in the presence of the inhibitors. Lifetime heterogeneities were observed in experiments averaged over multiple cells as well as within single cells, and these were followed over time. Cells in the resting state (induced by several hours of darkness), instead of the normal swimming state, show shortened lifetimes. The above results are discussed in terms of a superposition of effects on electron transfer and protonation rates, on the s

Topics: Animals; Chlamydomonas reinhardtii; Chlorophyll; Diuron; Energy Metabolism; Microscopy, Fluorescence; Models, Biological; Mutation; Nigericin; Paraquat; Photochemistry; Photosynthesis; Xanthophylls

In the present study we have examined the effects of grana stacking on the rate of violaxanthin (Vx) de-epoxidation and the extent of non-photochemical quenching of chlorophyll a fluorescence (NPQ) in isolated thylakoid membranes of spinach. Our results show that partial and complete unstacking of thylakoids in reaction media devoid of sorbitol and MgCl(2) did not significantly affect the efficiency of Vx de-epoxidation. Under high light (HL) illumination we found slightly higher values of Vx conversion in stacked membranes, whereas in thylakoids incubated at pH 5.2 in the dark, representing the pH-optimum of Vx de-epoxidase, de-epoxidation was slightly increased in the unstacked membranes. Partial and complete unstacking of grana membranes, however, had a dramatic effect on the HL-induced NPQ. High NPQ values could only be achieved in stacked thylakoid membranes in the presence of MgCl(2) and sorbitol. In unstacked membranes NPQ was drastically decreased. The effects of grana stacking on the xanthophyll cycle-dependent component of NPQ were even more pronounced, and complete unstacking of thylakoid membranes led to a total loss of this quenching component. Our data imply that grana stacking in the thylakoid membranes of higher plants is of high importance for the process of overall NPQ. For the xanthophyll cycle-dependent component of NPQ it may even be essential. Possible effects of grana stacking on the mechanism of zeaxanthin-dependent quenching are discussed.

Topics: Chlorophyll; Chlorophyll A; Chromatography, High Pressure Liquid; Circular Dichroism; Fluorescence; Hydrogen-Ion Concentration; Light; Magnesium Chloride; Plants; Sorbitol; Spinacia oleracea; Thylakoids; Xanthophylls

Arabidopsis thaliana plants grown from ethyl methane sulfonate-treated seeds were screened for so-called que mutants, which are affected in non-photochemical energy quenching. Based on video imaging of chlorophyll fluorescence an energy dissipation mutant, que1, was identified, isolated and characterized. Similar to the npq mutants, the que1 mutant showed a drastically reduced capacity for pH-dependent energy dissipation, qE, but without affecting the Delta pH-dependent conformational changes at 535 nm (DeltaA (535)), which have been supposed to be obligatorily correlated with qE and to reflect pH-regulated binding of zeaxanthin to the PsbS protein. Western blot and DNA sequence analysis revealed that neither a reduced expression of the PsbS protein nor a mutation in the PsbS gene was responsible for the missing qE in que1. Measurements of 9-aminoacridine fluorescence quenching showed that the acidification of the thylakoid lumen was also not affected in the mutant. Furthermore, que1 was able to convert violaxanthin to zeaxanthin. However, unusual characteristics of zeaxanthin formation in the mutant pointed at an altered availability of violaxanthin for de-epoxidation. This was further accompanied by a decrease of the photochemical quenching of chlorophyll fluorescence (qP), an increase of the portion of oxidized P700 and a reduction of the electron transport rate. These characteristics indicate changes in the organization of the thylakoid membrane that affect linear electron transport (but not lumen acidification) and the formation of energy dissipation in photosystem II. Preliminary genetic analysis revealed that the phenotype of que1 is related to two different mutations, mapped to the lower arms of chromosomes 1 and 4.

Topics: Absorption; Arabidopsis; Arabidopsis Proteins; Chlorophyll; Chromosome Mapping; Electron Transport; Fluorescence; Hydrogen-Ion Concentration; Light; Light-Harvesting Protein Complexes; Mutation; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Plant Leaves; Sequence Analysis, DNA; Thylakoids; Xanthophylls; Zeaxanthins

