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Sample records for membrane protein light-harvesting

  1. Revealing Linear Aggregates of Light Harvesting Antenna Proteins in Photosynthetic Membranes

    PubMed Central

    He, Yufan; Zeng, Xiaohua; Mukherjee, Saptarshi; Rajapaksha, Suneth; Kaplan, Samuel; Lu, H. Peter

    2010-01-01

    How light energy is harvested in a natural photosynthetic membrane through energy transfer is closely related to the stoichiometry and arrangement of light harvesting antenna proteins in the membrane. The specific photosynthetic architecture facilitates a rapid and efficient energy transfer among the light harvesting proteins (LH2 and LH1) and to the reaction center. Here we report the identification of linear aggregates of light harvesting proteins, LH2, in the photosynthetic membranes under ambient conditions by using atomic force microscopy (AFM) imaging and spectroscopic analysis. Our results suggest that the light harvesting protein, LH2, can exist as linear aggregates of 4±2 proteins in the photosynthetic membranes and that the protein distributions are highly heterogeneous. In the photosynthetic membranes examined in our measurements, the ratio of the aggregated to the non-aggregated LH2 proteins is about 3:1 to 5:1 depending on the intensity of the illumination used during sample incubation and on the bacterial species. AFM images further identify that the LH2 proteins in the linear aggregates are monotonically tilted at an angle 4°±2° from the plane of the photosynthetic membranes. The aggregates result in red-shifted absorption and emission spectra that are measured using various mutant membranes, including an LH2 knock-out, LH1 knock-out, and LH2 at different population densities. Measuring the fluorescence lifetimes of purified LH2 and LH2 in membranes, we have observed that the LH2 proteins in membranes exhibit biexponential lifetime decays whereas the purified LH2 proteins gave single exponential lifetime decays. We attribute that the two lifetime components originate from the existence of both aggregated and non-aggregated LH2 proteins in the photosynthetic membranes. PMID:19572507

  2. Biogenesis of light harvesting proteins.

    PubMed

    Dall'Osto, Luca; Bressan, Mauro; Bassi, Roberto

    2015-09-01

    The LHC family includes nuclear-encoded, integral thylakoid membrane proteins, most of which coordinate chlorophyll and xanthophyll chromophores. By assembling with the core complexes of both photosystems, LHCs form a flexible peripheral moiety for enhancing light-harvesting cross-section, regulating its efficiency and providing protection against photo-oxidative stress. Upon its first appearance, LHC proteins underwent evolutionary diversification into a large protein family with a complex genetic redundancy. Such differentiation appears as a crucial event in the adaptation of photosynthetic organisms to changing environmental conditions and land colonization. The structure of photosystems, including nuclear- and chloroplast-encoded subunits, presented the cell with a number of challenges for the control of the light harvesting function. Indeed, LHC-encoding messages are translated in the cytosol, and pre-proteins imported into the chloroplast, processed to their mature size and targeted to the thylakoids where are assembled with chromophores. Thus, a tight coordination between nuclear and plastid gene expression, in response to environmental stimuli, is required to adjust LHC composition during photoacclimation. In recent years, remarkable progress has been achieved in elucidating structure, function and regulatory pathways involving LHCs; however, a number of molecular details still await elucidation. In this review, we will provide an overview on the current knowledge on LHC biogenesis, ranging from organization of pigment-protein complexes to the modulation of gene expression, import and targeting to the photosynthetic membranes, and regulation of LHC assembly and turnover. Genes controlling these events are potential candidate for biotechnological applications aimed at optimizing light use efficiency of photosynthetic organisms. This article is part of a Special Issue entitled: Chloroplast biogenesis.

  3. Formation of crystalline arrays of chlorophyll a/b - light-harvesting protein by membrane reconstitution.

    PubMed Central

    Li, J; Hollingshead, C

    1982-01-01

    The structure of the major protein constituent of photosynthetic membranes in higher plants, the chlorophyll a/b-light harvesting complex (LHC), was studied by x-ray diffraction and electron microscopy. The LHC was purified from Triton X-100 solubilized thylakoid membranes of the pea, and contained 6 mol of chlorophylls a and b per mole of a polypeptide of 27,000 molecular weight. X-ray diffraction showed that in the presence of 10 mM MgCl2, purified LHC forms planar aggregates that stack with a period of 51 A. Within each layer, LHC molecules pack with a center-to-center distance of 85 A but without long-range order. However, when LHC is incorporated into single-walled vesicles of plant lecithin, the addition of NaCl above 10 mM, or MgCl2 above 2 mM, led to the formation of plaques of hexagonal lattices, with a lattice constant of 125 A. The large domain size and high degree of order in the plane of the membrane are evident from the sharp lattice lines observed to 7 A resolution on the equator of the x-ray pattern. Freeze-fracture electron micrographs demonstrated an aligned stacking of the lattices in adjacent membranes, resulting in crystallinity in the third dimension over short distances. Micrographs of negatively stained membranes revealed a hexagonal lattice of the same lattice constant, formed by surface-exposed parts of the LHC molecules which are probably responsible for the ordered stacking of lattices. In both the LHC aggregates and in the reconstituted membrane lattices the diffracted x-ray intensities at 10-A spacing on the equator indicate that the LHC molecule contains paralled alpha-helices or beta-sheets that are oriented perpendicular to the planar arrays. Images FIGURE 1 FIGURE 2 a FIGURE 2 b FIGURE 3 PMID:7034799

  4. Dissociation of the light-harvesting membrane protein complex I from Rhodobacter sphaeroides under high hydrostatic pressure

    NASA Astrophysics Data System (ADS)

    Puusepp, Marit; Kangur, Liina; Freiberg, Arvi

    2015-04-01

    The light-harvesting complex 1 (LH1) from Rhodobacter sphaeroides is an excellent model system for investigating the stability of oligomeric membrane proteins under high hydrostatic pressure. The currently investigated LH1 forms a 16-meric ring structure of B825 subunits. B825 is a heterodimer of transmembrane α- and β-polypeptide chains, which non-covalently binds two bacteriochlorophyll a molecules. These pigment molecules were used as intrinsic spectroscopic sensors to follow the dissociation reaction. Our results demonstrate that the LH1 dissociates into B825 subunits through an intermediary tetrameric unit B845. The dissociation mechanism depends on pressure. At ∼200-500 MPa the dissociation corresponds to a pseudo-first-order reaction, characterised by the apparent reaction rate at atmospheric pressure k0 = 3.10-5 s-1, activation volume ΔV‡ = -4 mL/mol, and free energy of activation ΔG‡ = 26 kJ/mol. Below 200 MPa and above 500 MPa, the reaction is more complex, including further dissociation of B825 into monomers B777. This paper was presented at the LIIth European High Pressure Research Group (EHPRG 52) Meeting in Lyon (France), 7-12 September 2014.

  5. Fluctuating Two-State Light Harvesting in a Photosynthetic Membrane

    SciTech Connect

    Pan, Duohai; Hu, Dehong; Liu, Ruchuan; Zeng, Xiaohua; Kaplan, Samuel; Lu, H. Peter

    2007-06-28

    How light is converted into chemical energy in a natural photosynthetic system is of great interest in energy sciences. Using single-molecule and single-vesicle fluorescence spectroscopy and imaging, we have observed fluctuating inter-molecular protein energy transfers in the photosynthetic membranes of R. sphaeroides. Our results suggest that there are dynamic coupled and non-coupled states in the light-harvesting protein assembly.

  6. Determination of the aggregate size in detergent solution of the light-harvesting chlorophyll a/b-protein complex from chloroplast membranes

    PubMed Central

    Butler, P. J. G.; Kühlbrandt, W.

    1988-01-01

    The molecular mass of an oligomeric integral membrane protein, the light-harvesting chlorophyll a/b-protein complex from the photosynthetic membranes of chloroplasts, has been determined in detergent solution by analytical ultracentrifugation and measurement of the density increment at constant chemical potential of all diffusible solutes. The technique used eliminates any problems resulting from detergent binding to the protein, is independent of the particular detergent used (in this case the nonionic n-octyl β-D-glucopyranoside), and gives the apparent weight-average molecular mass at different protein concentrations, allowing extrapolation to zero concentration. It means that the solutions of the complex must be brought to dialysis equilibrium with the solvent detergent solution and also requires a reliable method for measuring the protein concentration, for which amino acid analysis was used. The detergent-solubilized complex was a trimer that dissociated into monomers and dimers at low protein concentration. The accurate concentration determinations also allowed the molar chlorophyll-to-protein ratio to be measured as 15, corresponding to 8 chlorophyll a and 7 chlorophyll b molecules. PMID:16593931

  7. Complexation of copper and zinc ions with proteins of a light-harvesting complex (LHC-II) of chloroplast thylakoid membranes studied by FT-IR spectroscopy

    NASA Astrophysics Data System (ADS)

    Tajmir-Riahi, H. A.; Ahmed, A.

    1993-08-01

    The interaction of Zn(II) and Cu(II) ions with the light-harvesting proteins (LHC-II) of chloroplast thylakoid membranes was studied in aqueous solution with metal ion concentrations of 0.01 to 20mM, using Fourier transform-infrared (FT-IR) spectroscopy. Analyses of the metal ion binding mode and protein conformational variations were carried out and correlations between spectral changes and metal—protein complexation were established. Infrared difference spectroscopic results revealed the presence of a strong metal—protein interaction at high metal ion concentrations, while at low concentrations complexation was negligible. The binding of Zn and Cu ions was found to be with the protein carbonyl groups at low metal ion concentrations, whereas CO and CN groups were the main coordination sites at higher concentrations. A major conformational variation from α-helix to β-sheet and turn structures was observed in the presence of a concentrated metal ion solution.

  8. A Diatom Light-Harvesting Pigment-Protein Complex 1

    PubMed Central

    Friedman, Alan L.; Alberte, Randall S.

    1984-01-01

    A light-harvesting pigment-protein complex was isolated from the diatom Phaeodactylum tricornutum using the zwitterionic detergent CHAPS (3-[3-cholamidopropyl)dimethylammonio]-1-propanesulfonate). Detergent-solubilized membranes were fractionated by sucrose density gradient centrifugation into three components. The medium density fraction contained chlorophyll a, chlorophyll c, and fucoxanthin. This fraction was purified by DEAE-ion exchange chromatography, and contained chlorophyll a, chlorophyll c, and fucoxanthin in a molar ratio of 2.4:1.0:4.8. Fluorescence emission and excitation spectra of the isolated complex demonstrated that light energy absorbed by chlorophyll c and fucoxanthin was coupled to chlorophyll a fluorescence. Upon denaturation, the apoprotein yielded a polypeptide doublet at 17.5 to 18.0 kilodaltons which accounted for 30 to 40% of the toal membrane protein. These findings indicate that this pigment-protein complex is a major component of the diatom photosynthetic lammellae. The quantitative amino acid composition of the apoprotein was very similar to those reported for other membrane-bound pigment-protein complexes. Based on the protein to chlorophyll a ratio of 7700 grams protein per mole chlorophyll a for the complex, each apoprotein molecule contains, to the nearest integer, two chlorophyll a, one chlorophyll c, and five fucoxanthin molecules. Polyclonal antibodies raised against the 17.5 to 18.0 kilodaltons apoprotein showed a monospecific reaction with only the 17.5 to 18.0 protein zone from denatured P. tricornutum membranes as well as to the nondenatured pigment-protein complex. It appears that this complex is common to other diatom species. Images Fig. 2 Fig. 3 PMID:16663869

  9. Interaction of Mg(II), Ca(II) and Mn(II) ions with the light-harvesting proteins of chloroplast thylakoid membranes studied by FT-IR difference spectroscopy

    NASA Astrophysics Data System (ADS)

    Ahmed, A.; Tajmir-Riahi, H. A.

    1994-03-01

    The interaction of Mg(II), Ca(II) and Mn(II) ions with the light-harvesting (LHC-II) proteins of chloroplast thylakoid membranes was investigated in aqueous solution at different metal ion concentrations (0.01-20 mM), using Fourier transform-infrared (FT-IR) difference spectroscopy. The infrared difference spectroscopic results for the amide I and amide II regions (1800-1500 cm -1) have shown a strong metal—protein interaction at high metal ion concentrations (5-20 mM), whereas at very low concentrations (0.01-1 mM) the metal cation binding is negligible. The metal ion binding is mainly via the protein carbonyl group at low cation concentration, whereas metal ion coordination to the protein CO and CN groups were observed at higher cation concentrations. The Mn—tyrosine binding was also observed at high metal ion concentrations. Major conformational changes from α-helix (48% in uncomplexed protein) to β-sheet and turn structures were observed in the presence of these metal cations at high concentrations (10-20 mM).

  10. A thioredoxin-like/β-propeller protein maintains the efficiency of light harvesting in Arabidopsis.

    PubMed

    Brooks, Matthew D; Sylak-Glassman, Emily J; Fleming, Graham R; Niyogi, Krishna K

    2013-07-16

    The light-harvesting complexes of plants have evolved the ability to switch between efficient light harvesting and quenching forms to optimize photosynthesis in response to the environment. Several distinct mechanisms, collectively termed "nonphotochemical quenching" (NPQ), provide flexibility in this response. Here we report the isolation and characterization of a mutant, suppressor of quenching 1 (soq1), that has high NPQ even in the absence of photosystem II subunit S (PsbS), a protein that is necessary for the rapidly reversible component of NPQ. The formation of NPQ in soq1 was light intensity-dependent, and it exhibited slow relaxation kinetics and other characteristics that distinguish it from known NPQ components. Treatment with chemical inhibitors or an uncoupler, as well as crosses to mutants known to affect other NPQ components, showed that the NPQ in soq1 does not require a transthylakoid pH gradient, zeaxanthin formation, or the phosphorylation of light-harvesting complexes, and it appears to be unrelated to the photosystem II damage-and-repair cycle. Measurements of pigments and chlorophyll fluorescence lifetimes indicated that the additional NPQ in soq1 is the result of a decrease in chlorophyll excited-state lifetime and not pigment bleaching. The SOQ1 gene was isolated by map-based cloning, and it encodes a previously uncharacterized thylakoid membrane protein with thioredoxin-like and β-propeller domains located in the lumen and a haloacid-dehalogenase domain exposed to the chloroplast stroma. We propose that the role of SOQ1 is to prevent formation of a slowly reversible form of antenna quenching, thereby maintaining the efficiency of light harvesting.

  11. Light-harvesting chlorophyll a/b-binding protein inserted into isolated thylakoids binds pigments and is assembled into trimeric light-harvesting complex.

    PubMed Central

    Kuttkat, A; Grimm, R; Paulsen, H

    1995-01-01

    The light-harvesting chlorophyll a/b-binding protein (LHCP) is largely protected against protease (except for about 1 kD on the N terminus) in the thylakoid membrane; this protease resistance is often used to assay successful insertion of LHCP into isolated thylakoids in vitro. In this paper we show that this protease resistance is exhibited by trimeric light-harvesting complex of photosystem II (LHCII) but not by monomeric LHCII in which about 5 kD on the N terminus of LHCP are cleaved off by protease. When a mutant version of LHCP that is unable to trimerize in an in vitro reconstitution assay is inserted into isolated thylakoids, it gives rise to only the shorter protease digestion product indicative of monomeric LHCII. We conclude that more of the N-terminal domain of LHCP is shielded in trimeric than in monomeric LHCII and that this difference in protease sensitivity can be used to distinguish between LHCP assembled in LHCII monomers or trimers. The data presented prove that upon insertion of LHCP into isolated thylakoids at least part of the protein spontaneously binds pigments to form LHCII, which then is assembled in trimers. The dependence of the protease sensitivity of thylakoid-inserted LHCP on the oligomerization state of the newly formed LHCII justifies caution when using a protease assay to verify successful insertion of LHCP into the membrane. PMID:8539291

  12. Protein kinase that phosphorylates light-harvesting complex is autophosphorylated and is associated with photosystem II

    SciTech Connect

    Coughlan, S.J.; Hind, G.

    1987-10-06

    Thylakoid membranes were phosphorylated with (..gamma..-/sup 32/P)ATP and extracted with octyl glucoside and cholate. Among the radiolabeled phosphoproteins in the extract was a previously characterized protein kinase of 64-kDa apparent mass. The ability of this enzyme to undergo autophosphorylation in situ was used to monitor its distribution in the membrane. Fractionation studies showed that the kinase is confined to granal regions of the thylakoid, where it appears to be associated with the light-harvesting chlorophyll-protein complex of photosystem II. The kinetics of kinase autophosphorylation were investigated both in situ and in extracted, purified enzyme. In the membrane, autophosphorylation saturated within 20-30 min and was reversed with a half-time of 7-8 min upon removal of ATP or oxidative inactivation of the kinase; the accompanying dephosphorylation of light-harvesting complex was slower and kinetically complex. Fluoride (10 mM) inhibited these dephosphorylations. Autophosphorylation of the isolated kinase was independent of enzyme concentration, indicative of an intramolecular mechanism. A maximum of one serine residue per mole of kinase was esterified. Autophosphorylation was more rapid in the presence of histone IIIs, an exogenous substrate. Dephosphorylation of the isolated enzyme was not observed.

  13. A novel type of light-harvesting antenna protein of red algal origin in algae with secondary plastids

    PubMed Central

    2013-01-01

    Background Light, the driving force of photosynthesis, can be harmful when present in excess; therefore, any light harvesting system requires photoprotection. Members of the extended light-harvesting complex (LHC) protein superfamily are involved in light harvesting as well as in photoprotection and are found in the red and green plant lineages, with a complex distribution pattern of subfamilies in the different algal lineages. Results Here, we demonstrate that the recently discovered “red lineage chlorophyll a/b-binding-like proteins” (RedCAPs) form a monophyletic family within this protein superfamily. The occurrence of RedCAPs was found to be restricted to the red algal lineage, including red algae (with primary plastids) as well as cryptophytes, haptophytes and heterokontophytes (with secondary plastids of red algal origin). Expression of a full-length RedCAP:GFP fusion construct in the diatom Phaeodactylum tricornutum confirmed the predicted plastid localisation of RedCAPs. Furthermore, we observed that similarly to the fucoxanthin chlorophyll a/c-binding light-harvesting antenna proteins also RedCAP transcripts in diatoms were regulated in a diurnal way at standard light conditions and strongly repressed at high light intensities. Conclusions The absence of RedCAPs from the green lineage implies that RedCAPs evolved in the red lineage after separation from the the green lineage. During the evolution of secondary plastids, RedCAP genes therefore must have been transferred from the nucleus of the endocytobiotic alga to the nucleus of the host cell, a process that involved complementation with pre-sequences allowing import of the gene product into the secondary plastid bound by four membranes. Based on light-dependent transcription and on localisation data, we propose that RedCAPs might participate in the light (intensity and quality)-dependent structural or functional reorganisation of the light-harvesting antennae of the photosystems upon dark to light

  14. Molecular adaptation of photoprotection: triplet states in light-harvesting proteins.

    PubMed

    Gall, Andrew; Berera, Rudi; Alexandre, Maxime T A; Pascal, Andrew A; Bordes, Luc; Mendes-Pinto, Maria M; Andrianambinintsoa, Sandra; Stoitchkova, Katerina V; Marin, Alessandro; Valkunas, Leonas; Horton, Peter; Kennis, John T M; van Grondelle, Rienk; Ruban, Alexander; Robert, Bruno

    2011-08-17

    The photosynthetic light-harvesting systems of purple bacteria and plants both utilize specific carotenoids as quenchers of the harmful (bacterio)chlorophyll triplet states via triplet-triplet energy transfer. Here, we explore how the binding of carotenoids to the different types of light-harvesting proteins found in plants and purple bacteria provides adaptation in this vital photoprotective function. We show that the creation of the carotenoid triplet states in the light-harvesting complexes may occur without detectable conformational changes, in contrast to that found for carotenoids in solution. However, in plant light-harvesting complexes, the triplet wavefunction is shared between the carotenoids and their adjacent chlorophylls. This is not observed for the antenna proteins of purple bacteria, where the triplet is virtually fully located on the carotenoid molecule. These results explain the faster triplet-triplet transfer times in plant light-harvesting complexes. We show that this molecular mechanism, which spreads the location of the triplet wavefunction through the pigments of plant light-harvesting complexes, results in the absence of any detectable chlorophyll triplet in these complexes upon excitation, and we propose that it emerged as a photoprotective adaptation during the evolution of oxygenic photosynthesis.

  15. Biogenesis of a photosystem I light-harvesting complex. Evidence for a membrane intermediate.

    PubMed Central

    Adam, Z; Hoffman, N E

    1993-01-01

    CAB-7p is a chlorophyll a/b binding protein of photosystem I (PSI). It is found in light-harvesting complex I 680 (LHCI-680), one of the chlorophyll complexes produced by detergent solubilization of PSI. Two types of evidence are presented to indicate that assembly of CAB-7p into PSI proceeds through a membrane intermediate. First, when CAB-7p is briefly imported into chloroplasts or isolated thylakoids, we initially observe a fast-migrating membrane form of CAB-7p that is subsequently converted into PSI. The conversion of the fast-migrating form into PSI does not require stroma or ATP. Second, trypsin treatment of thylakoids containing radiolabeled CAB-7p indicates that there are at least two membrane forms of the mature 23-kD protein. The predominant form is completely resistant to proteolysis; a second form of the protein is cleaved by trypsin into 12- and 7-kD polypeptides. We interpret this to mean that the intermediate is a cleavable form that becomes protease resistant during assembly. This notion is supported by the observation that CAB-7p in LHCI-680 is largely cleaved by trypsin into 12- and 7-kD polypeptides, whereas CAB-7p in isolated PSI particles is trypsin resistant. In vitro, we generated a mutant form of CAB-7p, CAB-7/BgI2p, that was able to integrate into thylakoid membranes but was unable to assemble into PSI. The membrane form of CAB-7/BgI2p, like LHCI-680, was predominantly cleaved by trypsin into 12- and 7-kD fragments. We suggest that the mutant protein is arrested at an intermediate stage in the assembly pathway of PSI. Based on its mobility in nondenaturing gels and its susceptibility to protease cleavage, we suggest that the intermediate form is LHCI-680. We propose the following distinct stages in the biogenesis of LHCI: (a) apoprotein is integrated into the thylakoid, (b) chlorophyll is rapidly bound to apoprotein forming LHCI-680, and (c) LHCI-680 assembles into the native PSI complex. PMID:8108505

  16. Light-harvesting Complexes (LHCs) Cluster Spontaneously in Membrane Environment Leading to Shortening of Their Excited State Lifetimes.

    PubMed

    Natali, Alberto; Gruber, J Michael; Dietzel, Lars; Stuart, Marc C A; van Grondelle, Rienk; Croce, Roberta

    2016-08-01

    The light reactions of photosynthesis, which include light-harvesting and charge separation, take place in the amphiphilic environment of the thylakoid membrane. The light-harvesting complex II (LHCII) is the main responsible for light absorption in plants and green algae and is involved in photoprotective mechanisms that regulate the amount of excited states in the membrane. The dual function of LHCII has been extensively studied in detergent micelles, but recent results have indicated that the properties of this complex differ in a lipid environment. In this work we checked these suggestions by studying LHCII in liposomes. By combining bulk and single molecule measurements, we monitored the fluorescence characteristics of liposomes containing single complexes up to densely packed proteoliposomes. We show that the natural lipid environment per se does not alter the properties of LHCII, which for single complexes remain very similar to that in detergent. However, we show that LHCII has the strong tendency to cluster in the membrane and that protein interactions and the extent of crowding modulate the lifetimes of the excited state in the membrane. Finally, the presence of LHCII monomers at low concentrations of complexes per liposome is discussed. PMID:27252376

  17. Allelic variations of a light harvesting chlorophyll A/B protein gene (Lhcb1) associated with agronomic traits in Barley

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Light-harvesting chlorophyll a/b-binding protein (LHCP) is one of the most abundant chloroplast proteins in plants. Its main function is to collect and transfer light energy to photosynthetic reaction centers. However, the roles of different LHCPs in light-harvesting antenna systems remain obscure. ...

