Pigment Organization Research Articles

Molecular organisation of the chlorophyll a/c light-harvesting complex of Pleurochloris meiringensis (Xanthophyceae). Pigment binding and secondary structure of the protein

In our previous study by means of circular dichroism (CD) spectroscopy we have shown that the pigment organisation in the chlorophyll a/c light harvesting complex (Chl a/c LHC) isolated from Pleurochloris meiringensis significantly differs from the architecture of the main Chl a/b light harvesting antenna complexes of higher plants (Büchel and Garab. J. Photochem. Photobiol., B: Biol., 37 (1997) 118–124). In this work we measured the CD spectra in the far-UV, between 190 nm and 240 nm. and investigated the variations of the secondary structure of the protein upon treatments which affect the binding of the long wavelength Chl a. We found that low concentrations ( < 5%) of acetone, which had no noticeable effect on the (−)679 nm CD band, drastically reduced the α-helical content of the protein. In contrast, amounts of digitonin, which completely abolished this intense, non-conservative CD band of Chl a, induced only minor changes in the secondary structure of the protein complex. These data suggest that the binding site of the long-wavelength absorbing Chl a is found in a position deeply buried in the protein, and it is most likely co-ordinated by two helices.

Excitations and excitons in photosystem I

The elementary requirements for energy transfer in a photosynthetic antenna are discussed. Energy migration between chlorophyll molecules in the PSI antenna reaction centre complex was modelled using a geometry based on the recent X–ray structure. The migration is trap–limited and an intrinsic electron transfer rate constant of 1.3 ps−1 was obtained by modifying the model to account for experimental results obtained from antennae of different sizes. Exciton coupling between pairs of Chlorophyll molecules was calculated and it is found that to prevent self–quenching their interplanar separation must be greater than or equal to 5 A. In the PSI reaction centre, exciton coupling between adjacent chlorophylls was found to be around 300 cm−1 with no particularly large value between the special pair being necessary to describe the experimentally observed transient spectra of P +700− P 700 and A −− A . Electrochromic interaction is particularly important in reproducing the A −− A spectrum.

In vitro reconstitution of the photosystem I light-harvesting complex LHCI-730: heterodimerization is required for antenna pigment organization.

Here we describe the in vitro reconstitution of photosystem I light-harvesting complexes with pigments and proteins (Lhca1 and Lhca4) obtained by overexpression of tomato Lhca genes in Escherichia coli. Using Lhca1 and Lhca4 individually for reconstitution results in monomeric pigment-proteins, whereas a combination thereof yields a dimeric complex. Interactions of the apoproteins is highly specific, as reconstitution of either of the two constituent proteins in combination with a light-harvesting protein of photosystem II does not result in dimerization. The reconstituted Lhca1/4, but not complexes obtained with either Lhca1 or Lhca4 alone, closely resembles the native LHCI-730 dimer from tomato leaves with regard to spectroscopic properties, pigment composition, and stoichiometry. Monomeric complexes of Lhca1 or Lhca4 possess lower pigment/protein ratios, indicating that interactions of the two subunits not only facilitates pigment reorganization but also recruitment of additional pigments. In addition to higher averages of chlorophyll a/b ratios in monomeric complexes than in LHCI-730, comparative fluorescence and CD spectra demonstrate that heterodimerization involves preferential ligation of more chlorophyll b.

Pigment Organization and Transfer of Electronic Excitation in the Photosynthetic Unit of Purple Bacteria

Absorption of light by light-harvesting complexes and transfer of electronic excitation to the photosynthetic reaction center (RC) constitute the primary light-harvesting process of photosynthesis. This process is investigated on the basis of an atomic level structure of the so-called photosynthetic unit of the photosynthetic bacterium Rhodobacter sphaeroides. The photosynthetic unit combines in the intracytoplasmic membrane a nanometric assembly of three pigment−protein complexes: (i) the photosynthetic reaction center, (ii) a ring-shaped light-harvesting complex LH-I, and (iii) multiple copies of a similar complex, LH-II. The unit has been modeled using the known structure of (i) and for (ii) a model structure complexed appropriately with (i); for (iii) the structure of LH-II of Rhodospirillum molischianum is substituted. The model describes in detail the organization of chromophores involved in primary light absorption and excitation transfer: a hierarchy of ring-shaped bacteriochlorophyll aggregates that surround four centrally located bacteriochlorophylls of the photosynthetic reaction center. The bacteriochlorophylls involved in the overall transfer are found in a coplanar arrangement. On the basis of the modeled structure a quantum-mechanical description of the entire light-harvesting process is developed. For this purpose an effective Hamiltonian is established a priori and then employed to describe the LH-II → LH-II → LH-I → RC cascade of excitation transfer. The transfer times calculated are in agreement with measured transfer times. The results suggest that excitons are the key carriers of the excitation transferred; i.e., electronic excitations are coherently delocalized in the photosynthetic unit. This suggestion is corroborated by an investigation of the effect of inhomogeneous broadening on the predicted excitons in LH-II and LH-I, an effect that is found to be significant but small. A particularly important role is played by the lowest energy excitons to which the circular arrangement of bacteriochlorophylls imparts vanishing oscillator strength. Despite the lack of oscillator strength, the low-energy excitons are well suited for exciton transfer on a subpicosecond and picosecond time scale. The accessory bacteriochlorophylls of the photosynthetic reaction center are found to be critical for the LH-I → RC transfer, which would take several hundred picoseconds without these bacteriochlorophylls.