In the present study, we investigated the epoxidation reaction of the violaxanthin (Vx) cycle in intact cells of Chlorella vulgaris. Our results show that the overall epoxidation is slightly slower in darkness compared to the epoxidation during high light (HL) illumination. The calculation of the rate constants of the two epoxidation steps revealed that, for both conditions, the first epoxidation step from zeaxanthin (Zx) to antheraxanthin (Ax) is faster than the second epoxidation step from Ax to Vx. However, the most noteworthy result of our present study is that Ax, which is transiently formed during the epoxidation reaction, participates in non-photochemical quenching of chlorophyll fluorescence (NPQ). A correlation between NPQ and the de-epoxidized xanthophyll cycle pigments during the time-course of the epoxidation reaction can only be achieved when NPQ is plotted versus the sum of Zx and Ax. The accumulation of significant amounts of Ax during the epoxidation reaction further indicates that Ax-dependent quenching proceeds with a similar efficiency compared to the Zx-mediated NPQ. As the xanthophyll-dependent NPQ relies on the presence of de-epoxidized xanthophylls in the PS II antenna, Ax-dependent NPQ is only possible under the assumption that Ax rebinds to the light-harvesting complex (LHC) II during the epoxidation reaction.

Topics: Chlorella vulgaris; Chlorophyll; Darkness; Fluorescence; Kinetics; Light; Light-Harvesting Protein Complexes; Oxidoreductases; Xanthophylls; Zeaxanthins

We analyzed the herbicidal and antioxidant defense responses of transgenic rice plants that overexpressed the Myxococcus xanthus protoporphyrinogen oxidase gene. Leaf squares of the wild-type incubated with oxyfluorfen were characterized by necrotic leaf lesions and increases in conductivity and malonyldialdehyde levels, whereas transgenic lines M4 and M7 did not show any change with up to 100 microM oxyfluorfen. The wild-type had decreased F(v)/F(m) and produced a high level of H(2)O(2) at 18 h after foliar application of oxyfluorfen, whereas transgenic lines M4 and M7 were unaffected. In response to oxyfluorfen, violaxanthin, beta-carotene, and chlorophylls (Chls) decreased in wild-type plants, whereas antheraxanthin and zeaxanthin increased. Only a slight decline in Chls was observed in transgenic lines at 48 h after oxyfluorfen treatment. Noticeable increases of cytosolic Cu/Zn-superoxide dismutase, peroxidase isozymes 1 and 2, and catalase were observed after at 48 h of oxyfluorfen treatment in the wild-type. Non-enzymatic antioxidants appeared to respond faster to oxyfluorfen-induced photodynamic stress than did enzymatic antioxidants. Protective responses for the detoxification of active oxygen species were induced to counteract photodynamic stress in oxyfluorfen-treated, wild-type plants. However, oxyfluorfen-treated, transgenic plants suffered less oxidative stress, confirming increased herbicidal resistance resulted from dual expression of M. xanthus Protox in chloroplasts and mitochondria.

Topics: Antioxidants; beta Carotene; Catalase; Chlorophyll; Drug Resistance; Gene Expression Regulation, Enzymologic; Halogenated Diphenyl Ethers; Herbicides; Hydrogen Peroxide; Lipid Peroxidation; Myxococcus xanthus; Oryza; Oxidoreductases Acting on CH-CH Group Donors; Peroxidase; Phenotype; Phenyl Ethers; Plants, Genetically Modified; Protoporphyrinogen Oxidase; Superoxide Dismutase; Xanthophylls

The conversion of violaxanthin (Vx) to zeaxanthin (Zx) in the de-epoxidation reaction of the xanthophyll cycle plays an important role in the protection of chloroplasts against photooxidative damage. Vx is bound to the antenna proteins of both photosystems. In photosystem II, the formation of Zx is essential for the pH-dependent dissipation of excess light energy as heat. The function of Zx in photosystem I is still unclear. In this work we investigated the de-epoxidation characteristics of light-harvesting complex proteins of photosystem I (LHCI) under in vivo and in vitro conditions. Recombinant LHCI (Lhcal-4) proteins were reconstituted with Vx and lutein, and the convertibility of Vx was studied in an in vitro assay using partially purified Vx de-epoxidase isolated from spinach thylakoids. All four LHCI proteins exhibited unique de-epoxidation characteristics. An almost complete Vx conversion to Zx was observed only in Lhca3, whereas Zx formation in the other LHCI proteins decreased in the order Lhca4 > Lhca1 > Lhca2. Most likely, these differences in Vx de-epoxidation were related to the different accessibility of the respective carotenoid binding sites in the distinct antenna proteins. The results indicate that Vx bound to site V1 and N1 is easily accessible for de-epoxidation, whereas Vx bound to L2 is only partially and/or with the slower kinetics convertible to Zx. The de-epoxidation properties determined for the monomeric recombinant proteins were consistent with those obtained for isolated native LHCI-730 and LHCI-680 in the same in vitro assay and the de-epoxidation state found under in vivo conditions in native LHCIs.