  18. Plasmon enhanced light harvesting: multiscale modeling of the FMO protein coupled with gold nanoparticles.

    PubMed

    Andreussi, Oliviero; Caprasecca, Stefano; Cupellini, Lorenzo; Guarnetti-Prandi, Ingrid; Guido, Ciro A; Jurinovich, Sandro; Viani, Lucas; Mennucci, Benedetta

    2015-05-28

    Plasmonic systems, such as metal nanoparticles, are becoming increasingly important in spectroscopies and devices because of their ability to enhance, even by several orders of magnitude, the photophysical properties of neighboring systems. In particular, it has been shown both theoretically and experimentally that combining nanoplasmonic devices with natural light-harvesting proteins substantially increases the fluorescence and absorption properties of the system. This kind of biohybrid device can have important applications in the characterization and design of efficient light-harvesting systems. In the present work, the FMO light-harvesting protein was combined with gold nanoparticles of different sizes, and its photophysical properties were characterized using a multiscale quantum-mechanical classical-polarizable and continuum model (QM/MMPol/PCM). By optimal tuning of the plasmon resonance of the metal nanoparticles, fluorescence enhancements of up to 2 orders of magnitude were observed. Orientation effects were found to be crucial: amplifications by factors of up to 300 were observed for the absorption process, while the radiative decay of the emitting state increased at most by a factor of 10, mostly as a result of poor alignment of the emitting state with the considered metal aggregates. Despite being a limiting factor for high-fluorescence-enhancement devices, the strong orientation dependence may represent an important feature of the natural light-harvesting system that could allow selective enhancement of a specific excited state of the complex.

  19. Excitation energy transfer between Light-harvesting complex II and Photosystem I in reconstituted membranes.

    PubMed

    Akhtar, Parveen; Lingvay, Mónika; Kiss, Teréz; Deák, Róbert; Bóta, Attila; Ughy, Bettina; Garab, Győző; Lambrev, Petar H

    2016-04-01

    Light-harvesting complex II (LHCII), the major peripheral antenna of Photosystem II in plants, participates in several concerted mechanisms for regulation of the excitation energy and electron fluxes in thylakoid membranes. In part, these include interaction of LHCII with Photosystem I (PSI) enhancing the latter's absorption cross-section - for example in the well-known state 1 - state 2 transitions or as a long-term acclimation to high light. In this work we examined the capability of LHCII to deliver excitations to PSI in reconstituted membranes in vitro. Proteoliposomes with native plant thylakoid membrane lipids and different stoichiometric ratios of LHCII:PSI were reconstituted and studied by steady-state and time-resolved fluorescence spectroscopy. Fluorescence emission from LHCII was strongly decreased in PSI-LHCII membranes due to trapping of excitations by PSI. Kinetic modelling of the time-resolved fluorescence data revealed the existence of separate pools of LHCII distinguished by the time scale of energy transfer. A strongly coupled pool, equivalent to one LHCII trimer per PSI, transferred excitations to PSI with near-unity efficiency on a time scale of less than 10ps but extra LHCIIs also contributed significantly to the effective antenna size of PSI, which could be increased by up to 47% in membranes containing 3 LHCII trimers per PSI. The results demonstrate a remarkable competence of LHCII to increase the absorption cross-section of PSI, given the opportunity that the two types of complexes interact in the membrane.

  20. Partial purification of a spinach thylakoid protein kinase that can phosphorylate light-harvesting chlorophyll a/b proteins

    SciTech Connect

    Clark, R.D.; Hind, G.; Bennett, J.

    1985-01-01

    Protein phosphorylation in plant tissues is particularly marked in chloroplasts, protein kinase activity being associated with the outer envelope, the soluble stromal fraction, and the thylakoid membrane. Furthermore, thylakoid-bound activity probably includes several distinct kinases, as suggested by studies of divalent cation specificity and thermal lability carried out with intact thylakoids and by subfractionation of solubilized membranes. Illumination of thylakoids, particularly with red light, promotes the rapid and extensive phosphorylation of the light-harvesting chlorophyll a/b complex (LHCII) on a threonine residue near the amino terminus of the protein. This phosphorylation is thought to be involved in regulating the distribution of absorbed quanta between photosystems II and I and is modulated by the redox state of the thylakoid plastoquinone pool. Neither of the thylakoid kinases reported to date was capable of phosphorylating purified LHCII in vitro or of incorporating phosphate into threonyl residues of exogenous substrates, that some LHCII phosphorylation was catalyzed by a preliminary fraction led workers to suggest that at least one other kinase remained to be isolated. Here, the authors report the solubilization and partial purification of a protein kinase from spinach thylakoids that is capable of phosphorylating LHCII in vitro, and they show that the specific site of phosphorylation is very nearly the same as, if not identical with, the site phosphorylated in organello.

  1. Development of Scaffolds for Light Harvesting and Photocatalysis from the Coat Protein of Tobacco Mosaic Virus

    NASA Astrophysics Data System (ADS)

    Dedeo, Michel Toussaint

    The utility of a previously developed TMV-based light harvesting system has been dramatically expanded through the introduction of reactive handles for the site-specific modification of the interior and exterior surfaces. Further experiments to reengineer the coat protein have produced structures with unique, unexpected, and useful assembly properties that complement the newly available surface modifications. Energy transfer from chromophores in the RNA channel of self-assembled TMV structures to the exterior was made possible by conjugation of acceptor dyes and porphyrins to the N-terminus. By repositioning the N-terminus to the pore through circular permutation, this process was repeated to create structures that mimic the light harvesting 1 complex of photosynthetic bacteria. To study and improve upon natural photosynthesis, closely packed chromophore arrays and gold nanoparticles were tethered to the pore of stabilized TMV disks through introduction of a uniquely reactive lysine. Finally, a dimeric TMV coat protein was produced to control the distribution and arrangement of synthetic groups with synergistic activity.

  2. A Light Switch Based on Protein S-Nitrosylation Fine-Tunes Photosynthetic Light Harvesting in Chlamydomonas.

    PubMed

    Berger, Hanna; De Mia, Marcello; Morisse, Samuel; Marchand, Christophe H; Lemaire, Stéphane D; Wobbe, Lutz; Kruse, Olaf

    2016-06-01

    Photosynthetic eukaryotes are challenged by a fluctuating light supply, demanding for a modulated expression of nucleus-encoded light-harvesting proteins associated with photosystem II (LHCII) to adjust light-harvesting capacity to the prevailing light conditions. Here, we provide clear evidence for a regulatory circuit that controls cytosolic LHCII translation in response to light quantity changes. In the green unicellular alga Chlamydomonas reinhardtii, the cytosolic RNA-binding protein NAB1 represses translation of certain LHCII isoform mRNAs. Specific nitrosylation of Cys-226 decreases NAB1 activity and could be demonstrated in vitro and in vivo. The less active, nitrosylated form of NAB1 is found in cells acclimated to limiting light supply, which permits accumulation of light-harvesting proteins and efficient light capture. In contrast, elevated light supply causes its denitrosylation, thereby activating the repression of light-harvesting protein synthesis, which is needed to control excitation pressure at photosystem II. Denitrosylation of recombinant NAB1 is efficiently performed by the cytosolic thioredoxin system in vitro. To our knowledge, NAB1 is the first example of stimulus-induced denitrosylation in the context of photosynthetic acclimation. By identifying this novel redox cross-talk pathway between chloroplast and cytosol, we add a new key element required for drawing a precise blue print of the regulatory network of light harvesting. PMID:27208221

  3. Vibronic resonances facilitate excited-state coherence in light-harvesting proteins at room temperature.

    PubMed

    Novelli, Fabio; Nazir, Ahsan; Richards, Gethin H; Roozbeh, Ashkan; Wilk, Krystyna E; Curmi, Paul M G; Davis, Jeffrey A

    2015-11-19

    Until recently it was believed that photosynthesis, a fundamental process for life on earth, could be fully understood with semiclassical models. However, puzzling quantum phenomena have been observed in several photosynthetic pigment-protein complexes, prompting questions regarding the nature and role of these effects. Recent attention has focused on discrete vibrational modes that are resonant or quasi-resonant with excitonic energy splittings and strongly coupled to these excitonic states. Here we unambiguously identify excited state coherent superpositions in photosynthetic light-harvesting complexes using a new experimental approach. Decoherence on the time scale of the excited state lifetime allows low energy (56 cm(-1)) oscillations on the signal intensity to be observed. In conjunction with an appropriate model, these oscillations provide clear and direct experimental evidence that the persistent coherences observed originate from quantum superpositions among vibronic excited states. PMID:26528956

  4. Light-Harvesting Complex Stress-Related Proteins Catalyze Excess Energy Dissipation in Both Photosystems of Physcomitrella patens.

    PubMed

    Pinnola, Alberta; Cazzaniga, Stefano; Alboresi, Alessandro; Nevo, Reinat; Levin-Zaidman, Smadar; Reich, Ziv; Bassi, Roberto

    2015-11-01

    Two LHC-like proteins, Photosystem II Subunit S (PSBS) and Light-Harvesting Complex Stress-Related (LHCSR), are essential for triggering excess energy dissipation in chloroplasts of vascular plants and green algae, respectively. The mechanism of quenching was studied in Physcomitrella patens, an early divergent streptophyta (including green algae and land plants) in which both proteins are active. PSBS was localized in grana together with photosystem II (PSII), but LHCSR was located mainly in stroma-exposed membranes together with photosystem I (PSI), and its distribution did not change upon high-light treatment. The quenched conformation can be preserved by rapidly freezing the high-light-treated tissues in liquid nitrogen. When using green fluorescent protein as an internal standard, 77K fluorescence emission spectra on isolated chloroplasts allowed for independent assessment of PSI and PSII fluorescence yield. Results showed that both photosystems underwent quenching upon high-light treatment in the wild type in contrast to mutants depleted of LHCSR, which lacked PSI quenching. Due to the contribution of LHCII, P. patens had a PSI antenna size twice as large with respect to higher plants. Thus, LHCII, which is highly abundant in stroma membranes, appears to be the target of quenching by LHCSR.

  5. The light-harvesting chlorophyll a/b binding proteins Lhcb1 and Lhcb2 play complementary roles during state transitions in Arabidopsis.

    PubMed

    Pietrzykowska, Malgorzata; Suorsa, Marjaana; Semchonok, Dmitry A; Tikkanen, Mikko; Boekema, Egbert J; Aro, Eva-Mari; Jansson, Stefan

    2014-09-01

    Photosynthetic light harvesting in plants is regulated by phosphorylation-driven state transitions: functional redistributions of the major trimeric light-harvesting complex II (LHCII) to balance the relative excitation of photosystem I and photosystem II. State transitions are driven by reversible LHCII phosphorylation by the STN7 kinase and PPH1/TAP38 phosphatase. LHCII trimers are composed of Lhcb1, Lhcb2, and Lhcb3 proteins in various trimeric configurations. Here, we show that despite their nearly identical amino acid composition, the functional roles of Lhcb1 and Lhcb2 are different but complementary. Arabidopsis thaliana plants lacking only Lhcb2 contain thylakoid protein complexes similar to wild-type plants, where Lhcb2 has been replaced by Lhcb1. However, these do not perform state transitions, so phosphorylation of Lhcb2 seems to be a critical step. In contrast, plants lacking Lhcb1 had a more profound antenna remodeling due to a decrease in the amount of LHCII trimers influencing thylakoid membrane structure and, more indirectly, state transitions. Although state transitions are also found in green algae, the detailed architecture of the extant seed plant light-harvesting antenna can now be dated back to a time after the divergence of the bryophyte and spermatophyte lineages, but before the split of the angiosperm and gymnosperm lineages more than 300 million years ago. PMID:25194026

  6. The Light-Harvesting Chlorophyll a/b Binding Proteins Lhcb1 and Lhcb2 Play Complementary Roles during State Transitions in Arabidopsis[C][W][OPEN

    PubMed Central

    Pietrzykowska, Malgorzata; Suorsa, Marjaana; Semchonok, Dmitry A.; Tikkanen, Mikko; Boekema, Egbert J.; Aro, Eva-Mari

    2014-01-01

    Photosynthetic light harvesting in plants is regulated by phosphorylation-driven state transitions: functional redistributions of the major trimeric light-harvesting complex II (LHCII) to balance the relative excitation of photosystem I and photosystem II. State transitions are driven by reversible LHCII phosphorylation by the STN7 kinase and PPH1/TAP38 phosphatase. LHCII trimers are composed of Lhcb1, Lhcb2, and Lhcb3 proteins in various trimeric configurations. Here, we show that despite their nearly identical amino acid composition, the functional roles of Lhcb1 and Lhcb2 are different but complementary. Arabidopsis thaliana plants lacking only Lhcb2 contain thylakoid protein complexes similar to wild-type plants, where Lhcb2 has been replaced by Lhcb1. However, these do not perform state transitions, so phosphorylation of Lhcb2 seems to be a critical step. In contrast, plants lacking Lhcb1 had a more profound antenna remodeling due to a decrease in the amount of LHCII trimers influencing thylakoid membrane structure and, more indirectly, state transitions. Although state transitions are also found in green algae, the detailed architecture of the extant seed plant light-harvesting antenna can now be dated back to a time after the divergence of the bryophyte and spermatophyte lineages, but before the split of the angiosperm and gymnosperm lineages more than 300 million years ago. PMID:25194026

  7. Light harvesting proteins for solar fuel generation in bioengineered photoelectrochemical cells.

    PubMed

    Ihssen, Julian; Braun, Artur; Faccio, Greta; Gajda-Schrantz, Krisztina; Thöny-Meyer, Linda

    2014-01-01

    The sun is the primary energy source of our planet and potentially can supply all societies with more than just their basic energy needs. Demand of electric energy can be satisfied with photovoltaics, however the global demand for fuels is even higher. The direct way to produce the solar fuel hydrogen is by water splitting in photoelectrochemical (PEC) cells, an artificial mimic of photosynthesis. There is currently strong resurging interest for solar fuels produced by PEC cells, but some fundamental technological problems need to be solved to make PEC water splitting an economic, competitive alternative. One of the problems is to provide a low cost, high performing water oxidizing and oxygen evolving photoanode in an environmentally benign setting. Hematite, α-Fe2O3, satisfies many requirements for a good PEC photoanode, but its efficiency is insufficient in its pristine form. A promising strategy for enhancing photocurrent density takes advantage of photosynthetic proteins. In this paper we give an overview of how electrode surfaces in general and hematite photoanodes in particular can be functionalized with light harvesting proteins. Specifically, we demonstrate how low-cost biomaterials such as cyanobacterial phycocyanin and enzymatically produced melanin increase the overall performance of virtually no-cost metal oxide photoanodes in a PEC system. The implementation of biomaterials changes the overall nature of the photoanode assembly in a way that aggressive alkaline electrolytes such as concentrated KOH are not required anymore. Rather, a more environmentally benign and pH neutral electrolyte can be used. PMID:24678669

  8. Light harvesting proteins for solar fuel generation in bioengineered photoelectrochemical cells.

    PubMed

    Ihssen, Julian; Braun, Artur; Faccio, Greta; Gajda-Schrantz, Krisztina; Thöny-Meyer, Linda

    2014-01-01

    The sun is the primary energy source of our planet and potentially can supply all societies with more than just their basic energy needs. Demand of electric energy can be satisfied with photovoltaics, however the global demand for fuels is even higher. The direct way to produce the solar fuel hydrogen is by water splitting in photoelectrochemical (PEC) cells, an artificial mimic of photosynthesis. There is currently strong resurging interest for solar fuels produced by PEC cells, but some fundamental technological problems need to be solved to make PEC water splitting an economic, competitive alternative. One of the problems is to provide a low cost, high performing water oxidizing and oxygen evolving photoanode in an environmentally benign setting. Hematite, α-Fe2O3, satisfies many requirements for a good PEC photoanode, but its efficiency is insufficient in its pristine form. A promising strategy for enhancing photocurrent density takes advantage of photosynthetic proteins. In this paper we give an overview of how electrode surfaces in general and hematite photoanodes in particular can be functionalized with light harvesting proteins. Specifically, we demonstrate how low-cost biomaterials such as cyanobacterial phycocyanin and enzymatically produced melanin increase the overall performance of virtually no-cost metal oxide photoanodes in a PEC system. The implementation of biomaterials changes the overall nature of the photoanode assembly in a way that aggressive alkaline electrolytes such as concentrated KOH are not required anymore. Rather, a more environmentally benign and pH neutral electrolyte can be used.

  9. Third order nonlinear optical properties of stacked bacteriochlorophylls in bacterial photosynthetic light-harvesting proteins

    SciTech Connect

    Chen, L.X.; Laible, P.D.; Spano, F.C.; Manas, E.S.

    1997-09-01

    Enhancement of the nonresonant second order molecular hyperpolarizabilities {gamma} were observed in stacked macrocyclic molecular systems, previously in a {micro}-oxo silicon phthalocyanine (SiPcO) monomer, dimer and trimer series, and now in bacteriochlorophyll a (BChla) arrays of light harvesting (LH) proteins. Compared to monomeric BChla in a tetrahydrofuran (THF) solution, the <{gamma}> for each macrocycle was enhanced in naturally occurring stacked macrocyclic molecular systems in the bacterial photosynthetic LH proteins where BChla`s are arranged in tilted face-to-face arrays. In addition, the {gamma} enhancement is more significant in B875 of LH1 than in B850 in LH2. Theoretical modeling of the nonresonant {gamma} enhancement using simplified molecular orbitals for model SiPcO indicated that the energy level of the two photon state is crucial to the {gamma} enhancement when a two photon process is involved, whereas the charge transfer between the monomers is largely responsible when one photon near resonant process is involved. The calculated results can be extended to {gamma} enhancement in B875 and B850 arrays, suggesting that BChla in B875 are more strongly coupled than in B850. In addition, a 50--160 fold increase in <{gamma}> for the S{sub 1} excited state of relative to S{sub 0} of bacteriochlorophyll in vivo was observed which provides an alternative method for probing excited state dynamics and a potential application for molecular switching.

  10. Light Harvesting Proteins for Solar Fuel Generation in Bioengineered Photoelectrochemical Cells

    PubMed Central

    Ihssen, Julian; Braun, Artur; Faccio, Greta; Gajda-Schrantz, Krisztina; Thöny-Meyer, Linda

    2014-01-01

    The sun is the primary energy source of our planet and potentially can supply all societies with more than just their basic energy needs. Demand of electric energy can be satisfied with photovoltaics, however the global demand for fuels is even higher. The direct way to produce the solar fuel hydrogen is by water splitting in photoelectrochemical (PEC) cells, an artificial mimic of photosynthesis. There is currently strong resurging interest for solar fuels produced by PEC cells, but some fundamental technological problems need to be solved to make PEC water splitting an economic, competitive alternative. One of the problems is to provide a low cost, high performing water oxidizing and oxygen evolving photoanode in an environmentally benign setting. Hematite, α-Fe2O3, satisfies many requirements for a good PEC photoanode, but its efficiency is insufficient in its pristine form. A promising strategy for enhancing photocurrent density takes advantage of photosynthetic proteins. In this paper we give an overview of how electrode surfaces in general and hematite photoanodes in particular can be functionalized with light harvesting proteins. Specifically, we demonstrate how low-cost biomaterials such as cyanobacterial phycocyanin and enzymatically produced melanin increase the overall performance of virtually no-cost metal oxide photoanodes in a PEC system. The implementation of biomaterials changes the overall nature of the photoanode assembly in a way that aggressive alkaline electrolytes such as concentrated KOH are not required anymore. Rather, a more environmentally benign and pH neutral electrolyte can be used. PMID:24678669

  11. Single-residue insertion switches the quaternary structure and exciton states of cryptophyte light-harvesting proteins

    PubMed Central

    Harrop, Stephen J.; Wilk, Krystyna E.; Dinshaw, Rayomond; Collini, Elisabetta; Mirkovic, Tihana; Teng, Chang Ying; Oblinsky, Daniel G.; Green, Beverley R.; Hoef-Emden, Kerstin; Hiller, Roger G.; Scholes, Gregory D.; Curmi, Paul M. G.

    2014-01-01

    Observation of coherent oscillations in the 2D electronic spectra (2D ES) of photosynthetic proteins has led researchers to ask whether nontrivial quantum phenomena are biologically significant. Coherent oscillations have been reported for the soluble light-harvesting phycobiliprotein (PBP) antenna isolated from cryptophyte algae. To probe the link between spectral properties and protein structure, we determined crystal structures of three PBP light-harvesting complexes isolated from different species. Each PBP is a dimer of αβ subunits in which the structure of the αβ monomer is conserved. However, we discovered two dramatically distinct quaternary conformations, one of which is specific to the genus Hemiselmis. Because of steric effects emerging from the insertion of a single amino acid, the two αβ monomers are rotated by ∼73° to an “open” configuration in contrast to the “closed” configuration of other cryptophyte PBPs. This structural change is significant for the light-harvesting function because it disrupts the strong excitonic coupling between two central chromophores in the closed form. The 2D ES show marked cross-peak oscillations assigned to electronic and vibrational coherences in the closed-form PC645. However, such features appear to be reduced, or perhaps absent, in the open structures. Thus cryptophytes have evolved a structural switch controlled by an amino acid insertion to modulate excitonic interactions and therefore the mechanisms used for light harvesting. PMID:24979784

  12. Single-residue insertion switches the quaternary structure and exciton states of cryptophyte light-harvesting proteins.

    PubMed

    Harrop, Stephen J; Wilk, Krystyna E; Dinshaw, Rayomond; Collini, Elisabetta; Mirkovic, Tihana; Teng, Chang Ying; Oblinsky, Daniel G; Green, Beverley R; Hoef-Emden, Kerstin; Hiller, Roger G; Scholes, Gregory D; Curmi, Paul M G

    2014-07-01

    Observation of coherent oscillations in the 2D electronic spectra (2D ES) of photosynthetic proteins has led researchers to ask whether nontrivial quantum phenomena are biologically significant. Coherent oscillations have been reported for the soluble light-harvesting phycobiliprotein (PBP) antenna isolated from cryptophyte algae. To probe the link between spectral properties and protein structure, we determined crystal structures of three PBP light-harvesting complexes isolated from different species. Each PBP is a dimer of αβ subunits in which the structure of the αβ monomer is conserved. However, we discovered two dramatically distinct quaternary conformations, one of which is specific to the genus Hemiselmis. Because of steric effects emerging from the insertion of a single amino acid, the two αβ monomers are rotated by ∼73° to an "open" configuration in contrast to the "closed" configuration of other cryptophyte PBPs. This structural change is significant for the light-harvesting function because it disrupts the strong excitonic coupling between two central chromophores in the closed form. The 2D ES show marked cross-peak oscillations assigned to electronic and vibrational coherences in the closed-form PC645. However, such features appear to be reduced, or perhaps absent, in the open structures. Thus cryptophytes have evolved a structural switch controlled by an amino acid insertion to modulate excitonic interactions and therefore the mechanisms used for light harvesting.

  13. Light harvesting in photosystem II.

    PubMed

    van Amerongen, Herbert; Croce, Roberta

    2013-10-01

    Water oxidation in photosynthesis takes place in photosystem II (PSII). This photosystem is built around a reaction center (RC) where sunlight-induced charge separation occurs. This RC consists of various polypeptides that bind only a few chromophores or pigments, next to several other cofactors. It can handle far more photons than the ones absorbed by its own pigments and therefore, additional excitations are provided by the surrounding light-harvesting complexes or antennae. The RC is located in the PSII core that also contains the inner light-harvesting complexes CP43 and CP47, harboring 13 and 16 chlorophyll pigments, respectively. The core is surrounded by outer light-harvesting complexes (Lhcs), together forming the so-called supercomplexes, at least in plants. These PSII supercomplexes are complemented by some "extra" Lhcs, but their exact location in the thylakoid membrane is unknown. The whole system consists of many subunits and appears to be modular, i.e., both its composition and organization depend on environmental conditions, especially on the quality and intensity of the light. In this review, we will provide a short overview of the relation between the structure and organization of pigment-protein complexes in PSII, ranging from individual complexes to entire membranes and experimental and theoretical results on excitation energy transfer and charge separation. It will become clear that time-resolved fluorescence data can provide invaluable information about the organization and functioning of thylakoid membranes. At the end, an overview will be given of unanswered questions that should be addressed in the near future.