Identification of a photochemically inactive pheophytin molecule in the spinach D 1-D 2-cyt b559 complex

A photochemically inactive pheophytin (Pheo) a molecule in the spinach D 1-D 2-cyt b 559 complex, which contains approximately 6 chlorophyll (Chl) a molecules and two β-carotene molecules for every two Pheo a molecules, was identified by fluorescence excitation spectroscopy at 77K, and confirmed by absorption, linear dichroism (LD), and magnetic circular dichroism spectra. When the emission was monitored at 740 nm, photochemically active Pheo a was detected as peaks at 418, 513, 543, and 681 nm, as described in previous reports. However, when the emission was monitored at 665 nm, two bands were observed at 414 and 537 nm; these bands were insensitive to photochemical reduction of the primary acceptor and thus appear to be due to photochemically inactive Pheo a. The Qy band of this Pheo a molecule at 77K was estimated to be at 671 ± 1 nm, based on the magnitude of the band-shift in photochemically active Pheo a. Two β-carotene molecules in the complex were also discriminated by fluorescence excitation and LD spectra. These observations were discussed in the light of current understanding of the molecular organization of pigments and the relationship of pigment arrangement to optical properties in the reaction center of photosystem II.

Models for the Pigment Organization in the Chlorosomes of Photosynthetic Bacteria: Diastereoselective Control of in-Vitro Bacteriochlorophyll cs Aggregation. [Erratum to document cited in CA122:76986

Models for the Pigment Organization in the Chlorosomes of Photosynthetic Bacteria: Diastereoselective Control of in-Vitro Bacteriochlorophyll cs Aggregation. [Erratum to document cited in CA122:76986

Femtosecond probe of structural analogies between chlorosomes and bacteriochlorophyll c aggregates

Bacteriochlorophyll c pigments extracted from light harvesting chlorosomes in green photosynthetic bacteria are known to self-assemble into aggregates whose electronic spectroscopy resembles that of intact chlorosomes. Femtosecond optical experiments reveal that the chlorosomes and their reconstituted aggregates exhibit closely analogous internal energy transfer kinetics and exciton state evolution. These comparisons furnish compelling new evidence that proteins do not exert a major local role in the BChl c antenna pigment organization of intact chlorosomes.

Description of energy migration and trapping in photosystem I by a model with two distance scaling parameters

The energy transfer and trapping kinetics in the core antenna of Photosystem I are described in a new model in which the distance between the core antenna chlorophylls and P700 is proposed to be considerably longer than the distance between the chlorophylls within the antenna. Structurally, the model describes the Photosystem I core antenna as a regular sphere around P700, while energetically it consists of three levels representing the bulk antenna, P700 and the red-shifted antenna pigments absorbing at longer wavelength than P700, respectively. It is shown that the model explains experimental results obtained from the Photosystem I complex of the cyanobacterium Synechococcus sp. (A.R. Holzwarth, G. Schatz, H Brock, and E. Bittersman (1993) Biophys. J. 64: 1813-1826) quite well, and that no unrealistic charge separation rate and organization of the long-wavelength pigments has to be assumed. We suggest that excitation energy transfer and trapping in Photosystem I should be described as a 'transfer-to-the-trap'-limited process.