Topics: Apoproteins; beta Carotene; Binding Sites; Chlorophyll; Epoxy Compounds; Escherichia coli; Kinetics; Light-Harvesting Protein Complexes; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Pigments, Biological; Recombinant Proteins; Solanum lycopersicum; Spinacia oleracea; Thylakoids; Xanthophylls

Three plant xanthophylls are components of the xanthophyll cycle in which, upon exposure of leaves to high light, the enzyme violaxanthin de-epoxidase (VDE) transforms violaxanthin into zeaxanthin via the intermediate antheraxanthin. Previous work () showed that xanthophylls are bound to Lhc proteins and that substitution of violaxanthin with zeaxanthin induces conformational changes and fluorescence quenching by thermal dissipation. We have analyzed the efficiency of different Lhc proteins to exchange violaxanthin with zeaxanthin both in vivo and in vitro. Light stress of Zea mays leaves activates VDE, and the newly formed zeaxanthin is found primarily in CP26 and CP24, whereas other Lhc proteins show a lower exchange capacity. The de-epoxidation system has been reconstituted in vitro by using recombinant Lhc proteins, recombinant VDE, and monogalactosyl diacylglycerol (MGDG) to determine the intrinsic capacity for violaxanthin-to-zeaxanthin exchange of individual Lhc gene products. Again, CP26 was the most efficient in xanthophyll exchange. Biochemical and spectroscopic analysis of individual Lhc proteins after de-epoxidation in vitro showed that xanthophyll exchange occurs at the L2-binding site. Xanthophyll exchange depends on low pH, implying that access to the binding site is controlled by a conformational change via lumenal pH. These findings suggest that the xanthophyll cycle participates in a signal transduction system acting in the modulation of light harvesting versus thermal dissipation in the antenna system of higher plants.

Topics: beta Carotene; Chlorophyll; Chlorophyll A; Epoxide Hydrolases; Glycerides; Light; Light-Harvesting Protein Complexes; Oxidation-Reduction; Photosynthetic Reaction Center Complex Proteins; Plant Leaves; Plastids; Spectrophotometry; Xanthophylls; Zea mays

Under high-light conditions, photoprotective mechanisms minimize the damaging effects of excess light. A primary photoprotective mechanism is thermal dissipation of excess excitation energy within the light-harvesting complex of photosystem II (LHCII). Although roles for both carotenoids and specific polypeptides in thermal dissipation have been reported, neither the site nor the mechanism of this process has been defined precisely. Here, we describe the physiological and molecular characteristics of the Chlamydomonas reinhardtii npq5 mutant, a strain that exhibits little thermal dissipation. This strain is normal for state transition, high light-induced violaxanthin deepoxidation, and low light growth, but it is more sensitive to photoinhibition than the wild type. Furthermore, both pigment data and measurements of photosynthesis suggest that the photosystem II antenna in the npq5 mutant has one-third fewer light-harvesting trimers than do wild-type cells. The npq5 mutant is null for a gene designated Lhcbm1, which encodes a light-harvesting polypeptide present in the trimers of the photosystem II antennae. Based on sequence data, the Lhcbm1 gene is 1 of 10 genes that encode the major LHCII polypeptides in Chlamydomonas. Amino acid alignments demonstrate that these predicted polypeptides display a high degree of sequence identity but maintain specific differences in their N-terminal regions. Both physiological and molecular characterization of the npq5 mutant suggest that most thermal dissipation within LHCII of Chlamydomonas is dependent on the peripherally associated trimeric LHC polypeptides.

Topics: Algal Proteins; Amino Acid Sequence; Animals; beta Carotene; Carotenoids; Chlamydomonas; Chlorophyll; Chlorophyll A; Fluorescence; Light; Light-Harvesting Protein Complexes; Molecular Sequence Data; Mutation; Organisms, Genetically Modified; Oxygen; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Pigments, Biological; Sequence Alignment; Sequence Homology, Amino Acid; Temperature; Xanthophylls