  14. DNA-directed spatial assembly of photosynthetic light-harvesting proteins.

    PubMed

    Henry, Sarah L; Withers, Jamie M; Singh, Ishwar; Cooper, Jonathan M; Clark, Alasdair W; Burley, Glenn A; Cogdell, Richard J

    2016-01-28

    This manuscript describes the surface immobilization of a light-harvesting complex to prescribed locations directed by the sequence-selective recognition of duplex DNA. An engineered light-harvesting complex (RC-LH1) derived from Rhodopseudomonas (Rps.) palustris containing the zinc finger (ZF) domain zif268 was prepared. The zif268 domain directed the binding of zfRC-LH1 to target double-stranded DNA sequences both in solution and when immobilized on lithographically defined micro-patterns. Excitation energy transfer from the carotenoids to the bacteriochlorophyll pigments within zfRC-LH1 confirmed that the functional and structural integrity of the complex is retained after surface immobilization.

  15. The architecture and function of the light-harvesting apparatus of purple bacteria: from single molecules to in vivo membranes.

    PubMed

    Cogdell, Richard J; Gall, Andrew; Köhler, Jürgen

    2006-08-01

    This review describes the structures of the two major integral membrane pigment complexes, the RC-LH1 'core' and LH2 complexes, which together make up the light-harvesting system present in typical purple photosynthetic bacteria. The antenna complexes serve to absorb incident solar radiation and to transfer it to the reaction centres, where it is used to 'power' the photosynthetic redox reaction and ultimately leads to the synthesis of ATP. Our current understanding of the biosynthesis and assembly of the LH and RC complexes is described, with special emphasis on the roles of the newly described bacteriophytochromes. Using both the structural information and that obtained from a wide variety of biophysical techniques, the details of each of the different energy-transfer reactions that occur, between the absorption of a photon and the charge separation in the RC, are described. Special emphasis is given to show how the use of single-molecule spectroscopy has provided a more detailed understanding of the molecular mechanisms involved in the energy-transfer processes. We have tried, with the help of an Appendix, to make the details of the quantum mechanics that are required to appreciate these molecular mechanisms, accessible to mathematically illiterate biologists. The elegance of the purple bacterial light-harvesting system lies in the way in which it has cleverly exploited quantum mechanics.

  16. Light-Harvesting Complex Protein LHCBM9 Is Critical for Photosystem II Activity and Hydrogen Production in Chlamydomonas reinhardtii.

    PubMed

    Grewe, Sabrina; Ballottari, Matteo; Alcocer, Marcelo; D'Andrea, Cosimo; Blifernez-Klassen, Olga; Hankamer, Ben; Mussgnug, Jan H; Bassi, Roberto; Kruse, Olaf

    2014-04-01

    Photosynthetic organisms developed multiple strategies for balancing light-harvesting versus intracellular energy utilization to survive ever-changing environmental conditions. The light-harvesting complex (LHC) protein family is of paramount importance for this function and can form light-harvesting pigment protein complexes. In this work, we describe detailed analyses of the photosystem II (PSII) LHC protein LHCBM9 of the microalga Chlamydomonas reinhardtii in terms of expression kinetics, localization, and function. In contrast to most LHC members described before, LHCBM9 expression was determined to be very low during standard cell cultivation but strongly increased as a response to specific stress conditions, e.g., when nutrient availability was limited. LHCBM9 was localized as part of PSII supercomplexes but was not found in association with photosystem I complexes. Knockdown cell lines with 50 to 70% reduced amounts of LHCBM9 showed reduced photosynthetic activity upon illumination and severe perturbation of hydrogen production activity. Functional analysis, performed on isolated PSII supercomplexes and recombinant LHCBM9 proteins, demonstrated that presence of LHCBM9 resulted in faster chlorophyll fluorescence decay and reduced production of singlet oxygen, indicating upgraded photoprotection. We conclude that LHCBM9 has a special role within the family of LHCII proteins and serves an important protective function during stress conditions by promoting efficient light energy dissipation and stabilizing PSII supercomplexes. PMID:24706511

  17. The structure and function of bacterial light-harvesting complexes.

    PubMed

    Law, Christopher J; Roszak, Aleksander W; Southall, June; Gardiner, Alastair T; Isaacs, Neil W; Cogdell, Richard J

    2004-01-01

    The harvesting of solar radiation by purple photosynthetic bacteria is achieved by circular, integral membrane pigment-protein complexes. There are two main types of light-harvesting complex, termed LH2 and LH1, that function to absorb light energy and to transfer that energy rapidly and efficiently to the photochemical reaction centres where it is trapped. This mini-review describes our present understanding of the structure and function of the purple bacterial light-harvesting complexes.

  18. Incorporation of the chlorophyll d-binding light-harvesting protein from Acaryochloris marina and its localization within the photosynthetic apparatus of Synechocystis sp. PCC6803.

    PubMed

    Yang, Daping; Qing, Yang; Min, Chen

    2010-02-01

    The gene encoding a chlorophyll d-binding light-harvesting protein, pcbA from Acaryochloris marina (now called as accessory Chlorophyll Binding Protein CBPII) marked with a His-tag was transformed into the genome of Synechocystis PCC6803. Protein gel electrophoresis and western blotting confirmed that this foreign chlorophyll d-binding protein CBPII was expressed and integrated into the thylakoid membrane and bound with chlorophyll a, the only type of chlorophyll present in Synechocystis PCC 6803. Native electrophoresis suggested that CBPII interacts with photosystem II of Synechocystis PCC 6803. Surprisingly, spectral analyses showed that the phycobiliproteins were suppressed in the transformed Synechocystis pcbA(+), with a lower ratio of phycobilins to chlorophyll a. These results suggest that there are competitive interactions between the external antenna system of phycobiliproteins and the integral antenna system of chlorophyll-bound protein complexes.

  19. ARCHITECTURE OF A CHARGE-TRANSFER STATE REGULATING LIGHT HARVESTING IN A PLANT ANTENNA PROTEIN

    SciTech Connect

    Fleming, Graham; Ahn, Tae Kyu; Avenson, Thomas J.; Ballottari, Matteo; Cheng, Yuan-Chung; Niyogi, Krishna K.; Bassi, Roberto; Fleming, Graham R.

    2008-04-02

    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). In this work, we present 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 de-localized 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 chlorophylls-zeaxanthin charge-transfer state and switching on and off the charge-transfer quenching during qE.

  20. Architecture of a charge-transfer state regulating light harvesting in a plant antenna protein.

    PubMed

    Ahn, Tae Kyu; Avenson, Thomas J; Ballottari, Matteo; Cheng, Yuan-Chung; Niyogi, Krishna K; Bassi, Roberto; Fleming, Graham R

    2008-05-01

    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.

  1. Spectroscopic study of the light-harvesting protein C-phycocyanin associated with colorless linker peptides

    SciTech Connect

    Pizarro, Shelly A.

    2000-05-12

    The phycobilisome (PBS) light-harvesting antenna is composed of chromophore-containing biliproteins and 'colorless' linker peptides and is structurally designed to support unidirectional transfer of excitation energy from the periphery of the PBS to its core. The linker peptides have a unique role in this transfer process by modulating the spectral properties of the associated biliprotein. There is only one three-dimensional structure of a biliprotein/linker complex available to date (APC/LC7.8) and the mechanism of interaction between these two proteins remains unknown. This study brings together a detailed spectroscopic characterization of C-Phycocyanin (PC)-linker complexes (isolated from Synechococcus sp. PCC 7002) with proteomic analysis of the linker amino acid sequences to produce a model for biliprotein/linker interaction. The amino acid sequences of the rod linkers [LR8.9, LR32.3 and LRC28.5] were examined to identify evolutionarily conserved regions important to either the structure or function of this protein family. Although there is not one common homologous site among all the linkers, there are strong trends across each separate subset (LC, LR and LRC) and the N-terminal segments of both LR32.3 and LRC28.5 display multiple regions of similarity with other linkers. Predictions of the secondary structure of LR32.3 and LRC28.5, and comparison to the crystal structure of LC7.8, further narrowed the candidates for interaction sites with the PC chromophores. Measurements of the absorption, fluorescence, CD and excitation anisotropy of PC trimer, PC/LR32.3, and PC/LRC28.5, document the spectroscopic effect of each linker peptide on the PC chromophores at a series of temperatures (298 to 77 K). Because LR32.3 and LRC28.5 modulate the PC trimer spectral properties in distinct manners, it suggests different chromophore-interaction mechanisms for each linker. The low temperature absorbance spectrum of the PC trimer is consistent with an excitonic coupling

  2. Micelle-Induced Self-Assembling Protein Nanowires: Versatile Supramolecular Scaffolds for Designing the Light-Harvesting System.

    PubMed

    Sun, Hongcheng; Zhang, Xiyu; Miao, Lu; Zhao, Linlu; Luo, Quan; Xu, Jiayun; Liu, Junqiu

    2016-01-26

    Organic nanoparticle induced self-assembly of proteins with periodic nanostructures is a promising and burgeoning strategy to develop functional biomimetic nanomaterials. Cricoid proteins afford monodispersed and well-defined hollow centers, and can be used to multivalently interact with geometrically symmetric nanoparticles to form one-dimensional protein nanoarrays. Herein, we report that core-cross-linked micelles can direct cricoid stable protein one (SP1) to self-assembling nanowires through multiple electrostatic interactions. One micelle can act as an organic nanoparticle to interact with two central concaves of SP1 in an opposite orientation to form a sandwich structure, further controlling the assembly direction to supramolecular protein nanowires. The reported versatile supramolecular scaffolds can be optionally manipulated to develop multifunctional integrated or synergistic biomimetic nanomaterials. Artificial light-harvesting nanowires are further developed to mimic the energy transfer process of photosynthetic bacteria for their structural similarity, by means of labeling donor and acceptor chromophores to SP1 rings and spherical micelles, respectively. The absorbing energy can be transferred within the adjacent donors around the ring and shuttling the collected energy to the nearby acceptor chromophore. The artificial light-harvesting nanowires are designed by mimicking the structural characteristic of natural LH-2 complex, which are meaningful in exploring the photosynthesis process in vitro.

  3. Protein-Framed Multi-Porphyrin Micelles for a Hybrid Natural-Artificial Light-Harvesting Nanosystem.

    PubMed

    Liu, Yannan; Jin, Jiyang; Deng, Hongping; Li, Ke; Zheng, Yongli; Yu, Chunyang; Zhou, Yongfeng

    2016-07-01

    A micelle-like hybrid natural-artificial light-harvesting nanosystem was prepared through protein-framed electrostatic self-assembly of phycocyanin and a four-armed porphyrin star polymer. The nanosystem has a special structure of pomegranate-like unimolecular micelle aggregate with one phycocyanin acceptor in the center and multiple porphyrin donors in the shell. It can inhibit donor self-quenching effectively and display efficient transfer of excitation energy (about 80.1 %) in water. Furthermore, the number of donors contributing to a single acceptor could reach as high as about 179 in this nanosystem. PMID:27187799

  4. Tracking energy transfer between light harvesting complex 2 and 1 in photosynthetic membranes grown under high and low illumination.

    PubMed

    Lüer, Larry; Moulisová, Vladimíra; Henry, Sarah; Polli, Dario; Brotosudarmo, Tatas H P; Hoseinkhani, Sajjad; Brida, Daniele; Lanzani, Guglielmo; Cerullo, Giulio; Cogdell, Richard J

    2012-01-31

    Energy transfer (ET) between B850 and B875 molecules in light harvesting complexes LH2 and LH1/RC (reaction center) complexes has been investigated in membranes of Rhodopseudomonas palustris grown under high- and low-light conditions. In these bacteria, illumination intensity during growth strongly affects the type of LH2 complexes synthesized, their optical spectra, and their amount of energetic disorder. We used a specially built femtosecond spectrometer, combining tunable narrowband pump with broadband white-light probe pulses, together with an analytical method based on derivative spectroscopy for disentangling the congested transient absorption spectra of LH1 and LH2 complexes. This procedure allows real-time tracking of the forward (LH2 → LH1) and backward (LH2←LH1) ET processes and unambiguous determination of the corresponding rate constants. In low-light grown samples, we measured lower ET rates in both directions with respect to high-light ones, which is explained by reduced spectral overlap between B850 and B875 due to partial redistribution of oscillator strength into a higher energetic exciton transition. We find that the low-light adaptation in R. palustris leads to a reduced elementary backward ET rate, in accordance with the low probability of two simultaneous excitations reaching the same LH1/RC complex under weak illumination. Our study suggests that backward ET is not just an inevitable consequence of vectorial ET with small energetic offsets, but is in fact actively managed by photosynthetic bacteria.

  5. An atypical member of the light-harvesting complex stress-related protein family modulates diatom responses to light.

    PubMed

    Bailleul, Benjamin; Rogato, Alessandra; de Martino, Alessandra; Coesel, Sacha; Cardol, Pierre; Bowler, Chris; Falciatore, Angela; Finazzi, Giovanni

    2010-10-19

    Diatoms are prominent phytoplanktonic organisms that contribute around 40% of carbon assimilation in the oceans. They grow and perform optimally in variable environments, being able to cope with unpredictable changes in the amount and quality of light. The molecular mechanisms regulating diatom light responses are, however, still obscure. Using knockdown Phaeodactylum tricornutum transgenic lines, we reveal the key function of a member of the light-harvesting complex stress-related (LHCSR) protein family, denoted LHCX1, in modulation of excess light energy dissipation. In contrast to green algae, this gene is already maximally expressed in nonstressful light conditions and encodes a protein required for efficient light responses and growth. LHCX1 also influences natural variability in photoresponse, as evidenced in ecotypes isolated from different latitudes that display different LHCX1 protein levels. We conclude, therefore, that this gene plays a pivotal role in managing light responses in diatoms. PMID:20921421

  6. An atypical member of the light-harvesting complex stress-related protein family modulates diatom responses to light

    PubMed Central

    Bailleul, Benjamin; Rogato, Alessandra; de Martino, Alessandra; Coesel, Sacha; Cardol, Pierre; Bowler, Chris; Falciatore, Angela; Finazzi, Giovanni

    2010-01-01

    Diatoms are prominent phytoplanktonic organisms that contribute around 40% of carbon assimilation in the oceans. They grow and perform optimally in variable environments, being able to cope with unpredictable changes in the amount and quality of light. The molecular mechanisms regulating diatom light responses are, however, still obscure. Using knockdown Phaeodactylum tricornutum transgenic lines, we reveal the key function of a member of the light-harvesting complex stress-related (LHCSR) protein family, denoted LHCX1, in modulation of excess light energy dissipation. In contrast to green algae, this gene is already maximally expressed in nonstressful light conditions and encodes a protein required for efficient light responses and growth. LHCX1 also influences natural variability in photoresponse, as evidenced in ecotypes isolated from different latitudes that display different LHCX1 protein levels. We conclude, therefore, that this gene plays a pivotal role in managing light responses in diatoms. PMID:20921421

  7. Primary light-harvesting system: phycobilisomes and associated membranes. Progress report, January 1, 1981-December 31, 1981

    SciTech Connect

    Gantt, E.

    1981-01-01

    Phycobilisomes, serving as primary light harvesting complexes in cyanobacteria and red algae, were investigated. Structurally the phycobilisomes of both groups have the same fundamental phycobiliprotein arrangement. Allophycocyanin is in the center near the thylakoid. Stacked rods composed of phycocyanin, or phycocyanin-phycoerythrin radiate peripherally from the allophycocyanin core. Phycobilisomes of Nostoc sp. and Fremyella diplosiphon, after separation into separate allophycocyanin and phycoerythrin-phycocyanin fractions have been associated in vitro. Hybrid phycobilisomes, derived from mixtures of phycobiliprotein from these species were also obtained. The interaction is specific since reassociation was not obtained with phycobiliprotein complexes of some other algae. Phycobilisomes, whether native, or associated in vitro, were similar in their sedimentation, absorption, fluorescence excitation, fluorescence emission, and by electron microscopy. Furthermore, many of the colorless polypeptides were also highly similar between Nostoc and Fremyella. The similarity formed may reflect an evolutionary relationship between the two species. The polypeptide composition of Porphyridium cruentum phycobilisomes is the most complex of any thus far examined. The phycobiliprotein containing polypeptides comprised 84% of the total stainable protein, while the remaining were colorless. Most of the colorless polypeptides occurred in a pelletable fraction, which was enriched in allophycocyanin and phycocyanin, it is probable that some are involved in the linking of these phycobiliproteins.

  8. Tracking energy transfer between light harvesting complex 2 and 1 in photosynthetic membranes grown under high and low illumination

    PubMed Central

    Lüer, Larry; Moulisová, Vladimíra; Henry, Sarah; Polli, Dario; Brotosudarmo, Tatas H. P.; Hoseinkhani, Sajjad; Brida, Daniele; Lanzani, Guglielmo; Cerullo, Giulio; Cogdell, Richard J.

    2012-01-01

    Energy transfer (ET) between B850 and B875 molecules in light harvesting complexes LH2 and LH1/RC (reaction center) complexes has been investigated in membranes of Rhodopseudomonas palustris grown under high- and low-light conditions. In these bacteria, illumination intensity during growth strongly affects the type of LH2 complexes synthesized, their optical spectra, and their amount of energetic disorder. We used a specially built femtosecond spectrometer, combining tunable narrowband pump with broadband white-light probe pulses, together with an analytical method based on derivative spectroscopy for disentangling the congested transient absorption spectra of LH1 and LH2 complexes. This procedure allows real-time tracking of the forward (LH2 → LH1) and backward (LH2←LH1) ET processes and unambiguous determination of the corresponding rate constants. In low-light grown samples, we measured lower ET rates in both directions with respect to high-light ones, which is explained by reduced spectral overlap between B850 and B875 due to partial redistribution of oscillator strength into a higher energetic exciton transition. We find that the low-light adaptation in R. palustris leads to a reduced elementary backward ET rate, in accordance with the low probability of two simultaneous excitations reaching the same LH1/RC complex under weak illumination. Our study suggests that backward ET is not just an inevitable consequence of vectorial ET with small energetic offsets, but is in fact actively managed by photosynthetic bacteria. PMID:22307601

  9. Chlorophyll-binding proteins revisited - a multigenic family of light-harvesting and stress proteins from a brown algal perspective

    PubMed Central

    2010-01-01

    Background Chlorophyll-binding proteins (CBPs) constitute a large family of proteins with diverse functions in both light-harvesting and photoprotection. The evolution of CBPs has been debated, especially with respect to the origin of the LI818 subfamily, members of which function in non-photochemical quenching and have been found in chlorophyll a/c-containing algae and several organisms of the green lineage, but not in red algae so far. The recent publication of the Ectocarpus siliculosus genome represents an opportunity to expand on previous work carried out on the origin and function of CBPs. Results The Ectocarpus genome codes for 53 CBPs falling into all major families except the exclusively green family of chlorophyll a/b binding proteins. Most stress-induced CBPs belong to the LI818 family. However, we highlight a few stress-induced CBPs from Phaeodactylum tricornutum and Chondrus crispus that belong to different sub-families and are promising targets for future functional studies. Three-dimensional modeling of two LI818 proteins revealed features common to all LI818 proteins that are likely to interfere with their capacity to bind chlorophyll b and lutein, but may enable binding of chlorophyll c and fucoxanthin. In the light of this finding, we examined the possibility that LI818 proteins may have originated in a chlorophyll c/fucoxanthin containing organism and compared this scenario to three alternatives: an independent evolution of LI818 proteins in different lineages, an ancient origin together with the first CBPs, before the separation of the red and the green lineage, or an origin in the green lineage and a transfer to an ancestor of haptophytes and heterokonts during a cryptic endosymbiosis event. Conclusions Our findings reinforce the idea that the LI818 family of CBPs has a role in stress response. In addition, statistical analyses of phylogenetic trees show an independent origin in different eukaryotic lineages or a green algal origin of LI818

  10. Minimal Model of Quantum Kinetic Clusters for the Energy-Transfer Network of a Light-Harvesting Protein Complex.

    PubMed

    Wu, Jianlan; Tang, Zhoufei; Gong, Zhihao; Cao, Jianshu; Mukamel, Shaul

    2015-04-01

    The energy absorbed in a light-harvesting protein complex is often transferred collectively through aggregated chromophore clusters. For population evolution of chromophores, the time-integrated effective rate matrix allows us to construct quantum kinetic clusters quantitatively and determine the reduced cluster-cluster transfer rates systematically, thus defining a minimal model of energy-transfer kinetics. For Fenna-Matthews-Olson (FMO) and light-havrvesting complex II (LCHII) monomers, quantum Markovian kinetics of clusters can accurately reproduce the overall energy-transfer process in the long-time scale. The dominant energy-transfer pathways are identified in the picture of aggregated clusters. The chromophores distributed extensively in various clusters can assist a fast and long-range energy transfer.

  11. Mutations that affect structure and assembly of light-harvesting proteins in the cyanobacterium Synechocystis sp. strain 6701

    SciTech Connect

    Anderson, L.K.; Rayner, M.C.; Eiserling, F.A.

    1987-01-01

    The unicellular cyanobacterium Synechocystis sp. strain 6701 was mutagenized with UV irradiation and screened for pigment changes that indicated genetic lesions involving the light-harvesting proteins of the phycobilisome. A previous examination of the pigment mutant UV16 showed an assembly defect in the phycocyanin component of the phycobilisome. Mutagenesis of UV16 produced an additional double mutant, UV16-40, with decreased phycoerythrin content. Phycocyanin and phycoerythrin were isolated from UV16-40 and compared with normal biliproteins. The results suggested that the UV16 mutation affected the alpha subunit of phycocyanin, while the phycoerythrin beta subunit from UV16-40 had lost one of its three chromophores. Characterization of the unassembled phycobilisome components in these mutants suggests that these strains will be useful for probing in vivo the regulated expression and assembly of phycobilisomes.

  12. A light-harvesting antenna protein retains its folded conformation in the absence of protein-lipid and protein-pigment interactions.

    PubMed

    Kikuchi, J; Asakura, T; Loach, P A; Parkes-Loach, P S; Shimada, K; Hunter, C N; Conroy, M J; Williamson, M P

    1999-04-15

    The first study by nmr of the integral membrane protein, the bacterial light-harvesting (LH) antenna protein LH1 beta, is reported. The photosynthetic apparatus of purple bacteria contains two different kinds of antenna complexes (LH1 and LH2), which consist of two small integral membrane proteins alpha and beta, each of approximately 6 kDa, and bacteriochlorophyll and carotenoid pigments. We have purified the antenna polypeptide LH1 beta from Rhodobacter sphaeroides, and have recorded CD spectra and a series of two-dimensional nmr spectra. A comparison of CD spectra of LH1 beta observed in organic solvents and detergent micelles shows that the helical character of the peptide does not change appreciably between the two milieus. A significantly high-field shifted methyl signal was observed both in organic solvents and in detergent micelles, implying that a similar three-dimensional structure is present in each case. However, the 1H-nmr signals observed in organic solvents had a narrower line width and better resolution, and it is shown that in this case organic solvents provide a better medium for nmr studies than detergent micelles. A sequential assignment has been carried out on the C-terminal transmembrane region, which is the region in which the pigment is bound. The region is shown to have a helical structure by the chemical shift values of the alpha-CH protons and the presence of nuclear Overhauser effects characteristic of helices. An analysis of the amide proton chemical shifts of the residues surrounding the histidine chlorophyll ligand suggests that the local structure is well ordered even in the absence of protein-lipid and protein-pigment interactions. Its structure was determined from 348 nmr-derived constraints by using distance geometry calculations. The polypeptide contains an alpha-helix extending from Leu19 (position of cytoplasmic surface) to Trp44 (position of periplasmic surface). The helix is bent, as expected from the amide proton chemical

  13. Molecular topology of the photosynthetic light-harvesting pigment complex, peridinin-chlorophyll a-protein, from marine dinoflagellates.