Models for the Pigment Organization in the Chlorosomes of Photosynthetic Bacteria: Diastereoselective Control of in-vitro Bacteriochlorophyll cs Aggregation

Models for the Pigment Organization in the Chlorosomes of Photosynthetic Bacteria: Diastereoselective Control of in-vitro Bacteriochlorophyll cs Aggregation

ANTARCTIC CYANOBACTERIA: LIGHT, NUTRIENTS, AND PHOTOSYNTHESIS IN THE MICROBIAL MAT ENVIRONMENT1

ABSTRACTThe microenvironmental and photosynthetic characteristics of Antarctic microbial mats were measured in a series of ponds near McMurdo Sound. As elsewhere in Antarctica, these cold‐water benthic communities were dominated by oscillatoriacean cyanobacteria. Despite large variations in mat thickness, surface morphology, and color, all of the communities had a similar pigment organization, with a surface carotenoid‐rich layer that overlaid a deep chlorophyll maximum (DCM) enriched in phycocyanin as well as chlorophyll a. Spectroradiometric analyses showed that the DCM population inhabited an orange‐red shade environment. In several of the mats, the deep‐living trichomes migrated up to the surface of the mat within 2 h in response to a 10‐fold decrease in surface irradiance. The euphotic layer of the mats was supersaturated in oxygen and contained ammonium and dissolved reactive phosphorus concentrations in excess of 100 mg N·m−3 or P·m−3. Integral photosynthesis by core samples was saturated at low irradiances and varied two‐ to threefold throughout the continuous 24‐h radiation cycle. Oxygen microelectrode analyses showed that the photosynthetic rates were slow to negligible near the surface and maximal in the DCM. These compressed, nutrient‐rich euphotic zones have some properties analogous to planktonic systems, but the integrated photosynthetic responses of the community reflect the strong self‐shading within the mat and physiological dominance by the motile, DCM populations.

Community structure and pigment organisation of cyanobacteria-dominated microbial mats in Antarctica

Benthic microbial mat communities were sampled from 20 lakes, ponds and streams of the McMurdo Sound region, Antarctica. At least five distinct assemblages could be differentiated by their cyanobacterial species composition, pigment content and vertical structure. The most widely occurring freshwater communities were dominated by thin-trichome (0·5–3 µm) oscillatoriacean species that formed benthic films up to several millimetres thick. ‘Lift-off mats’ produced mucilaginous mats 1–5 cm thick at the surface and edge of certain ponds. Another group of oscillatoriacean communities was characteristic of hypersaline pond environments; these communities were dominated by species with thicker trichomes such as Oscillatoria priestleyi. Black mucilaginous layers of Nostoc commune were widely distributed in aquatic and semi-aquatic habitats. Dark brown sheath pigmentation was also characteristic of less cohesive mats and crusts dominated by Pleurocapsa, Gloeocapsa and Calothrix. High performance liquid chromatograp...

A linear dichroism study of photosynthetic pigment organisation in two fucoxanthin-containing algae

The orientation of the pigments in cells, thylakoids and isolated pigment-protein complexes of two chromophytic algae containing fucoxanthin as the principle carotenoid, has been investigated by linear dichroism spectroscopy. The Q y transition of both chlorophyll a (chl a ) and chlorophyll c (chl c ) is oriented at < 35° to the plane of the thylakoid membrane and to the long axis of the cells. The fucoxanthin transition, centred at 540 nm, is oriented in the same manner. In the isolated light-harvesting complexes, in compressed polyacrylamide gels, the linear dichroism specta suggest a different orientation of all pigments, i.e., < 55° to the direction of applied force or 35° to the long axis of the particle. It is demonstrated that this orientation is an artefact arising from decomposition of the light-harvesting complex during its preparation in the compressed polyacrylamide gel. The pigments of isolated PS I and PS II complexes retain the orientation they show in thylakoids and whole cells.

A linear dichroism study of photosynthetic pigment organisation in two fucoxanthin-containing algae

The orientation of the pigments in cells, thylakoids and isolated pigment-protein complexes of two chromophytic algae containing fucoxanthin as the principle carotenoid, has been investigated by linear dichroism spectroscopy. The Q y transition of both chlorophyll a (chl a) and chlorophyll c (chl c) is oriented at < 35° to the plane of the thylakoid membrane and to the long axis of the cells. The fucoxanthin transition, centred at 540 nm, is oriented in the same manner. In the isolated light-harvesting complexes, in compressed polyacrylamide gels, the linear dichroism specta suggest a different orientation of all pigments, i.e., < 55° to the direction of applied force or 35° to the long axis of the particle. It is demonstrated that this orientation is an artefact arising from decomposition of the light-harvesting complex during its preparation in the compressed polyacrylamide gel. The pigments of isolated PS I and PS II complexes retain the orientation they show in thylakoids and whole cells.