Photosystem I (PSI) of higher plants contains 18 subunits. Using Arabidopsis En insertion lines, we have isolated knockout alleles of the genes psaG, psaH2, and psaK, which code for PSI-G, -H, and -K. In the mutants psak-1 and psag-1.4, complete loss of PSI-K and -G, respectively, was confirmed, whereas the residual H level in psah2-1.4 is due to a second gene encoding PSI-H, psaH1. Double mutants, lacking PSI-G, and also -K, or a fraction of -H, together with the three single mutants were characterized for their growth phenotypes and PSI polypeptide composition. In general, the loss of each subunit has secondary, in some cases additive, effects on the abundance of other PSI polypeptides, such as D, E, H, L, N, and the light-harvesting complex I proteins Lhca2 and 3. In the G-less mutant psag-1.4, the variation in PSI composition suggests that PSI-G stabilizes the PSI-core. Levels of light-harvesting complex I proteins in plants, which lack simultaneously PSI-G and -K, indicate that PSI subunits other than G and K can also bind Lhca2 and 3. In the same single and double mutants, psag-1.4, psak-1, psah2-1.4, psag-1.4/psah2-1.4, and psag-1.4/psak-1 photosynthetic electron flow and excitation energy quenching were analyzed to address the roles of the various subunits in P700 reduction (mediated by PSI-F and -N) and oxidation (PSI-E), and state transitions (PSI-H). Based on the results, we also suggest for PSI-K a role in state transitions.

Topics: Alleles; Arabidopsis; Base Sequence; beta Carotene; Blotting, Western; Chlorophyll; Light-Harvesting Protein Complexes; Lutein; Mutation; Oxidation-Reduction; Oxygen; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Pigments, Biological; Plant Leaves; Plant Proteins; Reactive Oxygen Species; Sequence Homology, Nucleic Acid; Thylakoids; Xanthophylls; Zeaxanthins

Guard cells of the orchid genus, Paphiopedilum have been reported to lack developed chloroplasts and detectable chlorophyll a autofluorescence. Paphiopedilum stomata lack a photosynthesis-dependent opening response but have a blue light-specific opening. The present study found that low fluence rate green and red light elicited stomatal opening in Paphiopedilum and this opening was reversed by far red light, indicating the presence of a phytochrome-mediated opening response. Phytochrome-dependent, red light-stimulated opening was largest under low fluence rates and decreased to near zero as fluence rate increased. A recently discovered green light reversibility of blue light-specific stomatal opening was used to probe the properties of the blue light response in Paphiopedilum stomata. Blue light-stimulated opening was completely reversed by green light in the presence of far red light. Red light enhanced the blue light response of Paphiopedilum guard cells when given as a pretreatment or together with blue light. Analysis of guard cell pigments showed that guard cells have small amounts of chlorophyll a and b, zeaxanthin, violaxanthin, antheraxanthin and lutein. Zeaxanthin content increased in response to blue light or ascorbate and declined in the dark or under illumination in the presence of dithiothreitol, indicating the presence of an active xanthophyll cycle. Thus Paphiopedilum stomata possess both a blue light-mediated opening response with characteristics similar to species with normal chloroplast development and a novel phytochrome-mediated opening response.

Topics: Adaptation, Physiological; Ascorbic Acid; beta Carotene; Chlorophyll; Chlorophyll A; Dithiothreitol; Light; Lutein; Orchidaceae; Photosynthesis; Phytochrome; Pigments, Biological; Plant Epidermis; Xanthophylls; Zeaxanthins

Light-harvesting complexes (LHC) of higher plants are composed of at least 10 different proteins. Despite their pronounced amino acid sequence homology, the LHC of photosystem II show differences in pigment binding that are interpreted in terms of partly different functions. By contrast, there is only scarce knowledge about the pigment composition of LHC of photosystem I, and consequently no concept of potentially different functions of the various LHCI exists. For better insight into this issue, we isolated native LHCI-730 and LHCI-680. Pigment analyses revealed that LHCI-730 binds more chlorophyll and violaxanthin than LHCI-680. For the first time all LHCI complexes are now available in their recombinant form; their analysis allowed further dissection of pigment binding by individual LHCI proteins and analysis of pigment requirements for LHCI formation. By these different approaches a correlation between the requirement of a single chlorophyll species for LHC formation and the chlorophyll a/b ratio of LHCs could be detected, and indications regarding occupation of carotenoid-binding sites were obtained. Additionally the reconstitution approach allowed assignment of spectral features observed in native LHCI-680 to its components Lhca2 and Lhca3. It is suggested that excitation energy migrates from chlorophyll(s) fluorescing at 680 (Lhca3) via those fluorescing at 686/702 nm (Lhca2) or 720 nm (Lhca3) to the photosystem I core chlorophylls.