    PubMed

    Song, P S; Koka, P; Prézelin, B B; Haxo, F T

    1976-10-01

    The photosynthetic light-harvesting complex, peridinin-chlorophyll a-protein, was isolated from several marine dinoflagellates including Glenodinium sp. by Sephadex and ion-exchange chromatography. The carotenoid (peridinin)-chlorophyll a ratio in the complex is estimated to be 4:1. The fluorescence excitation spectrum of the complex indicates that energy absorbed by the carotenoid is transferred to the chlorophyll a molecule with 100% efficiency. Fluorescence lifetime measurements indicate that the energy transfer is much faster than fluorescence emission from chlorophyll a. The four peridinin molecules within the complex appear to form two allowed exciton bands which split the main absorption band of the carotenoid into two circular dichronic bands (with negative ellipticity band at 538 nm and positive band at 463 nm in the case of peridinin-chlorophyl a-protein complex from Glenodinium sp.). The fluorescence polarization of chlorophyll a in the complex at 200 K is about 0.1 in both circular dichroic excitation bands of the carotenoid chromophore. From these circular dichroic and fluorescence polarization data, a possible molecular arrangement of the four peridinin and chlorophyll molecules has been deduced for the complex. The structure of the complex deduced is also consistent with the magnitude of the exciton spliting (ca. greater than 3000 cm-1) at the intermolecular distance in the dimer pair of peridinin (ca. 12 A). This structural feature accounts for the efficient light-harvesting process of dinoflagellates as the exciton interaction lengthens the lifetime of peridinin (radiative) and the complex topology increases the energy transfer probability. The complex is, therefore, a useful molecular model for elucidating the mechanism and efficiency of solar energy conversion in vivo as well as in vitro. PMID:987799

  14. Heat- and light-induced detachment of the light-harvesting antenna complexes of photosystem I in isolated stroma thylakoid membranes.

    PubMed

    Krumova, S B; Várkonyi, Zs; Lambrev, P H; Kovács, L; Todinova, S J; Busheva, M C; Taneva, S G; Garab, G

    2014-08-01

    The multisubunit pigment-protein complex of photosystem I (PSI) consists of a core and peripheral light-harvesting antenna (LHCI). PSI is thought to be a rather rigid system and very little is known about its structural and functional flexibility. Recent data, however, suggest LHCI detachment from the PSI supercomplex upon heat and light treatments. Furthermore, it was suggested that the splitting off of LHCI acts as a safety valve for PSI core upon photoinhibition (Alboresi et al., 2009). In this work we analyzed the heat- and light-induced reorganizations in isolated PSI vesicles (stroma membrane vesicles enriched in PSI). Using differential scanning calorimetry we revealed a stepwise disassembly of PSI supercomplex above 50°C. Circular dichroism, sucrose gradient centrifugation and 77K fluorescence experiments identified the sequence of events of PSI destabilization: 3min heating at 60°C or 40min white light illumination at 25°C resulted in pronounced Lhca1/4 detachment from the PSI supercomplex, which is then followed by the degradation of Lhca2/3. The similarity of the main structural effects due to heat and light treatments supports the notion that thermo-optic mechanism, structural changes induced by ultrafast local thermal transients, which has earlier been shown to be responsible for structural changes in the antenna system of photosystem II, can also regulate the assembly and functioning of PSI antenna.

  15. Cyanobacterial flv4-2 Operon-Encoded Proteins Optimize Light Harvesting and Charge Separation in Photosystem II.

    PubMed

    Chukhutsina, Volha; Bersanini, Luca; Aro, Eva-Mari; van Amerongen, Herbert

    2015-05-01

    Photosystem II (PSII) complexes drive the water-splitting reaction necessary to transform sunlight into chemical energy. However, too much light can damage and disrupt PSII. In cyanobacteria, the flv4-2 operon encodes three proteins (Flv2, Flv4, and Sll0218), which safeguard PSII activity under air-level CO2 and in high light conditions. However, the exact mechanism of action of these proteins has not been clarified yet. We demonstrate that the PSII electron transfer properties are influenced by the flv4-2 operon-encoded proteins. Accelerated secondary charge separation kinetics was observed upon expression/overexpression of the flv4-2 operon. This is likely induced by docking of the Flv2/Flv4 heterodimer in the vicinity of the QB pocket of PSII, which, in turn, increases the QB redox potential and consequently stabilizes forward electron transfer. The alternative electron transfer route constituted by Flv2/Flv4 sequesters electrons from QB(-) guaranteeing the dissipation of excess excitation energy in PSII under stressful conditions. In addition, we demonstrate that in the absence of the flv4-2 operon-encoded proteins, about 20% of the phycobilisome antenna becomes detached from the reaction centers, thus decreasing light harvesting. Phycobilisome detachment is a consequence of a decreased relative content of PSII dimers, a feature observed in the absence of the Sll0218 protein.

  16. Cyanobacterial flv4-2 Operon-Encoded Proteins Optimize Light Harvesting and Charge Separation in Photosystem II.

    PubMed

    Chukhutsina, Volha; Bersanini, Luca; Aro, Eva-Mari; van Amerongen, Herbert

    2015-05-01

    Photosystem II (PSII) complexes drive the water-splitting reaction necessary to transform sunlight into chemical energy. However, too much light can damage and disrupt PSII. In cyanobacteria, the flv4-2 operon encodes three proteins (Flv2, Flv4, and Sll0218), which safeguard PSII activity under air-level CO2 and in high light conditions. However, the exact mechanism of action of these proteins has not been clarified yet. We demonstrate that the PSII electron transfer properties are influenced by the flv4-2 operon-encoded proteins. Accelerated secondary charge separation kinetics was observed upon expression/overexpression of the flv4-2 operon. This is likely induced by docking of the Flv2/Flv4 heterodimer in the vicinity of the QB pocket of PSII, which, in turn, increases the QB redox potential and consequently stabilizes forward electron transfer. The alternative electron transfer route constituted by Flv2/Flv4 sequesters electrons from QB(-) guaranteeing the dissipation of excess excitation energy in PSII under stressful conditions. In addition, we demonstrate that in the absence of the flv4-2 operon-encoded proteins, about 20% of the phycobilisome antenna becomes detached from the reaction centers, thus decreasing light harvesting. Phycobilisome detachment is a consequence of a decreased relative content of PSII dimers, a feature observed in the absence of the Sll0218 protein. PMID:25704162

  17. Grana-Localized Proteins, RIQ1 and RIQ2, Affect the Organization of Light-Harvesting Complex II and Grana Stacking in Arabidopsis[OPEN

    PubMed Central

    Yokoyama, Ryo; Yamamoto, Hiroshi; Kondo, Maki; Takeda, Satomi; Ifuku, Kentaro; Fukao, Yoichiro; Kamei, Yasuhiro; Nishimura, Mikio; Shikanai, Toshiharu

    2016-01-01

    Grana are stacked thylakoid membrane structures in land plants that contain PSII and light-harvesting complex II proteins (LHCIIs). We isolated two Arabidopsis thaliana mutants, reduced induction of non-photochemical quenching1 (riq1) and riq2, in which stacking of grana was enhanced. The curvature thylakoid 1a (curt1a) mutant was previously shown to lack grana structure. In riq1 curt1a, the grana were enlarged with more stacking, and in riq2 curt1a, the thylakoids were abnormally stacked and aggregated. Despite having different phenotypes in thylakoid structure, riq1, riq2, and curt1a showed a similar defect in the level of nonphotochemical quenching of chlorophyll fluorescence (NPQ). In riq curt1a double mutants, NPQ induction was more severely affected than in either single mutant. In riq mutants, state transitions were inhibited and the PSII antennae were smaller than in wild-type plants. The riq defects did not affect NPQ induction in the chlorophyll b-less mutant. RIQ1 and RIQ2 are paralogous and encode uncharacterized grana thylakoid proteins, but despite the high level of identity of the sequence, the functions of RIQ1 and RIQ2 were not redundant. RIQ1 is required for RIQ2 accumulation, and the wild-type level of RIQ2 did not complement the NPQ and thylakoid phenotypes in riq1. We propose that RIQ proteins link the grana structure and organization of LHCIIs. PMID:27600538

  18. Protein Configuration Landscape Fluctuations Revealed by Exciton Transition Polarizations in Single Light Harvesting Complexes.

    PubMed

    Tubasum, Sumera; Torbjörnsson, Magne; Yadav, Dheerendra; Camacho, Rafael; Söderlind, Gustaf; Scheblykin, Ivan G; Pullerits, Tõnu

    2016-02-01

    Protein is a flexible material with broad distribution of conformations forming an energy landscape of quasi-stationary states. Disentangling the system dynamics along this landscape is the key for understanding the functioning of the protein. Here we studied a photosynthetic antenna pigment-protein complex LH2 with single molecule two-dimensional polarization imaging. Modeling based on the Redfield relaxation theory well describes the observed polarization properties of LH2 fluorescence and fluorescence excitation, strongly suggesting that at 77 K the conformational subspace of the LH2 is limited to about three configurations with relatively frequent switching among each other. At room temperature the next level of fluctuations determines the conformational dynamics. The results support the multitier model of the energy landscape of proteins and demonstrate the potential of the method for the studies of structural dynamics in proteins.

  19. Zeaxanthin Binds to Light-Harvesting Complex Stress-Related Protein to Enhance Nonphotochemical Quenching in Physcomitrella patens[W

    PubMed Central

    Pinnola, Alberta; Dall’Osto, Luca; Gerotto, Caterina; Morosinotto, Tomas; Bassi, Roberto; Alboresi, Alessandro

    2013-01-01

    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. PMID:24014548

  20. Zeaxanthin binds to light-harvesting complex stress-related protein to enhance nonphotochemical quenching in Physcomitrella patens.

    PubMed

    Pinnola, Alberta; Dall'Osto, Luca; Gerotto, Caterina; Morosinotto, Tomas; Bassi, Roberto; Alboresi, Alessandro

    2013-09-01

    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.

  1. Photosynthetic acclimation: structural reorganisation of light harvesting antenna--role of redox-dependent phosphorylation of major and minor chlorophyll a/b binding proteins.

    PubMed

    Kargul, Joanna; Barber, James

    2008-03-01

    In order to carry out photosynthesis, plants and algae rely on the co-operative interaction of two photosystems: photosystem I and photosystem II. For maximum efficiency, each photosystem should absorb the same amount of light. To achieve this, plants and green algae have a mobile pool of chlorophyll a/b-binding proteins that can switch between being light-harvesting antenna for photosystem I or photosystem II, in order to maintain an optimal excitation balance. This switch, termed state transitions, involves the reversible phosphorylation of the mobile chlorophyll a/b-binding proteins, which is regulated by the redox state of the plastoquinone-mediating electron transfer between photosystem I and photosystem II. In this review, we will present the data supporting the function of redox-dependent phosphorylation of the major and minor chlorophyll a/b-binding proteins by the specific thylakoid-bound kinases (Stt7, STN7, TAKs) providing a molecular switch for the structural remodelling of the light-harvesting complexes during state transitions. We will also overview the latest X-ray crystallographic and electron microscopy-derived models for structural re-arrangement of the light-harvesting antenna during State 1-to-State 2 transition, in which the minor chlorophyll a/b-binding protein, CP29, and the mobile light-harvesting complex II trimer detach from the light-harvesting complex II-photosystem II supercomplex and associate with the photosystem I core in the vicinity of the PsaH/L/O/P domain.

  2. Regulation of Light-Harvesting Chlorophyll Protein Biosynthesis in Greening Seedlings 1

    PubMed Central

    Mathis, James N.; Burkey, Kent O.

    1987-01-01

    The biosynthesis of the chlorophyll a/b binding protein associated with photosystem II (LHC-II) was characterized during light-induced greening of etiolated barley (Hordeum vulgare [L.] cv Boone), maize (Zea mays [L.] Pioneer 3148), pea (Pisum sativum [L.] cv Progress 9), and soybean (Glycine max [L.] Merr. cv Ransom 2). Northern blot analysis revealed that pea LHC-II mRNA was present in dark-grown seedlings and accumulated rapidly within 1 hour following illumination with white light. In contrast, the accumulation of LHC-II mRNA was delayed in barley and soybean until 2 to 4 hours after illumination began. Single radial immunodiffusion analysis revealed that LHC-II polypeptides began to accumulate in all species between 4 and 8 hours although the protein was present in detectable levels at earlier times in certain species. In a pattern similar to the LHC-II protein accumulation, chlorophyll accumulated at increased rates between 4 and 8 hours of greening in all species following an initial delay. The absence of coordination between LHC-II mRNA and LHC-II protein accumulation that was clearly observed in pea suggested that transcription is not the factor that limits LHC-II complex formation during chloroplast development. The accumulation of chlorophyll and LHC-II protein appeared to coincide suggesting that chlorophyll biosynthesis may be a factor that limits LHC-II complex formation. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:16665840

  3. A PM6 study of Rhodopseudomonas Acidophila light harvesting center II B800 bacteriochlorophylls in representative protein environment.

    PubMed

    Türeli, Sina; Varnalı, Tereza

    2011-06-01

    Bacterial light-harvesting II (LH-II) centers contain two types of Bacteriochlorophylls (Bchl). One is named B800 and found as a single molecule within one monomer of the complex while the other named B850 is found as a dimer. Their names indicate their peak of UV absorbance around red spectrum. Both types of molecules are attached to the protein chain via ligation of their central Magnesium atom to an either Histidine or Deoxymethionine amino acid. They are also coordinated by peripheral hydrogen bonds that they accept with their carboxyl side group. Both the ligation and the hydrogen bonding are thought to have an effect on electronic structure of the Bchl hence its UV absorbance and energy transfer rate. Experiments and theoretic studies performed on this subject support the above idea. This theoretical molecular modeling study case aims to mimic the experimental mutations performed on certain amino acids in silico and study its effects on the electronic structure of Bchl. By comparison with experimental results it was observed that the likely place for the nearby Arginine is not below the plane of the Bchl as in the X-ray crystallographic structure but above the plane defined by the four nitrogen atoms and their rings. It was also seen that the coordination of the acetyl group is very sensitive to changes in ligation of the Bchl molecule.

  4. Structural Mechanism Underlying the Specific Recognition between the Arabidopsis State-Transition Phosphatase TAP38/PPH1 and Phosphorylated Light-Harvesting Complex Protein Lhcb1[OPEN

    PubMed Central

    Wei, Xuepeng; Guo, Jiangtao; Li, Mei; Liu, Zhenfeng

    2015-01-01

    During state transitions, plants regulate energy distribution between photosystems I and II through reversible phosphorylation and lateral migration of the major light-harvesting complex LHCII. Dephosphorylation of LHCII and the transition from state 2 to state 1 requires a thylakoid membrane-associated phosphatase named TAP38 or PPH1. TAP38/PPH1 specifically targets LHCII but not the core subunits of photosystem II, whereas the underlying molecular mechanism of their mutual recognition is currently unclear. Here, we present the structures of Arabidopsis thaliana TAP38/PPH1 in the substrate-free and substrate-bound states. The protein contains a type 2C serine/threonine protein phosphatase (PP2C) core domain, a Mn2+ (or Mg2+) binuclear center and two additional motifs contributing to substrate recognition. A 15-mer phosphorylated N-terminal peptide of Lhcb1 binds to TAP38/PPH1 on two surface clefts enclosed by the additional motifs. The first segment of the phosphopeptide is clamped by a pair of tooth-like arginine residues at Cleft 1 site. The binding adopts the lock-and-key mechanism with slight rearrangement of the substrate binding residues on TAP38/PPH1. Meanwhile, a more evident substrate-induced fitting occurs on Cleft 2 harboring the extended part of the phosphopeptide. The results unravel the bases for the specific recognition between TAP38/PPH1 and phosphorylated Lhcb1, a crucial step in state transitions. PMID:25888588

  5. Cytokinins modulate the expression of genes encoding the protein of the light-harvesting chlorophyll a/b complex.

    PubMed

    de la Serve, B T; Axelos, M; Péaud-Lenoël, C

    1985-05-01

    Tobacco cell suspension cultures responded to cytokinins (for instance kinetin) by full chloroplast differentiation. The hormone had the effect of stimulating the appearance of a few prominent plastid proteins. Synthesis of the light-harvesting chlorophyl a/b-binding protein (LHCP) in response to kinetin was noteworthy (Axelos M. et al.: Plant Sci Lett 33:201-212, 1984).Poly(A)(+)RNAs were prepared from cells grown in the presence of or without added kinetin. Poly(A)(+)RNA recovery and translation activity were not quantitatively altered by the hormone treatment. In vitro translation of polyadenylated mRNA into precursor polypeptides of LHCP (pLHCP) was quantified by immunoprecipitation and SDS-PAGE fractionation of pLHCP immunoprecipitates: pLHCP-mRNA translating activity was found to be stimulated in parallel to mature LHCP accumulation by kinetin-induced cells.Dot-blot and northern-blot hybridizations of poly(A)(+)RNA were carried out, using as a probe a pea LHCP-cDNA clone (Broglie R. et al.: Proc Natl Acad Sci USA 78: 7304-7308, 1981). A ten-fold increase of the level of pLHCP-encoding sequences was observed in poly(A)(+)RNA prepared from 9-d kinetin-stimulated cells, compared to control cells. Oligo(dT)-cellulose-excluded RNA fractions exhibited very low hybridization levels, in the same ratios as those obtained with poly(A)(+)RNA.Thus, the expression of LHCP-gene activity, in response to kinetin addition to tobacco cell suspension cultures, is regulated by the level of pLHCP-encoding mRNA rather than by translational or post-translational controls. re]19850218 rv]19850605 ac]19850613. PMID:24306651

  6. Heterologous Expression of Moss Light-harvesting Complex Stress-related 1 (LHCSR1), the Chlorophyll a-Xanthophyll Pigment-protein Complex Catalyzing Non-photochemical Quenching, in Nicotiana sp.*

    PubMed Central

    Pinnola, Alberta; Ghin, Leonardo; Gecchele, Elisa; Merlin, Matilde; Alboresi, Alessandro; Avesani, Linda; Pezzotti, Mario; Capaldi, Stefano; Cazzaniga, Stefano; Bassi, Roberto

    2015-01-01

    Oxygenic photosynthetic organisms evolved mechanisms for thermal dissipation of energy absorbed in excess to prevent formation of reactive oxygen species. The major and fastest component, called non-photochemical quenching, occurs within the photosystem II antenna system by the action of two essential light-harvesting complex (LHC)-like proteins, photosystem II subunit S (PSBS) in plants and light-harvesting complex stress-related (LHCSR) in green algae and diatoms. In the evolutionary intermediate Physcomitrella patens, a moss, both gene products are active. These proteins, which are present in low amounts, are difficult to purify, preventing structural and functional analysis. Here, we report on the overexpression of the LHCSR1 protein from P. patens in the heterologous systems Nicotiana benthamiana and Nicotiana tabacum using transient and stable nuclear transformation. We show that the protein accumulated in both heterologous systems is in its mature form, localizes in the chloroplast thylakoid membranes, and is correctly folded with chlorophyll a and xanthophylls but without chlorophyll b, an essential chromophore for plants and algal LHC proteins. Finally, we show that recombinant LHCSR1 is active in quenching in vivo, implying that the recombinant protein obtained is a good material for future structural and functional studies. PMID:26260788

  7. Heterologous expression of moss light-harvesting complex stress-related 1 (LHCSR1), the chlorophyll a-xanthophyll pigment-protein complex catalyzing non-photochemical quenching, in Nicotiana sp.

    PubMed

    Pinnola, Alberta; Ghin, Leonardo; Gecchele, Elisa; Merlin, Matilde; Alboresi, Alessandro; Avesani, Linda; Pezzotti, Mario; Capaldi, Stefano; Cazzaniga, Stefano; Bassi, Roberto

    2015-10-01

    Oxygenic photosynthetic organisms evolved mechanisms for thermal dissipation of energy absorbed in excess to prevent formation of reactive oxygen species. The major and fastest component, called non-photochemical quenching, occurs within the photosystem II antenna system by the action of two essential light-harvesting complex (LHC)-like proteins, photosystem II subunit S (PSBS) in plants and light-harvesting complex stress-related (LHCSR) in green algae and diatoms. In the evolutionary intermediate Physcomitrella patens, a moss, both gene products are active. These proteins, which are present in low amounts, are difficult to purify, preventing structural and functional analysis. Here, we report on the overexpression of the LHCSR1 protein from P. patens in the heterologous systems Nicotiana benthamiana and Nicotiana tabacum using transient and stable nuclear transformation. We show that the protein accumulated in both heterologous systems is in its mature form, localizes in the chloroplast thylakoid membranes, and is correctly folded with chlorophyll a and xanthophylls but without chlorophyll b, an essential chromophore for plants and algal LHC proteins. Finally, we show that recombinant LHCSR1 is active in quenching in vivo, implying that the recombinant protein obtained is a good material for future structural and functional studies. PMID:26260788

  8. Heterologous expression of moss light-harvesting complex stress-related 1 (LHCSR1), the chlorophyll a-xanthophyll pigment-protein complex catalyzing non-photochemical quenching, in Nicotiana sp.

    PubMed

    Pinnola, Alberta; Ghin, Leonardo; Gecchele, Elisa; Merlin, Matilde; Alboresi, Alessandro; Avesani, Linda; Pezzotti, Mario; Capaldi, Stefano; Cazzaniga, Stefano; Bassi, Roberto

    2015-10-01

    Oxygenic photosynthetic organisms evolved mechanisms for thermal dissipation of energy absorbed in excess to prevent formation of reactive oxygen species. The major and fastest component, called non-photochemical quenching, occurs within the photosystem II antenna system by the action of two essential light-harvesting complex (LHC)-like proteins, photosystem II subunit S (PSBS) in plants and light-harvesting complex stress-related (LHCSR) in green algae and diatoms. In the evolutionary intermediate Physcomitrella patens, a moss, both gene products are active. These proteins, which are present in low amounts, are difficult to purify, preventing structural and functional analysis. Here, we report on the overexpression of the LHCSR1 protein from P. patens in the heterologous systems Nicotiana benthamiana and Nicotiana tabacum using transient and stable nuclear transformation. We show that the protein accumulated in both heterologous systems is in its mature form, localizes in the chloroplast thylakoid membranes, and is correctly folded with chlorophyll a and xanthophylls but without chlorophyll b, an essential chromophore for plants and algal LHC proteins. Finally, we show that recombinant LHCSR1 is active in quenching in vivo, implying that the recombinant protein obtained is a good material for future structural and functional studies.

  9. Light Harvesting and White-Light Generation in a Composite of Carbon Dots and Dye-Encapsulated BSA-Protein-Capped Gold Nanoclusters.

    PubMed

    Barman, Monoj Kumar; Paramanik, Bipattaran; Bain, Dipankar; Patra, Amitava

    2016-08-01

    Several strategies have been adopted to design an artificial light-harvesting system in which light energy is captured by peripheral chromophores and it is subsequently transferred to the core via energy transfer. A composite of carbon dots and dye-encapsulated BSA-protein-capped gold nanoclusters (AuNCs) has been developed for efficient light harvesting and white light generation. Carbon dots (C-dots) act as donor and AuNCs capped with BSA protein act as acceptor. Analysis reveals that energy transfer increases from 63 % to 83 % in presence of coumarin dye (C153), which enhances the cascade energy transfer from carbon dots to AuNCs. Bright white light emission with a quantum yield of 19 % under the 375 nm excitation wavelength is achieved by changing the ratio of components. Interesting findings reveal that the efficient energy transfer in carbon-dot-metal-cluster nanocomposites may open up new possibilities in designing artificial light harvesting systems for future applications. PMID:27383453

  10. Reversible changes in macroorganization of the light-harvesting chlorophyll a/b pigment-protein complex detected by circular dichroism

    SciTech Connect

    Garab, G.; Leegood, R.C.; Walker, D.A.; Sutherland, J.C.; Hind, G.

    1988-04-05

    Light-induced changes in circular dichroism (CD) were studied in thylakoids isolated from spinach. The following features of CD responses occurring in the time range of 10 s to 1-3 min were noted: (i) The kinetics and relative amplitudes of the responses are similar over broad spectral ranges surrounding the major CD bands, i.e., between 670 and 760 nm and between 480 and 550 nm. This applies not only to randomly oriented samples but also to magnetically aligned membranes having markedly different CD spectra in the dark. (ii) Photosystem I is much more effective than photosystem II and can drive a 40-80% decrease in CD signal relative to the dark control level. (iii) Photosystem I driven changes are fully inhibited by nigericin or NH/sub 4/Cl but are largely insensitive to gramicidin. CD changes driven by photosystem II, on the other hand, are sensitive to all of these reagents. (iv) The CD responses can be shown to originate in circular differential scattering rather than in circular differential absorbance. They can also be distinguished from light-induced, nonpolarized scattering changes. The data are qualitatively evaluated with respect to the theory of circular differential scattering of large helically organized macroaggregates, the size of which is commensurate with the wavelength of the measuring beam. The observed decrease of the large CD signal is ascribed to a partial loss of macrohelicity in the light-harvesting chlorophyll a/b protein complex, in response to a proton gradient and/or surface electrical field generated most effectively by photosystem I.