Structure, function and organization of antenna polypeptides and antenna complexes from the three families of Rhodospirillaneae

Comparative primary structural analysis of polypeptides from antenna complexes from species of the three families of Rhodospirillaneae indicates the structural principles responsible for the formation of spectrally distinct light-harvesting complexes. In many of the characterized antenna systems the basic structural minimal unit is an α/β polypeptide pair. Specific clusters of amino acid residues, in particular aromatic residues in the C-terminal domain, identify the antenna polypeptides to specific types of antenna systems, such as B880 (strong circular dichroism (CD)), B870 (weak CD), B800–850 (high), B800–850 (low) or B800–820. The core complex B880 (B1020) of species from Ectothiorhodospiraceae and Chromatiaceae apparently consists of four (α 1α 2β 1β 2) or three (2αβ 1β 2) chemically dissimilar antenna polypeptides respectively. There is good evidence that the so-called variable antenna complexes, such as the B800–850 (high), B800–850 (low) or B800–820 of Rp. acidophila, Rp. palustris and Cr. vinosum, are comprised of multiple forms of peripheral light-harvesting polypeptides. Structural similarities between prokaryotic and eukaryotic antenna polypeptides are discussed in terms of similar pigment organization. The structural basis for the strict organization of pigment molecules (bacteriochlorophyll (BChl) cluster) in the antenna system of purple bacteria is the hierarchical organization of the α- and β-antenna polypeptides within and between the antenna complexes. On the basis of the three-domain structure of the antenna polypeptides with the central hydrophobic domain, forming a transmembrane α helix, possible arrangements of the antenna polypeptides in the three-dimensional structure of core and peripheral antenna complexes are discussed. Important structural and functional features of these polypeptides and therefore of the BChl cluster are the α/β heterodimers, the α 2,β 2 basic units and cyclic arrangements of these basic units. Equally important for the formation of the antenna complexes or the entire antenna are polypeptide-polypeptide, pigment-pigment and pigment-polypeptide interactions.

Temperature and Solvent Effects on Reaction Centers from ChlorofLexus aurantiacus and Chromatium tepidum1

Temperature and solvent effects on reaction center structures were examined in two thermophilic photosynthetic bacteria, Chloroflexus aurantiacus and Chromatium tepidum, in order to gain insight into the interactions among the reaction center proteins and pigment systems. Thermal stability of the reaction centers was found to be proportional to the optimum growth temperature. Circular dichroism (CD) spectra in the 250-300 nm region indicated that thermal denaturation destroyed tertiary structures (helix-to-helix interactions or amino acid residue conformation) in the native reaction center, keeping helical structures intact. Absorption and circular dichroism spectral changes showed that alcohol denatured the so-called special pair and the accessory BChl a independently. The alcohol denaturation further indicates that the coordination between BChl a and amino acid residue in the protein is one of the important interactions maintaining the pigment organization of the reaction centers.

Pigment organization and energy transfer in green bacteria. 3. Picosecond energy transfer kinetics within the B806-866 bacteriochlorophyll a antenna complex isolated from Chloroflexus aurantiacus

The B806-866 antenna protein complex from Chloroflexus aurantiacus has been isolated using the detergents lauryldimethylamine N-oxide (LDAO) and Deriphat-160. These preparations have been characterized spectroscopically. While the B806-866 complex was unstable for longer periods in 0.1% LDAO a high stability was observed when 0.1% Deriphat-160 was used as a detergent. In order to study the energy-transfer processes between the 806 and 866 nm absorbing excited states, fluorescence decays were measured with picosecond resolution in the detection wavelength range from 800 to 910 nm upon excitation at 790 nm. A fast energy transfer from the 806 nm to the 866 nm absorbing states was resolved, and was found to have an equilibration time of 5 ± 1 ps. This equilibration time is in line with previous estimates from steady-state fluorescence. Due to the fast equilibration time, resolution of this energy-transfer process has not been achieved in the past when measuring whole cells of C. aurantiacus.

Vibrational spectroscopy of excited electronic states in carotenoids in vivo. Picosecond time-resolved resonance Raman scattering

The vibrational spectroscopy and population dynamics of excited singlet (2(1)Ag), excited triplet (3B u), and the ground (1Ag) electronic states of carotenoids in chromatophores of Chromatium vinosum (mainly spirilloxanthin and rhodopin) and of the same carotenoids in benzene solutions are examined by picosecond time-resolved resonance Raman scattering. Coherent Stokes Raman scattering from the ground states of carotenoids in chromatophores also is observed. Resonance Raman spectra of in vitro rhodopin and spirilloxanthin when compared with in vivo data demonstrate that scattering from spirilloxanthin dominates the in vivo spectrum. Comparisons of the time-dependent intensities of 2(1)Ag and 1Ag resonance Raman bands from both in vitro and in vivo carotenoids suggest that vibrationally excited levels in 1Ag are populated directly by the decay of the 2(1)Ag state and that these levels relax into a thermalized distribution in less than 50 ps. The appearance of asymmetrically broadened, ground-state resonance Raman bands supports this conclusion. Formation of the 3Bu state is observed for carotenoids in chromatophores, but not for in vitro spirilloxanthin indicating that the 3Bu state is formed by fission processes originating from the spatial organization of pigments within chromatophores. The rate at which the intensities of 2(1)Ag resonance Raman bands decay is faster for the carotenoids in vivo than for those in vitro thereby indicating that additional relaxation channels (e.g., energy transfer to bacteriochlorophylls) are present in the chromatophore. The similarity of the in vivo and in vitro 2(1)Ag resonance Raman spectra shows that no significant modifications in the vibronic coupling has been caused by the chromatophore environment.