Topics: beta Carotene; Binding Sites; Chlorophyll; Chlorophyll A; Light-Harvesting Protein Complexes; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Photosystem II Protein Complex; Pigments, Biological; Plant Leaves; Solanum lycopersicum; Spectrometry, Fluorescence; Xanthophylls

The steady state absorption and fluorescence spectroscopic properties of the xanthophylls, violaxanthin, zeaxanthin, and lutein, and the efficiencies of singlet energy transfer from the individual xanthophylls to chlorophyll have been investigated in recombinant CP26 protein overexpressed in Escherichia coli and then refolded in vitro with purified pigments. Also, the effect of the different xanthophylls on the extents of static and dynamic quenching of chlorophyll fluorescence has been investigated. Absorption, fluorescence, and fluorescence excitation demonstrate that the efficiency of light harvesting from the xanthophylls to chlorophyll a is relatively high and insensitive to the particular xanthophyll that is present. A small effect of the different xanthophylls is observed on the extent of quenching of Chl fluorescence. The data provide the precise wavelengths of the absorption and fluorescence features of the bound pigments in the highly congested spectral profiles from these light-harvesting complexes. This information is important in assessing the mechanisms by which higher plants dissipate excess energy in light-harvesting proteins.

Topics: beta Carotene; Chlorophyll; Chlorophyll A; Energy Transfer; Escherichia coli; Light-Harvesting Protein Complexes; Lutein; Photochemistry; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Pigments, Biological; Recombinant Proteins; Spectrometry, Fluorescence; Spinacia oleracea; Xanthophylls; Zeaxanthins

Chlamydomonas reinhardtii double mutant npq2 lor1 lacks the beta, epsilon-carotenoids lutein and loroxanthin as well as all beta,beta-epoxycarotenoids derived from zeaxanthin (e.g. violaxanthin and neoxanthin). Thus, the only carotenoids present in the thylakoid membranes of the npq2 lor1 cells are beta-carotene and zeaxanthin. The effect of these mutations on the photochemical apparatus assembly and function was investigated. In cells of the mutant strain, the content of photosystem-II (PSII) and photosystem-I (PSI) was similar to that of the wild type, but npq2 lor1 had a significantly smaller PSII light-harvesting Chl antenna size. In contrast, the Chl antenna size of PSI was not truncated in the mutant. SDS-PAGE and Western blot analysis qualitatively revealed the presence of all LHCII and LHCI apoproteins in the thylakoid membrane of the mutant. The results showed that some of the LHCII and most of the LHCI were assembled and functionally connected with PSII and PSI, respectively. Photon conversion efficiency measurements, based on the initial slope of the light-saturation curve of photosynthesis and on the yield of Chl a fluorescence in vivo, showed similar efficiencies. However, a significantly greater light intensity was required for the saturation of photosynthesis in the mutant than in the wild type. It is concluded that zeaxanthin can successfully replace lutein and violaxanthin in most of the functional light-harvesting antenna of the npq2 lor1 mutant.

Topics: Animals; beta Carotene; Chlamydomonas; Chlorophyll; Chloroplasts; Darkness; Light; Light-Harvesting Protein Complexes; Lutein; Mutation; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Photosystem II Protein Complex; Plants; Proteins; Thylakoids; Xanthophylls

The xanthophyll cycle is an enzymatic, reversible process through which the carotenoids violaxanthin, antheraxanthin, and zeaxanthin are interconverted in response to the need to balance light absorption with the capacity to use the energy to drive the reactions of photosynthesis. The cycle is thought to be one of the main avenues for safely dissipating excitation energy absorbed by plants in excess of that needed for photosynthesis. One of the key factors needed to elucidate the molecular mechanism by which the potentially damaging excess energy is dissipated is the energy of the lowest excited singlet (S(1)) state of the xanthophyll pigments. Absorption from the ground state (S(0)) to S(1) is forbidden by symmetry, making a determination of the S(1) state energies of these molecules by absorption spectroscopy very difficult. Fluorescence spectroscopy is potentially the most direct method for obtaining the S(1) state energies. However, because of problems with sample purity, low emission quantum yields, and detection sensitivity, fluorescence spectra from these molecules, until now, have never been reported. In this work these technical obstacles have been overcome, and S(1) --> S(0) fluorescence spectra of violaxanthin and zeaxanthin are presented. The energies of the S(1) states deduced from the fluorescence spectra are 14 880 +/- 90 cm(-)(1) for violaxanthin and 14 550 +/- 90 cm(-)(1) for zeaxanthin. The results provide important insights into the mechanism of nonphotochemical dissipation of excess energy in plants.