  11. The P-700-chlorophyl alpha-protein complex and two major light-harvesting complexes of Acrocarpia paniculata and other brown seaweeds.

    PubMed

    Barrett, J; Anderson, J M

    1980-05-01

    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. PMID:7378391

  12. Normal mode analysis of the spectral density of the Fenna-Matthews-Olson light-harvesting protein: how the protein dissipates the excess energy of excitons.

    PubMed

    Renger, Thomas; Klinger, Alexander; Steinecker, Florian; Schmidt am Busch, Marcel; Numata, Jorge; Müh, Frank

    2012-12-20

    We report a method for the structure-based calculation of the spectral density of the pigment-protein coupling in light-harvesting complexes that combines normal-mode analysis with the charge density coupling (CDC) and transition charge from electrostatic potential (TrEsp) methods for the computation of site energies and excitonic couplings, respectively. The method is applied to the Fenna-Matthews-Olson (FMO) protein in order to investigate the influence of the different parts of the spectral density as well as correlations among these contributions on the energy transfer dynamics and on the temperature-dependent decay of coherences. The fluctuations and correlations in excitonic couplings as well as the correlations between coupling and site energy fluctuations are found to be 1 order of magnitude smaller in amplitude than the site energy fluctuations. Despite considerable amplitudes of that part of the spectral density which contains correlations in site energy fluctuations, the effect of these correlations on the exciton population dynamics and dephasing of coherences is negligible. The inhomogeneous charge distribution of the protein, which causes variations in local pigment-protein coupling constants of the normal modes, is responsible for this effect. It is seen thereby that the same building principle that is used by nature to create an excitation energy funnel in the FMO protein also allows for efficient dissipation of the excitons' excess energy.

  13. Polyclonal antibodies to light-harvesting CHL-protein of PSII (LHC II) in marine green algae Bryopsis corticulans

    NASA Astrophysics Data System (ADS)

    Wu, Xiaonan; Zhou, Baicheng; Tseng, C. K.

    1992-06-01

    Polyc·lonal antibodies raised against LHC II isolated from SDS-solubilized Bryopsis corticulans thylakiod membranes by SDS-PAGE, were characterised by double immunodiffusion, Rocket immunoelectrophoresis and antigen-antibody crossed immunoelectro-phoresis assays showed the antibodies had strong cross-reaction with all B. corticulans LHC II components (even with those which were incubated in boiling water) and showed immunological cross-reactivity with LHC II polypeptides of spinach and the marine green alga Codium fragile. The results suggested that LHC II of different species had similar antigenic determinants and also conservation of amino acid sequences of the polypeptides during evolution, and that the antibodies could cross react with apoproteins of D2 proteins (which contain P680) from B. corticulans, spinach and C. fragile, but not with apoproteins of P700 Chl-proteins. Our results indicated some similarities in primary structure between LHC II of different species, and between LHC II and D2 proteins of marine green algae and spinach. Our finding that D2 and P700 Chl-proteins are not immunologically related suggested that P700 Chl-proteins and D2 proteins pass through independent evolutionary pathways.

  14. The proteolysis adaptor, NblA, initiates protein pigment degradation by interacting with the cyanobacterial light-harvesting complexes.

    PubMed

    Sendersky, Eleonora; Kozer, Noga; Levi, Mali; Garini, Yuval; Shav-Tal, Yaron; Schwarz, Rakefet

    2014-07-01

    Degradation of the cyanobacterial protein pigment complexes, the phycobilisomes, is a central acclimation response that controls light energy capture. The small protein, NblA, is essential for proteolysis of these large complexes, which may reach a molecular mass of up to 4 MDa. Interactions of NblA in vitro supported the suggestion that NblA is a proteolysis adaptor that labels the pigment proteins for degradation. The mode of operation of NblA in situ, however, remained unresolved. Particularly, it was unclear whether NblA interacts with phycobilisome proteins while part of the large complex, or alternatively interaction with NblA, necessitates dissociation of pigment subunits from the assembly. Fluorescence intensity profiles demonstrated the preferential presence of NblA::GFP (green fluorescent protein) at the photosynthetic membranes, indicating co-localization with phycobilisomes. Furthermore, fluorescence lifetime imaging microscopy provided in situ evidence for interaction of NblA with phycobilisome protein pigments. Additionally, we demonstrated the role of NblA in vivo as a proteolysis tag based on the rapid degradation of the fusion protein NblA::GFP compared with free GFP. Taken together, these observations demonstrated in vivo the role of NblA as a proteolysis adaptor. Additionally, the interaction of NblA with phycobilisomes indicates that the dissociation of protein pigment subunits from the large complex is not a prerequisite for interaction with this adaptor and, furthermore, implicates NblA in the disassembly of the protein pigment complex. Thus, we suggest that, in the case of proteolysis of the phycobilisome, the adaptor serves a dual function: undermining the complex stability and designating the dissociated pigments for degradation.

  15. PS2013 Satellite Workshop on Photosynthetic Light-Harvesting Systems

    SciTech Connect

    Niederman, Robert A.; Blankenship, Robert E.; Frank, Harry A.

    2015-02-07

    These funds were used for partial support of the PS2013 Satellite Workshop on Photosynthetic Light-Harvesting Systems, that was held on 8-11 August, 2013, at Washington University, St. Louis, MO. This conference, held in conjunction with the 16th International Congress on Photosynthesis/St. Louis, continued a long tradition of light-harvesting satellite conferences that have been held prior to the previous six international photosynthesis congresses. In this Workshop, the basis was explored for the current interest in replacing fossil fuels with energy sources derived form direct solar radiation, coupled with light-driven electron transport in natural photosynthetic systems and how they offer a valuable blueprint for conversion of sunlight to useful energy forms. This was accomplished through sessions on the initial light-harvesting events in the biological conversion of solar energy to chemically stored energy forms, and how these natural photosynthetic processes serve as a guide to the development of robust bio-hybrid and artificial systems for solar energy conversion into both electricity or chemical fuels. Organized similar to a Gordon Research Conference, a lively, informal and collegial setting was established, highlighting the exchange of exciting new data and unpublished results from ongoing studies. A significant amount of time was set aside for open discussion and interactive poster sessions, with a special session devoted to oral presentations by talented students and postdoctoral fellows judged to have the best posters. This area of research has seen exceptionally rapid progress in recent years, with the availability of a number of antenna protein structures at atomic resolution, elucidation of the molecular surface architecture of native photosynthetic membranes by atomic force microscopy and the maturing of ultrafast spectroscopic and molecular biological techniques for the investigation and manipulation of photosynthetic systems. The conferees

  16. Identification of light-harvesting chlorophyll a/b-binding protein genes of Zostera marina L. and their expression under different environmental conditions

    NASA Astrophysics Data System (ADS)

    Kong, Fanna; Zhou, Yang; Sun, Peipei; Cao, Min; Li, Hong; Mao, Yunxiang

    2016-02-01

    Photosynthesis includes the collection of light and the transfer of solar energy using light-harvesting chlorophyll a/b-binding (LHC) proteins. In high plants, the LHC gene family includes LHCA and LHCB sub-families, which encode proteins constituting the light-harvesting complex of photosystems I and II. Zostera marina L. is a monocotyledonous angiosperm and inhabits submerged marine environments rather than land environments. We characterized the Lhca and Lhcb gene families of Z. marina from the expressed sequence tags (EST) database. In total, 13 unigenes were annotated as ZmLhc, 6 in Lhca family and 7 in ZmLhcb family. ZmLHCA and ZmLHCB contained the conservative LHC motifs and amino acid residues binding chlorophyll. The average similarity among mature ZmLHCA and ZmLHCB was 48.91% and 48.66%, respectively, which indicated a high degree of divergence within ZmLHChc gene family. The reconstructed phylogenetic tree showed that the tree topology and phylogenetic relationship were similar to those reported in other high plants, suggesting that the Lhc genes were highly conservative and the classification of ZmLhc genes was consistent with the evolutionary position of Z. marina. Real-time reverse transcription (RT) PCR analysis showed that different members of ZmLhca and ZmLhcb responded to a stress in different expression patterns. Salinity, temperature, light intensity and light quality may affect the expression of most ZmLhca and ZmLhcb genes. Inorganic carbon concentration and acidity had no obvious effect on ZmLhca and ZmLhcb gene expression, except for ZmLhca6.

  17. Genetic Complementation and Kinetic Analyses of Rhodobacter capsulatus ORF1696 Mutants Indicate that the ORF1696 Protein Enhances Assembly of the Light-Harvesting I Complex

    PubMed Central

    Young, C. S.; Reyes, R. C.; Beatty, J. T.

    1998-01-01

    Rhodobacter capsulatus ORF1696 mutant strains were created by insertion of antibiotic resistance cartridges at different sites within the ORF1696 gene in a strain that lacks the light-harvesting II (LHII) complex. Steady-state absorption spectroscopy profiles and the kinetics of the light-harvesting I (LHI) complex assembly and decay were used to evaluate the function of the ORF1696 protein in various strains. All of the mutant strains were found to be deficient in the LHI complex, including one (ΔNae) with a disruption located 13 codons before the 3′ end of the gene. A 5′-proximal disruption after the 31st codon of ORF1696 resulted in a mutant strain (ΔMun) with a novel absorption spectrum. The two strains with more 3′ disruptions (ΔStu and ΔNae) were restored nearly to the parental strain phenotype when trans complemented with a plasmid expressing the ORF1696 gene, but ΔMun was not. The absorption spectrum of ΔMun resembled that of a strain which had a polar mutation in ORF1696. We suggest that a rho-dependent transcription termination site exists between the MunI and proximal StuI sites of ORF1696. A comparison of LHI complex assembly kinetics showed that assembly occurred 2.6-fold faster in the parental strain than in strain ΔStu. In contrast, LHI complex decay occurred 1.7-fold faster in the ORF1696 parental strain than in ΔStu. These results indicate that the ORF1696 protein has a major effect on LHI complex assembly, and models of ORF1696 function are proposed. PMID:9537372

  18. Induction of efficient energy dissipation in the isolated light-harvesting complex of Photosystem II in the absence of protein aggregation.

    PubMed

    Ilioaia, Cristian; Johnson, Matthew P; Horton, Peter; Ruban, Alexander V

    2008-10-24

    Under excess illumination, the Photosystem II light-harvesting antenna of higher plants has the ability to switch into an efficient photoprotective mode, allowing safe dissipation of excitation energy into heat. In this study, we show induction of the energy dissipation state, monitored by chlorophyll fluorescence quenching, in the isolated major light-harvesting complex (LHCII) incorporated into a solid gel system. Removal of detergent caused strong fluorescence quenching, which was totally reversible. Singlet-singlet annihilation and gel electrophoresis experiments suggested that the quenched complexes were in the trimeric not aggregated state. Both the formation and recovery of this quenching state were inhibited by a cross-linker, implying involvement of conformational changes. Absorption and CD measurements performed on the samples in the quenched state revealed specific alterations in the spectral bands assigned to the red forms of chlorophyll a, neoxanthin, and lutein 1 molecules. The majority of these alterations were similar to those observed during LHCII aggregation. This suggests that not the aggregation process as such but rather an intrinsic conformational transition in the complex is responsible for establishment of quenching. 77 K fluorescence measurements showed red-shifted chlorophyll a fluorescence in the 690-705 nm region, previously observed in aggregated LHCII. The fact that all spectral changes associated with the dissipative mode observed in the gel were different from those of the partially denatured complex strongly argues against the involvement of protein denaturation in the observed quenching. The implications of these findings for proposed mechanisms of energy dissipation in the Photosystem II antenna are discussed.

  19. Evolutionary loss of light-harvesting proteins Lhcb6 and Lhcb3 in major land plant groups--break-up of current dogma.

    PubMed

    Kouřil, Roman; Nosek, Lukáš; Bartoš, Jan; Boekema, Egbert J; Ilík, Petr

    2016-05-01

    Photosynthesis in plants and algae relies on the coordinated function of photosystems (PS) I and II. Their efficiency is augmented by finely-tuned light-harvesting proteins (Lhcs) connected to them. The most recent Lhcs (in evolutionary terms), Lhcb6 and Lhcb3, evolved during the transition of plants from water to land and have so far been considered to be an essential characteristic of land plants. We used single particle electron microscopy and sequence analysis to study architecture and composition of PSII supercomplex from Norway spruce and related species. We have found that there are major land plant families that lack functional lhcb6 and lhcb3 genes, which notably changes the organization of PSII supercomplexes. The Lhcb6 and Lhcb3 proteins have been lost in the gymnosperm genera Picea and Pinus (family Pinaceae) and Gnetum (Gnetales). We also revealed that the absence of these proteins in Norway spruce modifies the PSII supercomplex in such a way that it resembles its counterpart in the alga Chlamydomonas reinhardtii, an evolutionarily older organism. Our results break a deep-rooted concept of Lhcb6 and Lhcb3 proteins being the essential characteristic of land plants, and beg the question of what the evolutionary benefit of their loss could be. PMID:27001142

  20. Identification of pH-sensing Sites in the Light Harvesting Complex Stress-related 3 Protein Essential for Triggering Non-photochemical Quenching in Chlamydomonas reinhardtii*

    PubMed Central

    Ballottari, Matteo; Truong, Thuy B.; De Re, Eleonora; Erickson, Erika; Stella, Giulio R.; Fleming, Graham R.; Bassi, Roberto; Niyogi, Krishna K.

    2016-01-01

    Light harvesting complex stress-related 3 (LHCSR3) is the protein essential for photoprotective excess energy dissipation (non-photochemical quenching, NPQ) in the model green alga Chlamydomonas reinhardtii. Activation of NPQ requires low pH in the thylakoid lumen, which is induced in excess light conditions and sensed by lumen-exposed acidic residues. In this work we have used site-specific mutagenesis in vivo and in vitro for identification of the residues in LHCSR3 that are responsible for sensing lumen pH. Lumen-exposed protonatable residues, aspartate and glutamate, were mutated to asparagine and glutamine, respectively. By expression in a mutant lacking all LHCSR isoforms, residues Asp117, Glu221, and Glu224 were shown to be essential for LHCSR3-dependent NPQ induction in C. reinhardtii. Analysis of recombinant proteins carrying the same mutations refolded in vitro with pigments showed that the capacity of responding to low pH by decreasing the fluorescence lifetime, present in the wild-type protein, was lost. Consistent with a role in pH sensing, the mutations led to a substantial reduction in binding the NPQ inhibitor dicyclohexylcarbodiimide. PMID:26817847

  1. Identification of pH-sensing Sites in the Light Harvesting Complex Stress-related 3 Protein Essential for Triggering Non-photochemical Quenching in Chlamydomonas reinhardtii.

    PubMed

    Ballottari, Matteo; Truong, Thuy B; De Re, Eleonora; Erickson, Erika; Stella, Giulio R; Fleming, Graham R; Bassi, Roberto; Niyogi, Krishna K

    2016-04-01

    Light harvesting complex stress-related 3 (LHCSR3) is the protein essential for photoprotective excess energy dissipation (non-photochemical quenching, NPQ) in the model green algaChlamydomonas reinhardtii Activation of NPQ requires low pH in the thylakoid lumen, which is induced in excess light conditions and sensed by lumen-exposed acidic residues. In this work we have used site-specific mutagenesisin vivoandin vitrofor identification of the residues in LHCSR3 that are responsible for sensing lumen pH. Lumen-exposed protonatable residues, aspartate and glutamate, were mutated to asparagine and glutamine, respectively. By expression in a mutant lacking all LHCSR isoforms, residues Asp(117), Glu(221), and Glu(224)were shown to be essential for LHCSR3-dependent NPQ induction inC. reinhardtii Analysis of recombinant proteins carrying the same mutations refoldedin vitrowith pigments showed that the capacity of responding to low pH by decreasing the fluorescence lifetime, present in the wild-type protein, was lost. Consistent with a role in pH sensing, the mutations led to a substantial reduction in binding the NPQ inhibitor dicyclohexylcarbodiimide. PMID:26817847

  2. Controlling Light Harvesting with Light.

    PubMed

    Gwizdala, Michal; Berera, Rudi; Kirilovsky, Diana; van Grondelle, Rienk; Krüger, Tjaart P J

    2016-09-14

    When exposed to intense sunlight, all organisms performing oxygenic photosynthesis implement various photoprotective strategies to prevent potentially lethal photodamage. The rapidly responding photoprotective mechanisms, occurring in the light-harvesting pigment-protein antennae, take effect within tens of seconds, while the dramatic and potentially harmful light intensity fluctuations manifest also on shorter time scales. Here we show that, upon illumination, individual phycobilisomes from Synechocystis PCC 6803, which, in vivo under low-light conditions, harvest solar energy, and have the built-in capacity to switch rapidly and reversibly into light-activated energy-dissipating states. Simultaneously measured fluorescence intensity, lifetime, and spectra, compared with a multicompartmental kinetic model, revealed that essentially any subunit of a phycobilisome can be quenched, and that the core complexes were targeted most frequently. Our results provide the first evidence for fluorescence blinking from a biologically active system at physiological light intensities and suggest that the light-controlled switches to intrinsically available energy-dissipating states are responsible for a novel type of photoprotection in cyanobacteria. We anticipate other photosynthetic organisms to employ similar strategies to respond instantly to rapid solar light intensity fluctuations. A detailed understanding of the photophysics of photosynthetic antenna complexes is of great interest for bioinspired solar energy technologies. PMID:27546794

  3. Controlling Light Harvesting with Light.

    PubMed

    Gwizdala, Michal; Berera, Rudi; Kirilovsky, Diana; van Grondelle, Rienk; Krüger, Tjaart P J

    2016-09-14

    When exposed to intense sunlight, all organisms performing oxygenic photosynthesis implement various photoprotective strategies to prevent potentially lethal photodamage. The rapidly responding photoprotective mechanisms, occurring in the light-harvesting pigment-protein antennae, take effect within tens of seconds, while the dramatic and potentially harmful light intensity fluctuations manifest also on shorter time scales. Here we show that, upon illumination, individual phycobilisomes from Synechocystis PCC 6803, which, in vivo under low-light conditions, harvest solar energy, and have the built-in capacity to switch rapidly and reversibly into light-activated energy-dissipating states. Simultaneously measured fluorescence intensity, lifetime, and spectra, compared with a multicompartmental kinetic model, revealed that essentially any subunit of a phycobilisome can be quenched, and that the core complexes were targeted most frequently. Our results provide the first evidence for fluorescence blinking from a biologically active system at physiological light intensities and suggest that the light-controlled switches to intrinsically available energy-dissipating states are responsible for a novel type of photoprotection in cyanobacteria. We anticipate other photosynthetic organisms to employ similar strategies to respond instantly to rapid solar light intensity fluctuations. A detailed understanding of the photophysics of photosynthetic antenna complexes is of great interest for bioinspired solar energy technologies.

  4. Spectroscopic Investigations of the Photophysics of Cryptophyte Light-Harvesting

    NASA Astrophysics Data System (ADS)

    Dinshaw, Rayomond

    The biological significance of photosynthesis is indisputable as it is necessary for nearly all life on earth. Photosynthesis provides chemical energy for plants, algae, and bacteria, while heterotrophic organisms rely on these species as their ultimate food source. The initial step in photosynthesis requires the absorption of sunlight to create electronic excitations. Light-harvesting proteins play the functional role of capturing solar radiation and transferring the resulting excitation to the reaction centers where it is used to carry out the chemical reactions of photosynthesis. Despite the wide variety of light-harvesting protein structures and arrangements, most light-harvesting proteins are able to utilize the captured solar energy for charge separation with near perfect quantum efficiency.1 This thesis will focus on understanding the energy transfer dynamics and photophysics of a specific subset of light-harvesting antennae known as phycobiliproteins. These proteins are extracted from cryptophyte algae and are investigated using steady-state and ultrafast spectroscopic techniques.

  5. Phycobilisome: architecture of a light-harvesting supercomplex.

    PubMed

    Watanabe, Mai; Ikeuchi, Masahiko

    2013-10-01

    The phycobilisome (PBS) is an extra-membrane supramolecular complex composed of many chromophore (bilin)-binding proteins (phycobiliproteins) and linker proteins, which generally are colorless. PBS collects light energy of a wide range of wavelengths, funnels it to the central core, and then transfers it to photosystems. Although phycobiliproteins are evolutionarily related to each other, the binding of different bilin pigments ensures the ability to collect energy over a wide range of wavelengths. Spatial arrangement and functional tuning of the different phycobiliproteins, which are mediated primarily by linker proteins, yield PBS that is efficient and versatile light-harvesting systems. In this review, we discuss the functional and spatial tuning of phycobiliproteins with a focus on linker proteins.

  6. Crystallization and Structure Analysis of Membrane Proteins

    NASA Astrophysics Data System (ADS)

    Newman, Richard

    In recent years, there has been great progress in the determination of high-resolution three-dimensional (3D) structures of membrane proteins. The first major breakthrough came with the crystallization (1) and X-ray crystallography (2,3) of the bacterial photosynthetic reaction center (see refs. 4 and 5 for reviews). The structure of another, entirely different membrane protein, the bacterial outer membrane porin from Rhodobacter capsulatus, has now been determined by X-ray crystallography (6). Recent results by electron crystallography of two-dimensional (2D) crystals have been most encouraging. The high-resolution 3D structure of bacteriorhodopsin (7) plant light-harvesting complex (8) and projection maps of several other membrane proteins at similar resolutions (9-11) have been obtained by this technique. Electron crystallography seems particularly appropriate for membrane proteins that are prone to form 2D crystals, and it is hoped that many more structures will be determined in this way.

  7. Light-dependent chloroplast development and expression of a light-harvesting chlorophyll a/b-binding protein gene in the gymnosperm Ginkgo biloba.

    PubMed Central

    Chinn, E; Silverthorne, J

    1993-01-01

    Unlike conifers, the gymnosperm Ginkgo biloba is dependent on light for chlorophyll (Chl) synthesis and initiation of chloroplast development. Dark-grown seedlings show complete etiolation, including no detectable Chl accumulation, no leaf expansion, and increased hypocotyl elongation. When dark-grown seedlings are placed in white light, Chl synthesis and leaf expansion are initiated, but unlike angiosperms, which initiate rapid photomorphogenesis, Ginkgo takes at least 1 week to change to a normal light-regulated pattern of growth. A cDNA clone (pLhcb*Gb1) encoding a Chl a/b-binding protein of light-harvesting complex II from Ginkgo mRNA has been used as a probe for the expression of this family of mRNAs. We have found that, in common with angiosperms but in marked contrast to pines, Lhcb mRNA is expressed in a highly light-dependent manner. In addition to being expressed in light-grown leaves, this sequence is also expressed in the green tissues of immature seeds. The Lhcb mRNA appears during greening in parallel with the onset of Chl synthesis. The complete sequence of pLhcb*Gb1 has been determined and the deduced amino acid sequence was found to be of type I based on comparison with signature sequences of angiosperm and gymnosperm sequences. PMID:8022934

  8. Intrinsically unstructured phosphoprotein TSP9 regulates light harvesting in Arabidopsis thaliana.

    PubMed

    Fristedt, Rikard; Carlberg, Inger; Zygadlo, Agnieszka; Piippo, Mirva; Nurmi, Markus; Aro, Eva-Mari; Scheller, Henrik Vibe; Vener, Alexander V

    2009-01-20

    Thylakoid-soluble phosphoprotein of 9 kDa, TSP9, is an intrinsically unstructured plant-specific protein [Song, J., et al. (2006) Biochemistry 45, 15633-15643] with unknown function but established associations with light-harvesting proteins and peripheries of both photosystems [Hansson, M., et al. (2007) J. Biol. Chem. 282, 16214-16222]. To investigate the function of this protein, we used a combination of reverse genetics and biochemical and fluorescence measurement methods in Arabidopsis thaliana. Differential gene expression analysis of plants with a T-DNA insertion in the TSP9 gene using an array of 24000 Arabidopsis genes revealed disappearance of high light-dependent induction of a specific set of mostly signaling and unknown proteins. TSP9-deficient plants had reduced levels of in vivo phosphorylation of light-harvesting complex II polypeptides. Recombinant TSP9 was phosphorylated in light by thylakoid membranes isolated from the wild-type and mutant plants lacking STN8 protein kinase but not by the thylakoids deficient in STN7 kinase, essential for photosynthetic state transitions. TSP9-lacking mutant and RNAi plants with downregulation of TSP9 showed reduced ability to perform state transitions. The nonphotochemical quenching of chlorophyll fluorescence at high light intensities was also less efficient in the mutant compared to wild-type plants. Blue native electrophoresis of thylakoid membrane protein complexes revealed that TSP9 deficiency increased relative stability of photosystem II dimers and supercomplexes. It is concluded that TSP9 regulates plant light harvesting acting as a membrane-binding protein facilitating dissociation of light-harvesting proteins from photosystem II. PMID:19113838

  9. Photosynthetic light harvesting: excitons and coherence

    PubMed Central

    Fassioli, Francesca; Dinshaw, Rayomond; Arpin, Paul C.; Scholes, Gregory D.