Pigment organization and energy transfer in green bacteria. 2. Circular and linear dichroism spectra of protein-containing and protein-free chlorosomes isolated from Chloroflexus aurantiacus strain Ok-70-fl

We have measured the circular dichroism (CD) and linear dichroism (LD) spectra of chlorosomes isolated from Chloroflexus aurantiacus strain Ok-70-fl obtained by two different isolation procedures. The gel-electrophoretic filtration procedure yields chlorosomes that are essentially free of BChl a 790 and proteins, while isolation by sucrose density gradient centrifugation yields the conventional chlorosome preparations. The LD spectra of the two kinds of preparation were very similar. In both cases the Q y LD signals correspond to an average angle between the BChl- c -Q y transition and the long axis of the chlorosome of approx. 15 ± 10°. In contrast to the LD spectra, the CD spectra of different preparations (membranes, BChl- a -free chlorosomes, BChl- a -containing chlorosomes) show pronounced differences both in the ellipticity as well as in the shape of the spectra and the number of maxima. However, these differences are not caused by the isolation procedure or the detergents used. We show that even freshly prepared membranes (of different, parallel grown batch cultures) give rise to very different CD spectra. The set of different CD spectra we obtained could be simulated well by linear combinations of two basic spectra. This strongly suggests that the variations in the CD spectra are caused by a variation in the relative amounts of two different species, two different types of chlorosome, or possibly by two different types of pigment aggregate within the chlorosomes.

Dimeric carotenoid interaction in the light-harvesting antenna of purple phototrophic bacteria

The carotenoid content of intracytoplasmic membrane vesicles isolated from purple phototrophic bacteria was reduced to a variable extent by mild extraction with light petroleum. Using preparations obtained from Rhodobacter capsulatus strains that contained the Light Harvesting System I (LHI) complex as the only major photosynthetic holochrome, it was shown that the visible circular dichroism of the carotenoids increased with the square of the membrane carotenoid content, as expected from being caused by dimeric exciton interaction. No chirality resulting from twists of the individual planar chromophore was detected. Therefore the contribution to carotenoid optical activity of non-degenerate interactions with bacteriochlorophyll or the apoprotein does not appear to be significant. The broadening of the absorption band of the bound pigment, caused by the splitting of the monomer transition, was demonstrated in membrane vesicles of both Rb, capsulatus and Rhodospirillum rubrum as a decrease of the fine structure of the band. Furthermore, the dimeric organization of the carotenoid pigments in the bacterial LHI complex accounted for the observed quantitative relationship between the fine structure of the band and the carotenoid content of the membrane.

The Ultrastructure of the Compound Eye of Two Species of Marine Ostracodes (Crustacea: Ostracoda: Cypridinidae)

AbstractUltrastructurally, the compound eyes of the luminescent marine ostracodes Vargula graminkola and V. tsujii are similar. These ostracodes have two lateral compound eyes, with relatively few ommatidia (13 and 20 respectively). They exhibit apposition type compound eyes as seen in many other arthropods. Each ommatidium includes: a flat, ectodermal cuticular covering, corneagen cells, two long cone cells that give rise to a large conspicuous crystalline cone, retinular cells, pigment cells, a microvillar rhabdom and proximal axonal neurons. The axons merge to form an optic nerve that extends into the brain through a short, muscular stalk that is surrounded externally by a cuticle. The number of retinular cells is typically six per ommatidium in V. graminicola and eight per ommatidium in V. tsujii. Screening pigment cells surround each ommatidium forming a layer that is about 5–15 pigment granules thick. In addition to pigment cells, the cytoplasm of the retinular cells includes numerous screening pigment granules. In light/dark adaptation, there are no obvious morphological differences in the orientation of the rhabdom or in the organization of the screening pigments. Both Vargula species studied are nocturnally active and bioluminescent suggesting that these eyes are capable receptors of the bright conspecific luminescence.