Topics: beta Carotene; Chlorophyll; Chlorophyll A; Energy Transfer; Lutein; Mathematical Computing; Normal Distribution; Photochemistry; Spectrometry, Fluorescence; Spinacia oleracea; Xanthophylls; Zeaxanthins

Non-photochemical quenching of chlorophyll fluorescence in plants occurs in the light harvesting antenna of photosystem II and is regulated by the xanthophyll cycle. A new in vitro model for this process has been developed. Purified light harvesting complexes above the detergent critical micelle concentration have a stable high fluorescence yield but a rapidly inducible fluorescence quenching occurs upon addition of zeaxanthin. Violaxanthin was without effect, lutein and antheraxanthin induced a marginal response, whereas the violaxanthin analogue, auroxanthin, induced strong quenching. Quenching was not caused by aggregation of the complexes but was accompanied by a spectral broadening and red shift, indicating a zeaxanthin-dependent alteration in the chlorophyll environment.

Topics: beta Carotene; Chlorophyll; Fluorescence; Light-Harvesting Protein Complexes; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Spinacia oleracea; Xanthophylls; Zeaxanthins

Infiltrating detached maize (Zea mays L.) leaves with L-galactono-1,4-lactone (L-GAL) resulted in a 4-fold increase in the content of leaf ascorbate. Upon exposure to high irradiance (1000 mumol photons m-2 s-1) at 5 degrees C, L-GAL leaves de-epoxidized the xanthophyll-cycle pigments faster than the control leaves; the maximal ratio of de-epoxidized xanthophyll-cycle pigments to the whole xanthophyll-cycle pool was the same in both leaf types. The elevated ascorbate content, together with the faster violaxanthin de-epoxidation, did not affect the degree of photoinhibition and the kinetics of the recovery from photoinhibition, assayed by monitoring the maximum quantum efficiency of photosystem II primary photochemistry (Fv/Fm). Under the experimental conditions, the thermal energy dissipation seems to be zeaxanthin-independent since, in contrast to the de-epoxidation, the decrease in the efficiency of excitation-energy capture by open photosystem II reaction centers (F'v/F'm) during the high-irradiance treatment at low temperature showed the same kinetic in both leaf types. This was also observed for the recovery of the maximal fluorescence after stress. Furthermore, the elevated ascorbate content did not diminish the degradation of pigments or alpha-tocopherol when leaves were exposed for up to 24 h to high irradiance at low temperature. Moreover, a higher content of ascorbate appeared to increase the requirement for reduced glutathione.

Topics: Ascorbic Acid; beta Carotene; Chlorophyll; Chlorophyll A; Cold Temperature; Energy Metabolism; Light; Light-Harvesting Protein Complexes; Oxidative Stress; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Pigments, Biological; Plant Leaves; Sugar Acids; Xanthophylls; Zea mays; Zeaxanthins

When light energy absorbed by plants becomes excessive relative to the capacity of photosynthesis, the xanthophyll violaxanthin is reversibly deepoxidized to zeaxanthin (violaxanthin cycle). The protective function of this phenomenon was investigated in a mutant of Arabidopsis thaliana, npq1, that has no functional violaxanthin deepoxidase. Two major consequences of the npq1 mutation are the absence of zeaxanthin formation in strong light and the partial inhibition of the quenching of singlet excited chlorophylls in the photosystem II light-harvesting complexes. Prolonged exposure of whole plants to bright light resulted in a limited photoinhibition of photosystem II in both npq1 and wild-type leaves, although CO(2) fixation and the linear electron transport in npq1 plants were reduced substantially. Lipid peroxidation was more pronounced in npq1 compared with the wild type, as measured by chlorophyll thermoluminescence, ethane production, and the total hydroperoxy fatty acids content. Lipid peroxidation was amplified markedly under chilling stress, and photooxidative damage ultimately resulted in leaf bleaching and tissue necrosis in npq1. The npq4 mutant, which possesses a normal violaxanthin cycle but has a limited capacity of quenching singlet excited chlorophylls, was rather tolerant to lipid peroxidation. The double mutant, npq4 npq1, which differs from npq4 only by the absence of the violaxanthin cycle, exhibited an increased susceptibility to photooxidative damage, similar to that of npq1. Our results demonstrate that the violaxanthin cycle specifically protects thylakoid membrane lipids against photooxidation. Part of this protection involves a mechanism other than quenching of singlet excited chlorophylls.