    2014-01-01

    Photosynthesis begins with light harvesting, where specialized pigment–protein complexes transform sunlight into electronic excitations delivered to reaction centres to initiate charge separation. There is evidence that quantum coherence between electronic excited states plays a role in energy transfer. In this review, we discuss how quantum coherence manifests in photosynthetic light harvesting and its implications. We begin by examining the concept of an exciton, an excited electronic state delocalized over several spatially separated molecules, which is the most widely available signature of quantum coherence in light harvesting. We then discuss recent results concerning the possibility that quantum coherence between electronically excited states of donors and acceptors may give rise to a quantum coherent evolution of excitations, modifying the traditional incoherent picture of energy transfer. Key to this (partially) coherent energy transfer appears to be the structure of the environment, in particular the participation of non-equilibrium vibrational modes. We discuss the open questions and controversies regarding quantum coherent energy transfer and how these can be addressed using new experimental techniques. PMID:24352671

  10. Photosynthetic light harvesting: excitons and coherence.

    PubMed

    Fassioli, Francesca; Dinshaw, Rayomond; Arpin, Paul C; Scholes, Gregory D

    2014-03-01

    Photosynthesis begins with light harvesting, where specialized pigment-protein complexes transform sunlight into electronic excitations delivered to reaction centres to initiate charge separation. There is evidence that quantum coherence between electronic excited states plays a role in energy transfer. In this review, we discuss how quantum coherence manifests in photosynthetic light harvesting and its implications. We begin by examining the concept of an exciton, an excited electronic state delocalized over several spatially separated molecules, which is the most widely available signature of quantum coherence in light harvesting. We then discuss recent results concerning the possibility that quantum coherence between electronically excited states of donors and acceptors may give rise to a quantum coherent evolution of excitations, modifying the traditional incoherent picture of energy transfer. Key to this (partially) coherent energy transfer appears to be the structure of the environment, in particular the participation of non-equilibrium vibrational modes. We discuss the open questions and controversies regarding quantum coherent energy transfer and how these can be addressed using new experimental techniques.

  11. Thermostability and Photostability of Photosystem II in Leaves of the Chlorina-f2 Barley Mutant Deficient in Light-Harvesting Chlorophyll a/b Protein Complexes.

    PubMed Central

    Havaux, M.; Tardy, F.

    1997-01-01

    The chlorophyll-b-less chlorina-f2 barley mutant is deficient in the major as well as some minor light-harvesting chlorophyll-protein complexes of photosystem II (LHCII). Although the LHCII deficiency had relatively minor repercussions on the leaf photosynthetic performances, the responses of photosystem II (PSII) to elevated temperatures and to bright light were markedly modified. The chlorina-f2 mutation noticeably reduced the thermostability of PSII, with thermal denaturation of PSII starting at about 35[deg]C and 38.5[deg]C in chlorina-f2 and in the wild type, respectively. The increased susceptibility of PSII to heat stress in chlorina-f2 leaves was due to the weakness of its electron donor side, with moderate heat stress causing detachment of the 33-kD extrinsic PSII protein from the oxygen-evolving complex. Prolonged dark adaptation of chlorina-f2 leaves was also observed to inhibit the PSII donor side. However, weak illumination slowly reversed the dark-induced inhibition of PSII in chlorina-f2 and cancelled the difference in PSII thermostability observed between chlorina-f2 and wild-type leaves. The mutant was more sensitive to photoinhibition than the wild type, with strong light stress impairing the PSII donor side in chlorina-f2 but not in the wild type. This difference was not observed in anaerobiosis or in the presence of 3-(3,4-dichlorophenyl)- 1,1-dimethylurea, diuron. The acceptor side of PSII was only slightly affected by the mutation and/or the aforementioned stress conditions. Taken together, our results indicate that LHCII stabilize the PSII complexes and maintain the water-oxidizing system in a functional state under varying environmental conditions. PMID:12223653

  12. Biogeography of Photosynthetic Light-Harvesting Genes in Marine Phytoplankton

    PubMed Central

    Bibby, Thomas S.; Zhang, Yinan; Chen, Min

    2009-01-01

    Background Photosynthetic light-harvesting proteins are the mechanism by which energy enters the marine ecosystem. The dominant prokaryotic photoautotrophs are the cyanobacterial genera Prochlorococcus and Synechococcus that are defined by two distinct light-harvesting systems, chlorophyll-bound protein complexes or phycobilin-bound protein complexes, respectively. Here, we use the Global Ocean Sampling (GOS) Project as a unique and powerful tool to analyze the environmental diversity of photosynthetic light-harvesting genes in relation to available metadata including geographical location and physical and chemical environmental parameters. Methods All light-harvesting gene fragments and their metadata were obtained from the GOS database, aligned using ClustalX and classified phylogenetically. Each sequence has a name indicative of its geographic location; subsequent biogeographical analysis was performed by correlating light-harvesting gene budgets for each GOS station with surface chlorophyll concentration. Conclusion/Significance Using the GOS data, we have mapped the biogeography of light-harvesting genes in marine cyanobacteria on ocean-basin scales and show that an environmental gradient exists in which chlorophyll concentration is correlated to diversity of light-harvesting systems. Three functionally distinct types of light-harvesting genes are defined: (1) the phycobilisome (PBS) genes of Synechococcus; (2) the pcb genes of Prochlorococcus; and (3) the iron-stress-induced (isiA) genes present in some marine Synechococcus. At low chlorophyll concentrations, where nutrients are limited, the Pcb-type light-harvesting system shows greater genetic diversity; whereas at high chlorophyll concentrations, where nutrients are abundant, the PBS-type light-harvesting system shows higher genetic diversity. We interpret this as an environmental selection of specific photosynthetic strategy. Importantly, the unique light-harvesting system isiA is found in the iron

  13. Absence of the major light harvesting antenna proteins alters the redox properties of photosystem II reaction centres in the chlorine F2 mutant of barley

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although the chlorina F2 mutant of barley specifically exhibits reduced levels of the major light harvesting polypeptides (Lhcb) associated with photosystem II, thermoluminescence measurements of photosystem reaction centre photochemistry revealed that S2/S3QB- charge recombinations were shifted to ...

  14. Synergistic Two-Photon Absorption Enhancement in Photosynthetic Light Harvesting

    NASA Astrophysics Data System (ADS)

    Chen, Kuo-Mei; Chen, Yu-Wei; Gao, Ting-Fong

    2012-06-01

    The grand scale fixation of solar energies into chemical substances by photosynthetic reactions of light-harvesting organisms provides Earth's other life forms a thriving environment. Scientific explorations in the past decades have unraveled the fundamental photophysical and photochemical processes in photosynthesis. Higher plants, green algae, and light-harvesting bacteria utilize organized pigment-protein complexes to harvest solar power efficiently and the resultant electronic excitations are funneled into a reaction center, where the first charge separation process takes place. Here we show experimental evidences that green algae (Chlorella vulgaris) in vivo display a synergistic two-photon absorption enhancement in their photosynthetic light harvesting. Their absorption coefficients at various wavelengths display dramatic dependence on the photon flux. This newly found phenomenon is attributed to a coherence-electronic-energy-transfer-mediated (CEETRAM) photon absorption process of light-harvesting pigment-protein complexes of green algae. Under the ambient light level, algae and higher plants can utilize this quantum mechanical mechanism to create two entangled electronic excitations adjacently in their light-harvesting networks. Concerted multiple electron transfer reactions in the reaction centers and oxygen evolving complexes can be implemented efficiently by the coherent motion of two entangled excitons from antennae to the charge separation reaction sites. To fabricate nanostructured, synthetic light-harvesting apparatus, the paramount role of the CEETRAM photon absorption mechanism should be seriously considered in the strategic guidelines.

  15. High Yield Non-detergent Isolation of Photosystem I-Light-harvesting Chlorophyll II Membranes from Spinach Thylakoids: IMPLICATIONS FOR THE ORGANIZATION OF THE PS I ANTENNAE IN HIGHER PLANTS.

    PubMed

    Bell, Adam J; Frankel, Laurie K; Bricker, Terry M

    2015-07-24

    Styrene-maleic acid copolymer was used to effect a non-detergent partial solubilization of thylakoids from spinach. A high density membrane fraction, which was not solubilized by the copolymer, was isolated and was highly enriched in the Photosystem (PS) I-light-harvesting chlorophyll (LHC) II supercomplex and depleted of PS II, the cytochrome b6/f complex, and ATP synthase. The LHC II associated with the supercomplex appeared to be energetically coupled to PS I based on 77 K fluorescence, P700 photooxidation, and PS I electron transport light saturation experiments. The chlorophyll (Chl) a/b ratio of the PS I-LHC II membranes was 3.2 ± 0.9, indicating that on average, three LHC II trimers may associate with each PS I. The implication of these findings within the context of higher plant PS I antenna organization is discussed.

  16. First solid-state NMR analysis of uniformly ¹³C-enriched major light-harvesting complexes from Chlamydomonas reinhardtii and identification of protein and cofactor spin clusters.

    PubMed

    Pandit, Anjali; Morosinotto, Tomas; Reus, Michael; Holzwarth, Alfred R; Bassi, Roberto; de Groot, Huub J M

    2011-04-01

    The light-harvesting complex II (LHCII) is the main component of the antenna system of plants and green algae and plays a major role in the capture of sun light for photosynthesis. The LHCII complexes have also been proposed to play a key role in the optimization of photosynthetic efficiency through the process of state 1-state 2 transitions and are involved in down-regulation of photosynthesis under excess light by energy dissipation through non-photochemical quenching (NPQ). We present here the first solid-state magic-angle spinning (MAS) NMR data of the major light-harvesting complex (LHCII) of Chlamydomonas reinhardtii, a eukaryotic green alga. We are able to identify nuclear spin clusters of the protein and of its associated chlorophyll pigments in ¹³C-¹³C dipolar homonuclear correlation spectra on a uniformly ¹³C-labeled sample. In particular, we were able to resolve several chlorophyll 13¹ carbon resonances that are sensitive to hydrogen bonding to the 13¹-keto carbonyl group. The data show that ¹³C NMR signals of the pigments and protein sites are well resolved, thus paving the way to study possible structural reorganization processes involved in light-harvesting regulation through MAS solid-state NMR. PMID:21276419

  17. Silica entrapment for significantly stabilized, energy-conducting light-harvesting complex (LHCII).

    PubMed

    Roeder, Sebastian; Hobe, Stephan; Paulsen, Harald

    2014-12-01

    The major light-harvesting chlorophyll a/b complex (LHCII) of the photosynthetic apparatus in green plants consists of a membrane protein and numerous noncovalently bound pigments that make up about one-third of the molecular mass of the pigment-protein complex. Due to this high pigment density, LHCII is potentially interesting as a light-harvesting component in synthetic constructs. However, for such applications its stability needs to be significantly improved. In this work, LHCII was dramatically stabilized by enclosing it within polymerizing colloidal silica. The entrapped LHCII stayed functional at 50 °C for up to 24 h instead of a few minutes in detergent solution and clearly showed energy transfer between complexes. Entrapment yield was enhanced by a polycationic peptide attached to the N terminus. Both the extent of stabilization and the yield of entrapment strongly increased with decreasing diameters of the silica particles.

  18. Molecular design of the photosystem II light-harvesting antenna: photosynthesis and photoprotection.

    PubMed

    Horton, Peter; Ruban, Alexander

    2005-01-01

    The photosystem II (PSII) light-harvesting system carries out two essential functions, the efficient collection of light energy for photosynthesis, and the regulated dissipation of excitation energy in excess of that which can be used. This dual function requires structural and functional flexibility, in which light-harvesting proteins respond to an external signal, the thylakoid DeltapH, to induce feedback control. This process, referred to as non-photochemical quenching (NPQ) depends upon the xanthophyll cycle and the PsbS protein. In nature, NPQ is heterogeneous in terms of kinetics and capacity, and this adapts photosynthetic systems to the specific dynamic features of the light environment. The molecular features of the thylakoid membrane which may enable this flexibility and plasticity are discussed.

  19. Up-converted fluorescence from photosynthetic light-harvesting complexes linearly dependent on excitation intensity.

    PubMed

    Leiger, Kristjan; Freiberg, Arvi

    2016-01-01

    Weak up-converted fluorescence related to bacteriochlorophyll a was recorded from various detergent-isolated and membrane-embedded light-harvesting pigment-protein complexes as well as from the functional membranes of photosynthetic purple bacteria under continuous-wave infrared laser excitation at 1064 nm, far outside the optically allowed singlet absorption bands of the chromophore. The fluorescence increases linearly with the excitation power, distinguishing it from the previously observed two-photon excited fluorescence upon femtosecond pulse excitation. Possible mechanisms of this excitation are discussed.

  20. A structure-based model of energy transfer reveals the principles of light harvesting in photosystem II supercomplexes.

    PubMed

    Bennett, Doran I G; Amarnath, Kapil; Fleming, Graham R

    2013-06-19

    Photosystem II (PSII) initiates photosynthesis in plants through the absorption of light and subsequent conversion of excitation energy to chemical energy via charge separation. The pigment binding proteins associated with PSII assemble in the grana membrane into PSII supercomplexes and surrounding light harvesting complex II trimers. To understand the high efficiency of light harvesting in PSII requires quantitative insight into energy transfer and charge separation in PSII supercomplexes. We have constructed the first structure-based model of energy transfer in PSII supercomplexes. This model shows that the kinetics of light harvesting cannot be simplified to a single rate limiting step. Instead, substantial contributions arise from both excitation diffusion through the antenna pigments and transfer from the antenna to the reaction center (RC), where charge separation occurs. Because of the lack of a rate-limiting step, fitting kinetic models to fluorescence lifetime data cannot be used to derive mechanistic insight on light harvesting in PSII. This model will clarify the interpretation of chlorophyll fluorescence data from PSII supercomplexes, grana membranes, and leaves.

  1. The multiple roles of light-harvesting chlorophyll a/b-protein complexes define structure and optimize function of Arabidopsis chloroplasts: a study using two chlorophyll b-less mutants.

    PubMed

    Kim, Eun-Ha; Li, Xiao-Ping; Razeghifard, Reza; Anderson, Jan M; Niyogi, Krishna K; Pogson, Barry J; Chow, Wah Soon

    2009-08-01

    The multiple roles of light-harvesting chlorophyll a/b-protein complexes in the structure and function of Arabidopsis chloroplasts were investigated using two chlorophyll b-less mutants grown under metal halide lamps with a significant far-red component. In ch1-3, all six light-harvesting proteins of photosystem (PS) II were greatly decreased; in ch1-3lhcb5, Lhcb5 was completely absent while the other five proteins were further decreased. The thylakoids of ch1-3 were less negatively-charged than the wild type, and those of ch1-3lhcb5 were even less so. Despite the expected weaker electrostatic repulsion, however, thylakoids in leaves of the mutants were not well stacked, an effect we attribute to lower van der Waals attraction, lower electrostatic attraction between opposite charges, and the absence or instability of PSII supercomplexes and peripheral light-harvesting trimers. The quantum yield of oxygen evolution in leaves decreased from 0.109 (wild type) to 0.087 (ch1-3) and 0.081 (ch1-3lhcb5) O(2) (photon absorbed)(-1); we attribute this decrease to an excessive spillover from PSII to PSI, a limited PSII antenna, and increased light-independent thermal dissipation in PSII in the mutants. Destabilization of the donor side of PSII, indicated by slower electron donation to the redox-active tyrosine Y(Z)(*) in ch1-3, probably enhanced PSII susceptibility to photoinactivation, increased the non-functional PSII complexes in vivo, and further inactivated PSII complexes in vitro. The evolution of chlorophyll b-containing chloroplasts seems to fine-tune oxygenic photosynthesis.

  2. In Vitro Reconstitution of Light-harvesting Complexes of Plants and Green Algae

    PubMed Central

    Natali, Alberto; Roy, Laura M.; Croce, Roberta

    2014-01-01

    In plants and green algae, light is captured by the light-harvesting complexes (LHCs), a family of integral membrane proteins that coordinate chlorophylls and carotenoids. In vivo, these proteins are folded with pigments to form complexes which are inserted in the thylakoid membrane of the chloroplast. The high similarity in the chemical and physical properties of the members of the family, together with the fact that they can easily lose pigments during isolation, makes their purification in a native state challenging. An alternative approach to obtain homogeneous preparations of LHCs was developed by Plumley and Schmidt in 19871, who showed that it was possible to reconstitute these complexes in vitro starting from purified pigments and unfolded apoproteins, resulting in complexes with properties very similar to that of native complexes. This opened the way to the use of bacterial expressed recombinant proteins for in vitro reconstitution. The reconstitution method is powerful for various reasons: (1) pure preparations of individual complexes can be obtained, (2) pigment composition can be controlled to assess their contribution to structure and function, (3) recombinant proteins can be mutated to study the functional role of the individual residues (e.g., pigment binding sites) or protein domain (e.g., protein-protein interaction, folding). This method has been optimized in several laboratories and applied to most of the light-harvesting complexes. The protocol described here details the method of reconstituting light-harvesting complexes in vitro currently used in our laboratory,and examples describing applications of the method are provided. PMID:25350712

  3. Molecular basis of photoprotection and control of photosynthetic light-harvesting.

    PubMed

    Pascal, Andrew A; Liu, Zhenfeng; Broess, Koen; van Oort, Bart; van Amerongen, Herbert; Wang, Chao; Horton, Peter; Robert, Bruno; Chang, Wenrui; Ruban, Alexander

    2005-07-01

    In order to maximize their use of light energy in photosynthesis, plants have molecules that act as light-harvesting antennae, which collect light quanta and deliver them to the reaction centres, where energy conversion into a chemical form takes place. The functioning of the antenna responds to the extreme changes in the intensity of sunlight encountered in nature. In shade, light is efficiently harvested in photosynthesis. However, in full sunlight, much of the energy absorbed is not needed and there are vitally important switches to specific antenna states, which safely dissipate the excess energy as heat. This is essential for plant survival, because it provides protection against the potential photo-damage of the photosynthetic membrane. But whereas the features that establish high photosynthetic efficiency have been highlighted, almost nothing is known about the molecular nature of the dissipative states. Recently, the atomic structure of the major plant light-harvesting antenna protein, LHCII, has been determined by X-ray crystallography. Here we demonstrate that this is the structure of a dissipative state of LHCII. We present a spectroscopic analysis of this crystal form, and identify the specific changes in configuration of its pigment population that give LHCII the intrinsic capability to regulate energy flow. This provides a molecular basis for understanding the control of photosynthetic light-harvesting.

  4. Environmentally modulated phosphorylation and dynamics of proteins in photosynthetic membranes.

    PubMed

    Vener, Alexander V

    2007-06-01

    Recent advances in vectorial proteomics of protein domains exposed to the surface of photosynthetic thylakoid membranes of plants and the green alga Chlamydomonas reinhardtii allowed mapping of in vivo phosphorylation sites in integral and peripheral membrane proteins. In plants, significant changes of thylakoid protein phosphorylation are observed in response to stress, particularly in photosystem II under high light or high temperature stress. Thylakoid protein phosphorylation in the algae is much more responsive to the ambient redox and light conditions, as well as to CO(2) availability. The light-dependent multiple and differential phosphorylation of CP29 linker protein in the green algae is suggested to control photosynthetic state transitions and uncoupling of light harvesting proteins from photosystem II under high light. The similar role for regulation of the dynamic distribution of light harvesting proteins in plants is proposed for the TSP9 protein, which together with other recently discovered peripheral proteins undergoes specific environment- and redox-dependent phosphorylation at the thylakoid surface. This review focuses on the environmentally modulated reversible phosphorylation of thylakoid proteins related to their membrane dynamics and affinity towards particular photosynthetic protein complexes. PMID:17184728

  5. Principles of light harvesting from single photosynthetic complexes

    PubMed Central

    Schlau-Cohen, G. S.

    2015-01-01

    Photosynthetic systems harness sunlight to power most life on Earth. In the initial steps of photosynthetic light harvesting, absorbed energy is converted to chemical energy with near-unity quantum efficiency. This is achieved by an efficient, directional and regulated flow of energy through a network of proteins. Here, we discuss the following three key principles of this flow and of photosynthetic light harvesting: thermal fluctuations of the protein structure; intrinsic conformational switches with defined functional consequences; and environmentally triggered conformational switches. Through these principles, photosynthetic systems balance two types of operational costs: metabolic costs, or the cost of maintaining and running the molecular machinery, and opportunity costs, or the cost of losing any operational time. Understanding how the molecular machinery and dynamics are designed to balance these costs may provide a blueprint for improved artificial light-harvesting devices. With a multi-disciplinary approach combining knowledge of biology, this blueprint could lead to low-cost and more effective solar energy conversion. Photosynthetic systems achieve widespread light harvesting across the Earth's surface; in the face of our growing energy needs, this is functionality we need to replicate, and perhaps emulate. PMID:26052423

  6. Principles of light harvesting from single photosynthetic complexes.

    PubMed

    Schlau-Cohen, G S

    2015-06-01

    Photosynthetic systems harness sunlight to power most life on Earth. In the initial steps of photosynthetic light harvesting, absorbed energy is converted to chemical energy with near-unity quantum efficiency. This is achieved by an efficient, directional and regulated flow of energy through a network of proteins. Here, we discuss the following three key principles of this flow and of photosynthetic light harvesting: thermal fluctuations of the protein structure; intrinsic conformational switches with defined functional consequences; and environmentally triggered conformational switches. Through these principles, photosynthetic systems balance two types of operational costs: metabolic costs, or the cost of maintaining and running the molecular machinery, and opportunity costs, or the cost of losing any operational time. Understanding how the molecular machinery and dynamics are designed to balance these costs may provide a blueprint for improved artificial light-harvesting devices. With a multi-disciplinary approach combining knowledge of biology, this blueprint could lead to low-cost and more effective solar energy conversion. Photosynthetic systems achieve widespread light harvesting across the Earth's surface; in the face of our growing energy needs, this is functionality we need to replicate, and perhaps emulate. PMID:26052423

  7. Energizing the light harvesting antenna: Insight from CP29.

    PubMed

    Ioannidis, Nikolaos E; Papadatos, Sotiris; Daskalakis, Vangelis

    2016-10-01

    How do plants cope with excess light energy? Crop health and stress tolerance are governed by molecular photoprotective mechanisms. Protective exciton quenching in plants is activated by membrane energization, via unclear conformational changes in proteins called antennas. Here we show that pH and salt gradients stimulate the response of such an antenna under low and high energization by all-atom Molecular Dynamics Simulations. Novel insight establishes that helix-5 (H5) conformation in CP29 from spinach is regulated by chemiosmotic factors. This is selectively correlated with the chl-614 macrocycle deformation and interactions with nearby pigments, that could suggest a role in plant photoprotection. Adding to the significance of our findings, H5 domain is conserved among five antennas (LHCB1-5). These results suggest that light harvesting complexes of Photosystem II, one of the most abundant proteins on earth, can sense chemiosmotic gradients via their H5 domains in an upgraded role from a solar detector to also a chemiosmotic sensor. PMID:27438094

  8. Energizing the light harvesting antenna: Insight from CP29.

    PubMed

    Ioannidis, Nikolaos E; Papadatos, Sotiris; Daskalakis, Vangelis

    2016-10-01

    How do plants cope with excess light energy? Crop health and stress tolerance are governed by molecular photoprotective mechanisms. Protective exciton quenching in plants is activated by membrane energization, via unclear conformational changes in proteins called antennas. Here we show that pH and salt gradients stimulate the response of such an antenna under low and high energization by all-atom Molecular Dynamics Simulations. Novel insight establishes that helix-5 (H5) conformation in CP29 from spinach is regulated by chemiosmotic factors. This is selectively correlated with the chl-614 macrocycle deformation and interactions with nearby pigments, that could suggest a role in plant photoprotection. Adding to the significance of our findings, H5 domain is conserved among five antennas (LHCB1-5). These results suggest that light harvesting complexes of Photosystem II, one of the most abundant proteins on earth, can sense chemiosmotic gradients via their H5 domains in an upgraded role from a solar detector to also a chemiosmotic sensor.

  9. Topological analysis of the Rhodobacter capsulatus PucC protein and effects of C-terminal deletions on light-harvesting complex II.

    PubMed Central

    LeBlanc, H N; Beatty, J T

    1996-01-01

    A theoretical model for the cytoplasmic membrane topology of the Rhodobacter capsulatus PucC protein was derived and tested experimentally with pucC'::pho'A gene fusions. The alkaline phosphatase (AP) activities of selected fusions were assayed, and the resultant pattern of high and low activity was compared with that of the theoretical model. High AP activity correlated well with fusion joints located in regions predicted to be periplasmic, and most fusions in predicted cytoplasmic loops yield approximately 1/20th as much activity. Replacement of pho'A with lac'Z in nine of the fusions confirmed the topology, as beta-galactosidase activities were generally reciprocal to the corresponding AP activity. On the basis of the theoretical analysis and the information provided by the activities of fusions, a model for PucC topology in which there are 12 membrane-spanning segments and both the N and C termini are located in the cytoplasm is proposed. Translationally out-of-frame pucC::phoA fusions were expressed in an R. capsulatus delta pucC strain. None of the fusions missing only one or two of the proposed C-terminal transmembrane segments restored the wild-type phenotype, suggesting that the C terminus of PucC is important for function. PMID:8759841

  10. The terminal phycobilisome emitter, LCM: A light-harvesting pigment with a phytochrome chromophore.

    PubMed

    Tang, Kun; Ding, Wen-Long; Höppner, Astrid; Zhao, Cheng; Zhang, Lun; Hontani, Yusaku; Kennis, John T M; Gärtner, Wolfgang; Scheer, Hugo; Zhou, Ming; Zhao, Kai-Hong

    2015-12-29

    Photosynthesis relies on energy transfer from light-harvesting complexes to reaction centers. Phycobilisomes, the light-harvesting antennas in cyanobacteria and red algae, attach to the membrane via the multidomain core-membrane linker, L(CM). The chromophore domain of L(CM) forms a bottleneck for funneling the harvested energy either productively to reaction centers or, in case of light overload, to quenchers like orange carotenoid protein (OCP) that prevent photodamage. The crystal structure of the solubly modified chromophore domain from Nostoc sp. PCC7120 was resolved at 2.2 Å. Although its protein fold is similar to the protein folds of phycobiliproteins, the phycocyanobilin (PCB) chromophore adopts ZZZssa geometry, which is unknown among phycobiliproteins but characteristic for sensory photoreceptors (phytochromes and cyanobacteriochromes). However, chromophore photoisomerization is inhibited in L(CM) by tight packing. The ZZZssa geometry of the chromophore and π-π stacking with a neighboring Trp account for the functionally relevant extreme spectral red shift of L(CM). Exciton coupling is excluded by the large distance between two PCBs in a homodimer and by preservation of the spectral features in monomers. The structure also indicates a distinct flexibility that could be involved in quenching. The conclusions from the crystal structure are supported by femtosecond transient absorption spectra in solution. PMID:26669441

  11. The terminal phycobilisome emitter, LCM: A light-harvesting pigment with a phytochrome chromophore.

    PubMed

    Tang, Kun; Ding, Wen-Long; Höppner, Astrid; Zhao, Cheng; Zhang, Lun; Hontani, Yusaku; Kennis, John T M; Gärtner, Wolfgang; Scheer, Hugo; Zhou, Ming; Zhao, Kai-Hong

    2015-12-29

    Photosynthesis relies on energy transfer from light-harvesting complexes to reaction centers. Phycobilisomes, the light-harvesting antennas in cyanobacteria and red algae, attach to the membrane via the multidomain core-membrane linker, L(CM). The chromophore domain of L(CM) forms a bottleneck for funneling the harvested energy either productively to reaction centers or, in case of light overload, to quenchers like orange carotenoid protein (OCP) that prevent photodamage. The crystal structure of the solubly modified chromophore domain from Nostoc sp. PCC7120 was resolved at 2.2 Å. Although its protein fold is similar to the protein folds of phycobiliproteins, the phycocyanobilin (PCB) chromophore adopts ZZZssa geometry, which is unknown among phycobiliproteins but characteristic for sensory photoreceptors (phytochromes and cyanobacteriochromes). However, chromophore photoisomerization is inhibited in L(CM) by tight packing. The ZZZssa geometry of the chromophore and π-π stacking with a neighboring Trp account for the functionally relevant extreme spectral red shift of L(CM). Exciton coupling is excluded by the large distance between two PCBs in a homodimer and by preservation of the spectral features in monomers. The structure also indicates a distinct flexibility that could be involved in quenching. The conclusions from the crystal structure are supported by femtosecond transient absorption spectra in solution.

  12. The terminal phycobilisome emitter, LCM: A light-harvesting pigment with a phytochrome chromophore

    PubMed Central

    Tang, Kun; Ding, Wen-Long; Höppner, Astrid; Zhao, Cheng; Zhang, Lun; Hontani, Yusaku; Kennis, John T. M.; Gärtner, Wolfgang; Scheer, Hugo; Zhou, Ming; Zhao, Kai-Hong

    2015-01-01

    Photosynthesis relies on energy transfer from light-harvesting complexes to reaction centers. Phycobilisomes, the light-harvesting antennas in cyanobacteria and red algae, attach to the membrane via the multidomain core-membrane linker, LCM. The chromophore domain of LCM forms a bottleneck for funneling the harvested energy either productively to reaction centers or, in case of light overload, to quenchers like orange carotenoid protein (OCP) that prevent photodamage. The crystal structure of the solubly modified chromophore domain from Nostoc sp. PCC7120 was resolved at 2.2 Å. Although its protein fold is similar to the protein folds of phycobiliproteins, the phycocyanobilin (PCB) chromophore adopts ZZZssa geometry, which is unknown among phycobiliproteins but characteristic for sensory photoreceptors (phytochromes and cyanobacteriochromes). However, chromophore photoisomerization is inhibited in LCM by tight packing. The ZZZssa geometry of the chromophore and π-π stacking with a neighboring Trp account for the functionally relevant extreme spectral red shift of LCM. Exciton coupling is excluded by the large distance between two PCBs in a homodimer and by preservation of the spectral features in monomers. The structure also indicates a distinct flexibility that could be involved in quenching. The conclusions from the crystal structure are supported by femtosecond transient absorption spectra in solution. PMID:26669441

  13. Comparison of the rate constants for energy transfer in the light-harvesting protein, C-phycocyanin, calculated from Foerster`s theory and experimentally measured by time-resolved fluorescence spectroscopy

    SciTech Connect

    Debreczeny, M.P.

    1994-05-01

    We have measured and assigned rate constants for energy transfer between chromophores in the light-harvesting protein C-phycocyanin (PC), in the monomeric and trimeric aggregation states, isolated from Synechococcus sp. PCC 7002. In order to compare the measured rate constants with those predicted by Fdrster`s theory of inductive resonance in the weak coupling limit, we have experimentally resolved several properties of the three chromophore types ({beta}{sub 155} {alpha}{sub 84}, {beta}{sub 84}) found in PC monomers, including absorption and fluorescence spectra, extinction coefficients, fluorescence quantum yields, and fluorescence lifetimes. The cpcB/C155S mutant, whose PC is missing the {beta}{sub 155} chromophore, was, useful in effecting the resolution of the chromophore properties and in assigning the experimentally observed rate constants for energy transfer to specific pathways.

  14. Interference lithographic nanopatterning of plant and bacterial light-harvesting complexes on gold substrates

    PubMed Central

    Patole, Samson; Vasilev, Cvetelin; El-Zubir, Osama; Wang, Lin; Johnson, Matthew P.; Cadby, Ashley J.; Leggett, Graham J.; Hunter, C. Neil

    2015-01-01

    We describe a facile approach for nanopatterning of photosynthetic light-harvesting complexes over macroscopic areas, and use optical spectroscopy to demonstrate retention of native properties by both site-specifically and non-specifically attached photosynthetic membrane proteins. A Lloyd's mirror dual-beam interferometer was used to expose self-assembled monolayers of amine-terminated alkylthiolates on gold to laser irradiation. Following exposure, photo-oxidized adsorbates were replaced by oligo(ethylene glycol) terminated thiols, and the remaining intact amine-functionalized regions were used for attachment of the major light-harvesting chlorophyll–protein complex from plants, LHCII. These amine patterns could be derivatized with nitrilotriacetic acid (NTA), so that polyhistidine-tagged bacteriochlorophyll–protein complexes from phototrophic bacteria could be attached with a defined surface orientation. By varying parameters such as the angle between the interfering beams and the laser irradiation dose, it was possible to vary the period and widths of NTA and amine-functionalized lines on the surfaces; periods varied from 1200 to 240 nm and linewidths as small as 60 nm (λ/4) were achieved. This level of control over the surface chemistry was reflected in the surface topology of the protein nanostructures imaged by atomic force microscopy; fluorescence imaging and spectral measurements demonstrated that the surface-attached proteins had retained their native functionality. PMID:26464784

  15. In Vivo Identification of Photosystem II Light Harvesting Complexes Interacting with PHOTOSYSTEM II SUBUNIT S.

    PubMed

    Gerotto, Caterina; Franchin, Cinzia; Arrigoni, Giorgio; Morosinotto, Tomas

    2015-08-01

    Light is the primary energy source for photosynthetic organisms, but in excess, it can generate reactive oxygen species and lead to cell damage. Plants evolved multiple mechanisms to modulate light use efficiency depending on illumination intensity to thrive in a highly dynamic natural environment. One of the main mechanisms for protection from intense illumination is the dissipation of excess excitation energy as heat, a process called nonphotochemical quenching. In plants, nonphotochemical quenching induction depends on the generation of a pH gradient across thylakoid membranes and on the presence of a protein called PHOTOSYSTEM II SUBUNIT S (PSBS). Here, we generated Physcomitrella patens lines expressing histidine-tagged PSBS that were exploited to purify the native protein by affinity chromatography. The mild conditions used in the purification allowed copurifying PSBS with its interactors, which were identified by mass spectrometry analysis to be mainly photosystem II antenna proteins, such as LIGHT-HARVESTING COMPLEX B (LHCB). PSBS interaction with other proteins appears to be promiscuous and not exclusive, although the major proteins copurified with PSBS were components of the LHCII trimers (LHCB3 and LHCBM). These results provide evidence of a physical interaction between specific photosystem II light-harvesting complexes and PSBS in the thylakoids, suggesting that these subunits are major players in heat dissipation of excess energy.

  16. Distinct roles of the photosystem II protein PsbS and zeaxanthin in the regulation of light harvesting in plants revealed by fluorescence lifetime snapshots

    DOE PAGES

    Sylak-Glassman, Emily J.; Malnoë, Alizée; De Re, Eleonora; Brooks, Matthew D.; Fischer, Alexandra Lee; Niyogi, Krishna K.; Fleming, Graham R.

    2014-10-01

    The photosystem II (PSII) protein PsbS and the enzyme violaxanthin deepoxidase (VDE) are known to influence the dynamics of energy-dependent quenching (qE), the component of nonphotochemical quenching (NPQ) that allows plants to respond to fast fluctuations in light intensity. Although the absence of PsbS and VDE has been shown to change the amount of quenching, there have not been any measurements that can detect whether the presence of these proteins alters the type of quenching that occurs. The chlorophyll fluorescence lifetime probes the excited-state chlorophyll relaxation dynamics and can be used to determine the amount of quenching as well asmore » whether two different genotypes with the same amount of NPQ have similar dynamics of excited-state chlorophyll relaxation. We measured the fluorescence lifetimes on whole leaves of Arabidopsis thaliana throughout the induction and relaxation of NPQ for wild type and the qE mutants, npq4, which lacks PsbS; npq1, which lacks VDE and cannot convert violaxanthin to zeaxanthin; and npq1 npq4, which lacks both VDE and PsbS. These measurements show that although PsbS changes the amount of quenching and the rate at which quenching turns on, it does not affect the relaxation dynamics of excited chlorophyll during quenching. In addition, the data suggest that PsbS responds not only to ΔpH but also to the Δψ across the thylakoid membrane. In contrast, the presence of VDE, which is necessary for the accumulation of zeaxanthin, affects the excited-state chlorophyll relaxation dynamics.« less

  17. Distinct roles of the photosystem II protein PsbS and zeaxanthin in the regulation of light harvesting in plants revealed by fluorescence lifetime snapshots

    SciTech Connect

    Sylak-Glassman, Emily J.; Malnoë, Alizée; De Re, Eleonora; Brooks, Matthew D.; Fischer, Alexandra Lee; Niyogi, Krishna K.; Fleming, Graham R.

    2014-10-01

    The photosystem II (PSII) protein PsbS and the enzyme violaxanthin deepoxidase (VDE) are known to influence the dynamics of energy-dependent quenching (qE), the component of nonphotochemical quenching (NPQ) that allows plants to respond to fast fluctuations in light intensity. Although the absence of PsbS and VDE has been shown to change the amount of quenching, there have not been any measurements that can detect whether the presence of these proteins alters the type of quenching that occurs. The chlorophyll fluorescence lifetime probes the excited-state chlorophyll relaxation dynamics and can be used to determine the amount of quenching as well as whether two different genotypes with the same amount of NPQ have similar dynamics of excited-state chlorophyll relaxation. We measured the fluorescence lifetimes on whole leaves of Arabidopsis thaliana throughout the induction and relaxation of NPQ for wild type and the qE mutants, npq4, which lacks PsbS; npq1, which lacks VDE and cannot convert violaxanthin to zeaxanthin; and npq1 npq4, which lacks both VDE and PsbS. These measurements show that although PsbS changes the amount of quenching and the rate at which quenching turns on, it does not affect the relaxation dynamics of excited chlorophyll during quenching. In addition, the data suggest that PsbS responds not only to ΔpH but also to the Δψ across the thylakoid membrane. In contrast, the presence of VDE, which is necessary for the accumulation of zeaxanthin, affects the excited-state chlorophyll relaxation dynamics.

  18. Chromophore-chromophore and chromophore-protein interactions in monomeric light-harvesting complex II of green plants studied by spectral hole burning and fluorescence line narrowing.

    PubMed

    Pieper, Jörg; Rätsep, Margus; Irrgang, Klaus-Dieter; Freiberg, Arvi

    2009-08-01

    Persistent nonphotochemical hole burning and delta-FLN spectra obtained at 4.5 K are reported for monomeric chlorophyll (Chl) a/b light-harvesting complexes of photosystem II (LHC II) of green plants. The hole burned spectra of monomeric LHC II appear to be similar to those obtained before for trimeric LHC II (Pieper et al. J. Phys. Chem. B 1999, 103, 2412). They are composed of three main features: (i) a homogeneously broadened zero-phonon hole coincident with the burn wavelength, (ii) an intense, broad hole in the vicinity of approximately 680 nm as a result of efficient excitation energy transfer to a low-energy trap state, and (iii) a satellite hole at approximately 649 nm which is correlated with the low-energy 680 nm hole. Zero-phonon hole action spectroscopy reveals that the low-energy absorption band is located at 679.6 nm and possesses a width of approximately 110 cm(-1) which is predominantly due to inhomogeneous broadening at 4.5 K. The electron-phonon coupling of the above-mentioned low-energy state(s) is weak with a Huang-Rhys factor S in the order of 0.6 and a peak phonon frequency (omega(m)) of approximately 22 cm(-1) within a broad and strongly asymmetric one-phonon profile. The resulting Stokes shift 2S omega(m) of approximately 26.4 cm(-1) readily explains the position of the fluorescence origin band at 680.8 nm. Thus, we conclude that the 679.6 nm state(s) is (are) the fluorescent state(s) of monomeric LHC II at 4.5 K. The absorption intensity of the lowest Q(y) state is shown to roughly correspond to that of one out of the eight Chl a molecules bound in the monomeric subunit. In addition, the satellite hole structure produced by hole burning within the 679.6 nm state is weak with only one shallow satellite hole observed in the Chl b spectral range at 648.8 nm. These results suggest that the 679.6 nm state is widely localized on a Chl a molecule, which may belong to a Chl a/b heterodimer. These characteristics are different from those expected

  19. Macroorganization of Chlorophyll a/b light-harvesting complex in thylakoids and aggregates: information from circular differential scattering

    SciTech Connect

    Garab, G.; Faludi-Daniel, A.; Sutherland, J.C.; Hind, G.

    1988-04-05

    Circular dichroism (CD) and magnetic circular dichroism (MCD) spectra were recorded for spinach thylakoids and for isolated, aggregated chlorophyll a/b light-harvesting pigment-protein complex, in random and magnetically aligned states of orientation at room and low temperatures. The shape and magnitude of the CD signal of most bands strongly depended on the orientation of the thylakoid membranes or the aggregated pigment-protein complex. In both thylakoids and aggregated light-harvesting complexes, however, the MCD spectra of the two different orientations were almost identical. Random and magnetically aligned samples exhibited anomalous, large CD signals outside the bands of pigment absorbance. Lack of similarity between the corresponding MCD and CD spectra showed that the large CD signals are not produced as a distortion of CD of absorbance by light scattering. Instead, these anomalous spectral features are believed to originate in differential selective scattering of circularly polarized light. The results lead to the conclusion that the light-harvesting pigment-protein complex in thylakoid grana forms a helical macroarray with dimensions commensurate with the wavelengths of the anomalous circular dichroism signals. A hypothesis is put forward suggesting a role for these macrodomains in granal organization.

  20. Zeaxanthin Radical Cation Formation in Minor Light-Harvesting Complexes of Higher Plant Antenna

    SciTech Connect

    Avenson, Thomas H.; Ahn, Tae Kyu; Zigmantas, Donatas; Niyogi, Krishna K.; Li, Zhirong; Ballottari, Matteo; Bassi, Roberto; Fleming, Graham R.

    2008-01-31

    Previous work on intact thylakoid membranes showed that transient formation of a zeaxanthin radical cation was correlated with regulation of photosynthetic light-harvesting via energy-dependent quenching. A molecular mechanism for such quenching was proposed to involve charge transfer within a chlorophyll-zeaxanthin heterodimer. Using near infrared (880-1100 nm) transient absorption spectroscopy, we demonstrate that carotenoid (mainly zeaxanthin) radical cation generation occurs solely in isolated minor light-harvesting complexes that bind zeaxanthin, consistent with the engagement of charge transfer quenching therein. We estimated that less than 0.5percent of the isolated minor complexes undergo charge transfer quenching in vitro, whereas the fraction of minor complexes estimated to be engaged in charge transfer quenching in isolated thylakoids was more than 80 times higher. We conclude that minor complexes which bind zeaxanthin are sites of charge transfer quenching in vivo and that they can assume Non-quenching and Quenching conformations, the equilibrium LHC(N)<--> LHC(Q) of which is modulated by the transthylakoid pH gradient, the PsbS protein, and protein-protein interactions.

  1. Solar cells incorporating light harvesting arrays

    DOEpatents

    Lindsey, Jonathan S.; Meyer, Gerald J.

    2002-01-01

    A solar cell incorporates a light harvesting array that comprises: (a) a first substrate comprising a first electrode; and (b) a layer of light harvesting rods electrically coupled to the first electrode, each of the light harvesting rods comprising a polymer of Formula I: X.sup.1.paren open-st.X.sup.m+1).sub.m (I) wherein m is at least 1, and may be from two, three or four to 20 or more; X.sup.1 is a charge separation group (and preferably a porphyrinic macrocycle, which may be one ligand of a double-decker sandwich compound) having an excited-state of energy equal to or lower than that of X.sup.2 ; and X.sup.2 through X.sup.m+1 are chromophores (and again are preferably porphyrinic macrocycles).

  2. Solar cells incorporating light harvesting arrays

    DOEpatents

    Lindsey, Jonathan S.; Meyer, Gerald J.

    2003-07-22

    A solar cell incorporates a light harvesting array that comprises: (a) a first substrate comprising a first electrode; and (b) a layer of light harvesting rods electrically coupled to the first electrode, each of the light harvesting rods comprising a polymer of Formula I: ##EQU1## wherein m is at least 1, and may be from two, three or four to 20 or more; X.sup.1 is a charge separation group (and preferably a porphyrinic macrocycle, which may be one ligand of a double-decker sandwich compound) having an excited-state of energy equal to or lower than that of X.sup.2 ; and X.sup.2 through X.sup.m+1 are chromophores (and again are preferably porphyrinic macrocycles).

  3. Natural strategies for photosynthetic light harvesting.

    PubMed

    Croce, Roberta; van Amerongen, Herbert

    2014-07-01

    Photosynthetic organisms are crucial for life on Earth as they provide food and oxygen and are at the basis of most energy resources. They have a large variety of light-harvesting strategies that allow them to live nearly everywhere where sunlight can penetrate. They have adapted their pigmentation to the spectral composition of light in their habitat, they acclimate to slowly varying light intensities and they rapidly respond to fast changes in light quality and quantity. This is particularly important for oxygen-producing organisms because an overdose of light in combination with oxygen can be lethal. Rapid progress is being made in understanding how different organisms maximize light harvesting and minimize deleterious effects. Here we summarize the latest findings and explain the main design principles used in nature. The available knowledge can be used for optimizing light harvesting in both natural and artificial photosynthesis to improve light-driven production processes.

  4. Structure of Light-Harvesting Aggregates in Individual Chlorosomes.

    PubMed

    Günther, Lisa M; Jendrny, Marc; Bloemsma, Erik A; Tank, Marcus; Oostergetel, Gert T; Bryant, Donald A; Knoester, Jasper; Köhler, Jürgen

    2016-06-23

    Among all photosynthetic organisms, green bacteria have evolved one of the most efficient light-harvesting antenna, the chlorosome, that contains hundreds of thousands of bacteriochlorophyll molecules, allowing these bacteria to grow photosynthetically by absorbing only a few photons per bacteriochlorophyll molecule per day. In contrast to other photosynthetic light-harvesting antenna systems, for which a protein scaffold imposes the proper positioning of the chromophores with respect to each other, in chlorosomes, this is accomplished solely by self-assembly. This has aroused enormous interest in the structure-function relations of these assemblies, as they can serve as blueprints for artificial light harvesting systems. In spite of these efforts, conclusive structural information is not available yet, reflecting the sample heterogeneity inherent to the natural system. Here we combine mutagenesis, polarization-resolved single-particle fluorescence-excitation spectroscopy, cryo-electron microscopy, and theoretical modeling to study the chlorosomes of the green sulfur bacterium Chlorobaculum tepidum. We demonstrate that only the combination of these techniques yields unambiguous information on the structure of the bacteriochlorophyll aggregates within the chlorosomes. Moreover, we provide a quantitative estimate of the curvature variation of these aggregates that explains ongoing debates concerning the chlorosome structure. PMID:27240572

  5. Optimal Energy Transfer in Light-Harvesting Systems.

    PubMed

    Chen, Lipeng; Shenai, Prathamesh; Zheng, Fulu; Somoza, Alejandro; Zhao, Yang

    2015-08-20

    Photosynthesis is one of the most essential biological processes in which specialized pigment-protein complexes absorb solar photons, and with a remarkably high efficiency, guide the photo-induced excitation energy toward the reaction center to subsequently trigger its conversion to chemical energy. In this work, we review the principles of optimal energy transfer in various natural and artificial light harvesting systems. We begin by presenting the guiding principles for optimizing the energy transfer efficiency in systems connected to dissipative environments, with particular attention paid to the potential role of quantum coherence in light harvesting systems. We will comment briefly on photo-protective mechanisms in natural systems that ensure optimal functionality under varying ambient conditions. For completeness, we will also present an overview of the charge separation and electron transfer pathways in reaction centers. Finally, recent theoretical and experimental progress on excitation energy transfer, charge separation, and charge transport in artificial light harvesting systems is delineated, with organic solar cells taken as prime examples.