Topics: Arabidopsis; beta Carotene; Carotenoids; Chlorophyll; Ethane; Light; Light-Harvesting Protein Complexes; Lipid Peroxidation; Membrane Lipids; Oxidative Stress; Oxidoreductases; Photons; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Vitamin E; Xanthophylls; Zeaxanthins

Violaxanthin de-epoxidase (VDE) is a lumen-localized enzyme that catalyzes the de-epoxidation of violaxanthin in the thylakoid membrane upon formation of a transthylakoid pH gradient. We investigated the developmental expression of VDE in leaves of mature tobacco (Nicotiana tabacum) plants grown under high-light conditions (in the field) and low-light conditions (in a growth chamber). The difference in light conditions was evident by the increased pool size (violaxanthin + antheraxanthin + zeaxanthin, VAZ) throughout leaf development in field-grown plants. VDE activity based on chlorophyll or leaf area was low in the youngest leaves, with the levels increasing with increasing leaf age in both high- and low-light-grown plants. However, in high-light-grown plants, the younger leaves in early leaf expansion showed a more rapid increase in VDE activity and maintained higher levels of VDE transcript in more leaves, indicating that high light may induce greater levels of VDE. VDE transcript levels decreased substantially in leaves of mid-leaf expansion, while the levels of enzyme continued to increase, suggesting that the VDE enzyme does not turn over rapidly. The level of VDE changed in an inverse, nonlinear relationship with respect to the VAZ pool, suggesting that enzyme levels could be indirectly regulated by the VAZ pool.

Topics: beta Carotene; Chlorophyll; Chloroplasts; Chromatography, High Pressure Liquid; Dose-Response Relationship, Radiation; Gene Expression Regulation, Plant; Kinetics; Light; Nicotiana; Oxidoreductases; Pigments, Biological; Plant Leaves; Plants, Toxic; RNA, Messenger; RNA, Plant; Time Factors; Xanthophylls

Singlet energy transfer between the carotenoids (Cars) and chlorophylls (Chls) in the light-harvesting complex II (LHC II) from higher plants has been studied using ultrafast transient absorption spectroscopy by exciting the Cars directly in the 475-515 nm wavelength range. LHC II trimers from Arabidopsis thaliana with well-defined Car compositions have been used. From HPLC, the wild type (WT) monomer contains two luteins (Ls), one neoxanthin (N), and a trace of violaxanthin (V) per 12 Chls. The ABA-3 mutant contains 1.4 Ls and 0.6 zeaxanthin (Z) per monomer. Though exploitation of the difference in Car constitution and exciting the WT at 475 and 490 nm, and the ABA-3 mutant at 490 and 515 nm, the different Car contributions to energy transfer have been probed. Evidence for energy transfer mainly from the Car to Chl b is observed in the WT. In the mutant, additional transfer from Car to Chl a correlates with the presence of Z. The results imply predominant energy transfer from the central Ls to Chl b which requires a modification of the currently accepted arrangement of Chl pigments in LHC II.

Topics: Arabidopsis; beta Carotene; Carotenoids; Chlorophyll; Energy Transfer; Kinetics; Light-Harvesting Protein Complexes; Lutein; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Spectrophotometry; Time Factors; Xanthophylls

Excess light energy absorbed by the chloroplast membranes of green plants is dissipated by nonradiative de-excitation in order to protect against photodamage. This is observed as the nonphotochemical quenching of chlorophyll fluorescence, which has been suggested to result from an alteration in the structure and function of the chlorophyll a/b light-harvesting complexes of photosystem II (LHCII) due to the combined effects of protonation and the de-epoxidation of bound violaxanthin to form zeaxanthin. In agreement with this hypothesis, it is shown that the light-harvesting chlorophyll a/b proteins purified from spinach leaves exhibit pH-stimulated quenching of chlorophyll fluorescence; this quenching shares all the key features observed for the nonphotochemical quenching of chlorophyll fluorescence in vivo. In the case of the two minor complexes, LHCIIa (CP29) and LHCIIc (CP26), quenching is much greater than in the bulk complex LHCIIb and is strongly inhibited by the reagent dicyclohexylcarbodiimide. The carotenoids violaxanthin and zeaxanthin cause strong inhibition and stimulation of quenching, respectively, in these complexes. The results of this study are consistent with the suggestion that the minor light-harvesting complexes play a crucial role in photoprotective energy dissipation in the photosynthetic membrane of green plants. Moreover, for the first time, a system using isolated LHCIIa and LHCIIc for the study of the regulation of light harvesting is described.

Topics: beta Carotene; Carotenoids; Carrier Proteins; Chlorophyll; Chlorophyll A; Fluorescence; Light-Harvesting Protein Complexes; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Plants; Xanthophylls

In higher plants non-photochemical dissipation of excess light, trapped by the pigment pool of photosystem II, prevents photodamage to the photosynthetic apparatus. We report here that an algal virus infecting Chlorella strain Pbi induces non-photochemical quenching of photosystem II fluorescence, indicating enhanced loss of absorbed light energy from photosystem II. This phenomenon occurs soon after the establishment of the virus infection cycle and is observed at low irradiance (20 micromol quanta m-2 s-1). At low light, infection associated non-photochemical quenching is not linked to extensive conversion of violaxanthin to antheraxanthin and zeaxanthin. However, such conversion occurs rapidly (2-10 min) in infected cells under conditions of high irradiance (100-300 micromol quanta m-2 s-1). Under similar conditions uninfected Chlorella cells do not display significant changes in non-photochemical quenching.

Topics: beta Carotene; Carotenoids; Chlorella; Chlorophyll; Cycloheximide; Dithiothreitol; Epoxy Compounds; Fluorescence; Genes, Viral; Light; Light-Harvesting Protein Complexes; Lutein; Paraquat; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Phycodnaviridae; Pigments, Biological; Xanthophylls; Zeaxanthins

Changes in carotenoid composition, CO2 assimilation and chlorophyll fluorescence due to photoinhibition at 5 degrees C and 20 degrees C were studied in 12 day and 30 day old sorghum leaves. The old leaves had a higher violaxanthin (V) content and less beta-carotene. Photoinhibition at both temperatures caused significant increases in zeaxanthin (Z) and decreases in violaxanthin. However, in young leaves the increase in zeaxanthin was greater than the decrease in violaxanthin. In young leaves the V + A + Z pool size (A = antheraxanthin) almost doubled under photoinhibitory conditions (compared to controls) while in old leaves the V + A + Z pool remained approximately constant. After photoinhibition treatment changes in the levels of the xanthophylls were restored during a recovery period both in young and old leaves. When rephotoinhibited after a 48 hr recovery period, the young plants showed better protection against photoinhibition. We suggest that in young leaves zeaxanthin is newly synthesized under photoinhibitory conditions besides being de-epoxidized from violaxanthin and that the synthesis of V + A + Z pool is higher at 20 degrees C than at 5 degrees C in both young and old leaves.

Topics: beta Carotene; Carbon Dioxide; Carotenoids; Chlorophyll; Chloroplasts; Fluorescence; Light; Lutein; Pigments, Biological; Plant Leaves; Temperature; Time Factors; Xanthophylls; Zeaxanthins

Acrocarpia paniculata thylakoids were fragmented with Triton X-100 and the pigment-protein complexes so released were isolated by sucrose density gradient centrifugation. Three main chlorophyll-carotenoid-protein complexes with distinct pigment compositions were isolated. (1) A P-700-chlorophyll a-protein complex, with a ratio of 1 P-700: 38 chlorophyll a: 4 beta-carotene molecules, had similar absorption and fluorescence characteristics to the chlorophyll-protein complex 1 isolated with Triton X-100 from higher plants, green algae and Ecklonia radiata. (2) an orange-brown complex had a chlorophyll a : c2 : fucoxanthin molar ratio of 2 : 1 : 2. this complex had no chlorophyll c1 and contained most of the fucoxanthin present in the chloroplasts. This pigment complex is postulated to be the main light-harvesting complex of brown seaweeds. (3) A green complex had a chlorophyll a : c1 : c2 : violaxanthin molar ratio of 8 : 1 : 1. This also is a light-harvesting complex. the absorption and fluorescence spectral characteristics and other physical properties were consistent with the pigments of these three major complexes being bound to protein. Differential extraction of brown algal thylakoids with Triton X-100 showed that a chlorophyll c2-fucoxanthin-protein complex was a minor pigment complex of these thylakoids.

Topics: beta Carotene; Carotenoids; Centrifugation, Density Gradient; Chlorophyll; Chloroplasts; Phaeophyceae; Photochemistry; Pigments, Biological; Plant Proteins; Seaweed; Spectrometry, Fluorescence; Spectrum Analysis; Xanthophylls