  6. Optimal Energy Transfer in Light-Harvesting Systems.

    PubMed

    Chen, Lipeng; Shenai, Prathamesh; Zheng, Fulu; Somoza, Alejandro; Zhao, Yang

    2015-01-01

    Photosynthesis is one of the most essential biological processes in which specialized pigment-protein complexes absorb solar photons, and with a remarkably high efficiency, guide the photo-induced excitation energy toward the reaction center to subsequently trigger its conversion to chemical energy. In this work, we review the principles of optimal energy transfer in various natural and artificial light harvesting systems. We begin by presenting the guiding principles for optimizing the energy transfer efficiency in systems connected to dissipative environments, with particular attention paid to the potential role of quantum coherence in light harvesting systems. We will comment briefly on photo-protective mechanisms in natural systems that ensure optimal functionality under varying ambient conditions. For completeness, we will also present an overview of the charge separation and electron transfer pathways in reaction centers. Finally, recent theoretical and experimental progress on excitation energy transfer, charge separation, and charge transport in artificial light harvesting systems is delineated, with organic solar cells taken as prime examples. PMID:26307957

  7. Evolution of a light-harvesting protein by addition of new subunits and rearrangement of conserved elements: crystal structure of a cryptophyte phycoerythrin at 1.63-A resolution.

    PubMed

    Wilk, K E; Harrop, S J; Jankova, L; Edler, D; Keenan, G; Sharples, F; Hiller, R G; Curmi, P M

    1999-08-01

    Cryptophytes are unicellular photosynthetic algae that use a lumenally located light-harvesting system, which is distinct from the phycobilisome structure found in cyanobacteria and red algae. One of the key components of this system is water-soluble phycoerythrin (PE) 545 whose expression is enhanced by low light levels. The crystal structure of the heterodimeric alpha(1)alpha(2)betabeta PE 545 from the marine cryptophyte Rhodomonas CS24 has been determined at 1.63-A resolution. Although the beta-chain structure is similar to the alpha and beta chains of other known phycobiliproteins, the overall structure of PE 545 is novel with the alpha chains forming a simple extended fold with an antiparallel beta-ribbon followed by an alpha-helix. The two doubly linked beta50/beta61 chromophores (one on each beta subunit) are in van der Waals contact, suggesting that exciton-coupling mechanisms may alter their spectral properties. Each alpha subunit carries a covalently linked 15,16-dihydrobiliverdin chromophore that is likely to be the final energy acceptor. The architecture of the heterodimer suggests that PE 545 may dock to an acceptor protein via a deep cleft and that energy may be transferred via this intermediary protein to the reaction center.

  8. Biomolecular membrane protein crystallization

    NASA Astrophysics Data System (ADS)

    Reddy Bolla, Jani; Su, Chih-Chia; Yu, Edward W.

    2012-07-01

    Integral membrane proteins comprise approximately 30% of the sequenced genomes, and there is an immediate need for their high-resolution structural information. Currently, the most reliable approach to obtain these structures is X-ray crystallography. However, obtaining crystals of membrane proteins that diffract to high resolution appears to be quite challenging, and remains a major obstacle in structural determination. This brief review summarizes a variety of methodologies for use in crystallizing these membrane proteins. Hopefully, by introducing the available methods, techniques, and providing a general understanding of membrane proteins, a rational decision can be made about now to crystallize these complex materials.

  9. The size of the light-harvesting antenna of higher plant photosystem II is regulated by illumination intensity through transcription of antenna protein genes.

    PubMed

    Borisova-Mubarakshina, M M; Vetoshkina, D V; Rudenko, N N; Shirshikova, G N; Fedorchuk, T P; Naydov, I A; Ivanov, B N

    2014-06-01

    In arabidopsis plants, with an increase in illumination intensity during growth the extent of reduction of the plastoquinone pool in the photosynthetic electron transport chain increased, whereas the effective quantum yield of photosynthesis decreased. After 5 days of growth under high illumination intensity, these parameters in high light returned to values observed in "shade-adapted" plants in low light. During the same period, the size of the antenna decreased, correlating with a decrease in the amounts of proteins of peripheral pigment-protein complexes. It was found that the decrease in the amounts of these proteins occurred due to suppression of transcription of their genes.

  10. Investigation of Detergent Effects on the Solution Structure of Spinach Light Harvesting Complex II

    SciTech Connect

    Cardoso, Mateus B; Smolensky, Dmitriy; Heller, William T; O'Neill, Hugh Michael

    2010-01-01

    The properties of spinach light harvesting complex II (LHC II), stabilized in the detergents Triton X-100 (TX100) and n-Octyl-{beta}-D-Glucoside (BOG), were investigated by small-angle neutron scattering (SANS). The LHC II-BOG scattering curve overlaid well with the theoretical scattering curve generated from the crystal structure of LHC II indicating that the protein preparation was in its native functional state. On the other hand, the simulated LHC II curve deviated significantly from the LHC II-TX100 experimental data. Analysis by circular dichroism spectroscopy supported the SANS analysis and showed that LHC II-TX100 is inactivated. This investigation has implications for extracting and stabilizing photosynthetic membrane proteins for the development of biohybrid photoconversion devices.

  11. Investigation of detergent effects on the solution structure of spinach Light Harvesting Complex II

    NASA Astrophysics Data System (ADS)

    Cardoso, Mateus B.; Smolensky, Dmitriy; Heller, William T.; O'Neill, Hugh

    2010-11-01

    The properties of spinach light harvesting complex II (LHC II), stabilized in the detergents Triton X-100 (TX100) and n-Octyl-β-D-Glucoside (BOG), were investigated by small-angle neutron scattering (SANS). The LHC II-BOG scattering curve overlaid well with the theoretical scattering curve generated from the crystal structure of LHC II indicating that the protein preparation was in its native functional state. On the other hand, the simulated LHC II curve deviated significantly from the LHC II-TX100 experimental data. Analysis by circular dichroism spectroscopy supported the SANS analysis and showed that LHC II-TX100 is inactivated. This investigation has implications for extracting and stabilizing photosynthetic membrane proteins for the development of biohybrid photoconversion devices.

  12. Discrete cyclic porphyrin arrays as artificial light-harvesting antenna.

    PubMed

    Aratani, Naoki; Kim, Dongho; Osuka, Atsuhiro

    2009-12-21

    The importance of photosynthesis has driven researchers to seek ways to mimic its fundamental features in simplified systems. The absorption of a photon by light-harvesting (antenna) complexes made up of a large number of protein-embedded pigments initiates photosynthesis. Subsequently the many pigments within the antenna system shuttle that photon via an efficient excitation energy transfer (EET) until it encounters a reaction center. Since the 1995 discovery of the circularly arranged chromophoric assemblies in the crystal structure of light-harvesting antenna complex LH2 of purple bacteria Rps. Acidophila, many designs of light-harvesting antenna systems have focused on cyclic porphyrin wheels that allow for efficient EET. In this Account, we review recent research in our laboratories in the synthesis of covalently and noncovalently linked discrete cyclic porphyrin arrays as models of the photosynthetic light-harvesting antenna complexes. On the basis of the silver(I)-promoted oxidative coupling strategy, we have prepared a series of extremely long yet discrete meso-meso-linked porphyrin arrays and covalently linked large porphyrin rings. We examined the photophysical properties of these molecules using steady-state absorption, fluorescence, fluorescence lifetime, fluorescence anisotropy decay, and transient absorption measurements. Both the pump-power dependence on the femtosecond transient absorption and the transient absorption anisotropy decay profiles are directly related to the EET processes within the porphyrin rings. Within these structures, the exciton-exciton annihilation time and the polarization anisotropy rise time are well-described in terms of the Forster-type incoherent energy hopping model. In noncoordinating solvents such as CHCl(3), meso-pyridine-appended zinc(II) porphyrins and their meso-meso-linked dimers spontaneously assemble to form tetrameric porphyrin squares and porphyrin boxes, respectively. In the latter case, we have demonstrated

  13. A lycopene β-cyclase/lycopene ε-cyclase/light-harvesting complex-fusion protein from the green alga Ostreococcus lucimarinus can be modified to produce α-carotene and β-carotene at different ratios.

    PubMed

    Blatt, Andreas; Bauch, Matthias E; Pörschke, Yvonne; Lohr, Martin

    2015-05-01

    Biosynthesis of asymmetric carotenoids such as α-carotene and lutein in plants and green algae involves the two enzymes lycopene β-cyclase (LCYB) and lycopene ε-cyclase (LCYE). The two cyclases are closely related and probably resulted from an ancient gene duplication. While in most plants investigated so far the two cyclases are encoded by separate genes, prasinophyte algae of the order Mamiellales contain a single gene encoding a fusion protein comprised of LCYB, LCYE and a C-terminal light-harvesting complex (LHC) domain. Here we show that the lycopene cyclase fusion protein from Ostreococcus lucimarinus catalyzed the simultaneous formation of α-carotene and β-carotene when heterologously expressed in Escherichia coli. The stoichiometry of the two products in E. coli could be altered by gradual truncation of the C-terminus, suggesting that the LHC domain may be involved in modulating the relative activities of the two cyclase domains in the algae. Partial deletions of the linker region between the cyclase domains or replacement of one or both cyclase domains with the corresponding cyclases from the green alga Chlamydomonas reinhardtii resulted in pronounced shifts of the α-carotene-to-β-carotene ratio, indicating that both the relative activities of the cyclase domains and the overall structure of the fusion protein have a strong impact on the product stoichiometry. The possibility to tune the product ratio of the lycopene cyclase fusion protein from Mamiellales renders it useful for the biotechnological production of the asymmetric carotenoids α-carotene or lutein in bacteria or fungi.

  14. Excitation energy transfer and charge separation are affected in Arabidopsis thaliana mutants lacking light-harvesting chlorophyll a/b binding protein Lhcb3.

    PubMed

    Adamiec, Małgorzata; Gibasiewicz, Krzysztof; Luciński, Robert; Giera, Wojciech; Chełminiak, Przemysław; Szewczyk, Sebastian; Sipińska, Weronika; van Grondelle, Rienk; Jackowski, Grzegorz

    2015-12-01

    The composition of LHCII trimers as well as excitation energy transfer and charge separation in grana cores of Arabidopsis thaliana mutant lacking chlorophyll a/b binding protein Lhcb3 have been investigated and compared to those in wild-type plants. In grana cores of lhcb3 plants we observed increased amounts of Lhcb1 and Lhcb2 apoproteins per PSII core. The additional copies of Lhcb1 and Lhcb2 are expected to substitute for Lhcb3 in LHCII trimers M as well as in the LHCII "extra" pool, which was found to be modestly enlarged as a result of the absence of Lhcb3. Time-resolved fluorescence measurements reveal a deceleration of the fast phase of excitation dynamics in grana cores of the mutant by ~15 ps, whereas the average fluorescence lifetime is not significantly altered. Monte Carlo modeling predicts a slowing down of the mean hopping time and an increased stabilization of the primary charge separation in the mutant. Thus our data imply that absence of apoprotein Lhcb3 results in detectable differences in excitation energy transfer and charge separation.

  15. Molecular Factors Controlling Photosynthetic Light-Harvesting by Carotenoids

    PubMed Central

    Polívka, Tomáš; Frank, Harry A.

    2010-01-01

    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 photo-induced damage caused by excess light absorption. To carry out these functions in nature, carotenoids are bound in discrete pigment-protein complexes in close proximity to chlorophylls. A few 3D 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 making the 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

  16. Structures of membrane proteins

    PubMed Central

    Vinothkumar, Kutti R.; Henderson, Richard

    2010-01-01

    In reviewing the structures of membrane proteins determined up to the end of 2009, we present in words and pictures the most informative examples from each family. We group the structures together according to their function and architecture to provide an overview of the major principles and variations on the most common themes. The first structures, determined 20 years ago, were those of naturally abundant proteins with limited conformational variability, and each membrane protein structure determined was a major landmark. With the advent of complete genome sequences and efficient expression systems, there has been an explosion in the rate of membrane protein structure determination, with many classes represented. New structures are published every month and more than 150 unique membrane protein structures have been determined. This review analyses the reasons for this success, discusses the challenges that still lie ahead, and presents a concise summary of the key achievements with illustrated examples selected from each class. PMID:20667175

  17. Drugging Membrane Protein Interactions

    PubMed Central

    Yin, Hang; Flynn, Aaron D.

    2016-01-01

    The majority of therapeutics target membrane proteins, accessible on the surface of cells, to alter cellular signaling. Cells use membrane proteins to transduce signals into cells, transport ions and molecules, bind the cell to a surface or substrate, and catalyze reactions. Newly devised technologies allow us to drug conventionally “undruggable” regions of membrane proteins, enabling modulation of protein–protein, protein–lipid, and protein–nucleic acid interactions. In this review, we survey the state of the art in high-throughput screening and rational design in drug discovery, and we evaluate the advances in biological understanding and technological capacity that will drive pharmacotherapy forward against unorthodox membrane protein targets. PMID:26863923

  18. Light-harvesting complex Lhcb9 confers a green alga-type photosystem I supercomplex to the moss Physcomitrella patens.

    PubMed

    Iwai, Masakazu; Yokono, Makio; Kono, Masaru; Noguchi, Ko; Akimoto, Seiji; Nakano, Akihiko

    2015-01-19

    Light-harvesting complex (LHC) proteins in chloroplast thylakoid membranes not only transfer absorbed light energy to the two photosystems but also regulate the rate of energy transfer to avoid photodamage. Here we demonstrate that Lhcb9, a recently discovered LHC protein in the moss Physcomitrella patens, functions to connect LHC proteins with photosystem I (PSI), resulting in the formation of two different types of PSI supercomplexes in thylakoid membranes. We observed that the Lhcb9-containing PSI supercomplex is disassembled in response to excess light conditions. On the basis of our phylogenetic analysis, it appears that P. patens acquired Lhcb9 by horizontal gene transfer from the earlier green algal lineage, leading to the presence of both green alga-type and vascular plant-type PSI supercomplexes, which would have been crucial for conquering the dynamic environmental interface between aquatic and terrestrial conditions it faced during evolution.

  19. Rod-like nano-light harvester.

    PubMed

    Ling, Jun; Zheng, Zhicheng; Köhler, Anna; Müller, Axel H E

    2014-01-01

    Imitating the natural "energy cascade" architecture, we present a single-molecular rod-like nano-light harvester (NLH) based on a cylindrical polymer brush. Block copolymer side chains carrying (9,9-diethylfluoren-2-yl)methyl methacrylate units as light absorbing antennae (energy donors) are tethered to a linear polymer backbone containing 9-anthracenemethyl methacrylate units as emitting groups (energy acceptors). These NLHs exhibit very efficient energy absorption and transfer. Moreover, we manipulate the energy transfer by tuning the donor-acceptor distance.

  20. Regulation of photosystem I light harvesting by zeaxanthin.

    PubMed

    Ballottari, Matteo; Alcocer, Marcelo J P; D'Andrea, Cosimo; Viola, Daniele; Ahn, Tae Kyu; Petrozza, Annamaria; Polli, Dario; Fleming, Graham R; Cerullo, Giulio; Bassi, Roberto

    2014-06-10

    In oxygenic photosynthetic eukaryotes, the hydroxylated carotenoid zeaxanthin is produced from preexisting violaxanthin upon exposure to excess light conditions. Zeaxanthin binding to components of the photosystem II (PSII) antenna system has been investigated thoroughly and shown to help in the dissipation of excess chlorophyll-excited states and scavenging of oxygen radicals. However, the functional consequences of the accumulation of the light-harvesting complex I (LHCI) proteins in the photosystem I (PSI) antenna have remained unclarified so far. In this work we investigated the effect of zeaxanthin binding on photoprotection of PSI-LHCI by comparing preparations isolated from wild-type Arabidopsis thaliana (i.e., with violaxanthin) and those isolated from the A. thaliana nonphotochemical quenching 2 mutant, in which violaxanthin is replaced by zeaxanthin. Time-resolved fluorescence measurements showed that zeaxanthin binding leads to a previously unrecognized quenching effect on PSI-LHCI fluorescence. The efficiency of energy transfer from the LHCI moiety of the complex to the PSI reaction center was down-regulated, and an enhanced PSI resistance to photoinhibition was observed both in vitro and in vivo. Thus, zeaxanthin was shown to be effective in inducing dissipative states in PSI, similar to its well-known effect on PSII. We propose that, upon acclimation to high light, PSI-LHCI changes its light-harvesting efficiency by a zeaxanthin-dependent quenching of the absorbed excitation energy, whereas in PSII the stoichiometry of LHC antenna proteins per reaction center is reduced directly.

  1. Photophysics in single light-harvesting complexes II: from micelle to native nanodisks

    NASA Astrophysics Data System (ADS)

    Gruber, J. Michael; Scheidelaar, Stefan; van Roon, Henny; Dekker, Jan P.; Killian, J. Antoinette; van Grondelle, Rienk

    2016-02-01

    Most photosynthetic pigment-protein complexes of algae and higher plants are integral membrane proteins and are thus usually isolated in the presence of detergent to provide a hydrophobic interface and prevent aggregation. It was recently shown that the styrene maleic acid (SMA) copolymer can be used instead to solubilize and isolate protein complexes with their native lipid environment into nanodisk particles. We isolated LHCII complexes in SMA and compared their photophysics with trimeric LHCII complexes in β-DM detergent micelles to understand the effect of the native environment on the function of light-harvesting antennae. The triplet state kinetics and the calculated relative absorption cross section of single complexes indicate the successful isolation of trimeric complexes in SMA nanodisks, confirming the trimeric structure as the likely native configuration. The survival time of complexes before they photobleach is increased in SMA compared to detergent which might be explained by a stabilizing effect of the co-purified lipids in nanodisks. We furthermore find an unquenched fluorescence lifetime of 3.5 ns for LHCII in SMA nanodisks which coincides with detergent isolated complexes and notably differs from 2 ns typically found in native thylakoid structures. A large dynamic range of partially quenched complexes both in detergent micelles and lipid nanodisks is demonstrated by correlating the fluorescence lifetime with the intensity and likely reflects the conformational freedom of these complexes. This further supports the hypothesis that fluorescence intermittency is an intrinsic property of LHCII that may be involved in excess energy dissipation in native light-harvesting.

  2. Structural relationship in chloroplast membranes. Final report, August 1, 1977-August 31, 1980

    SciTech Connect

    1980-09-01

    Methodology has been developed for the isolation and characterization of pigment-proteins from chloroplast membranes. Characterization of these pigment-proteins has increased our understanding of mechanisms regulating the efficiency of photosynthetic light harvesting during photosynthesis. Incorporation of isolated pigment-protein complexes into model membranes has allowed simulation of grana stacks; these structural features of chloroplasts play a key role in maintaining appropriate interactions among light-harvesting assembles to regulate photosynthetic solar energy conversion.

  3. Quantum mechanical light harvesting mechanisms in photosynthesis

    NASA Astrophysics Data System (ADS)

    Scholes, Gregory

    2012-02-01

    More than 10 million billion photons of light strike a leaf each second. Incredibly, almost every red-coloured photon is captured by chlorophyll pigments and initiates steps to plant growth. Last year we reported that marine algae use quantum mechanics in order to optimize photosynthesis [1], a process essential to its survival. These and other insights from the natural world promise to revolutionize our ability to harness the power of the sun. In a recent review [2] we described the principles learned from studies of various natural antenna complexes and suggested how to utilize that knowledge to shape future technologies. We forecast the need to develop ways to direct and regulate excitation energy flow using molecular organizations that facilitate feedback and control--not easy given that the energy is only stored for a billionth of a second. In this presentation I will describe new results that explain the observation and meaning of quantum-coherent energy transfer. [4pt] [1] Elisabetta Collini, Cathy Y. Wong, Krystyna E. Wilk, Paul M. G. Curmi, Paul Brumer, and Gregory D. Scholes, ``Coherently wired light-harvesting in photosynthetic marine algae at ambient temperature'' Nature 463, 644-648 (2010).[0pt] [2] Gregory D. Scholes, Graham R. Fleming, Alexandra Olaya-Castro and Rienk van Grondelle, ``Lessons from nature about solar light harvesting'' Nature Chem. 3, 763-774 (2011).

  4. Heat- and light-induced detachment of the light harvesting complex from isolated photosystem I supercomplexes.

    PubMed

    Nellaepalli, Sreedhar; Zsiros, Ottó; Tóth, Tünde; Yadavalli, Venkateswarlu; Garab, Győző; Subramanyam, Rajagopal; Kovács, László

    2014-08-01

    In a previous study, using photosystem I enriched stroma thylakoid membrane vesicles, we have shown that the light harvesting complexes of this photosystem are prone to heat- and light-induced, thermo-optically driven detachment from the supercomplex [43]. We have also shown that the splitting of the supercomplex occurs in a gradual and specific manner, selectively affecting the different constituents of the antenna complexes. Here we further analyse these heat- and light-induced processes in isolated Photosystem I supercomplex using circular dichroism and 77K fluorescence emission spectroscopy and immuno blotting, and obtain further details on the sequence of events of the dissociation process as well as on the thermal stability of the different components. Our absorption and circular dichroism spectroscopy and immuno blotting data show that the dissociation of LHCI from PSI-LHCI supercomplex starts above 50°C. Also, the low temperature fluorescence emission spectra depicts decrease of maximum fluorescence emission at 730nm and an increase of the intensity at 685nm, and about 10nm blue-shifts, from 730 to 720nm and from 685 to 676nm, respectively, indicating the heat (50°C) induced detachment of LHCI from PSI core complexes. The reaction centre proteins are highly stable even at high temperatures. Lhca2 is more heat stable than the other light harvesting protein complexes of PSI, whereas Lhca4 and Lhca3 are rather labile. Combined heat and light treatments significantly enhances the disorganization of PSI-LHCI supercomplexes, indicating a thermo-optic mechanism, which might have significant role under combined heat and light stress conditions.

  5. Membrane protein secretases.

    PubMed Central

    Hooper, N M; Karran, E H; Turner, A J

    1997-01-01

    A diverse range of membrane proteins of Type 1 or Type II topology also occur as a circulating, soluble form. These soluble forms are often derived from the membrane form by proteolysis by a group of enzymes referred to collectively as 'secretases' or 'sheddases'. The cleavage generally occurs close to the extracellular face of the membrane, releasing physiologically active protein. This secretion process also provides a mechanism for down-regulating the protein at the cell surface. Examples of such post-translational proteolysis are seen in the Alzheimer's amyloid precursor protein, the vasoregulatory enzyme angiotensin converting enzyme, transforming growth factor-alpha, the tumour necrosis factor ligand and receptor superfamilies, certain cytokine receptors, and others. Since the proteins concerned are involved in pathophysiological processes such as neurodegeneration, apoptosis, oncogenesis and inflammation, the secretases could provide novel therapeutic targets. Recent characterization of these individual secretases has revealed common features, particularly sensitivity to certain metalloprotease inhibitors and upregulation of activity by phorbol esters. It is therefore likely that a closely related family of metallosecretases controls the surface expression of multiple integral membrane proteins. Current knowledge of the various secretases are compared in this Review, and strategies for cell-free assays of such proteases are outlined as a prelude to their ultimate purification and cloning. PMID:9020855

  6. Applications of textured surfaces for light harvesting

    NASA Astrophysics Data System (ADS)

    Cocilovo, Byron

    Surface textures add another dimension to optical design. They can be used to redirect light, isolate spectral bands, and enhance optical fields. They effectively take up no space, so can be applied to any optical surface -- from intermediary elements to substrates. Here I present three applications of textured surfaces for light harvesting. The first project places scattering textures inside a film that can be applied to windows to scatter infrared light towards solar cells at the edges. The collected energy is then used to power tinting films. The second project uses modular diffractive structures to increase the absorption in solar cells. Lastly, structured silver surfaces are used to enhance plasmonics fields and increase two-photon excitation fluorescence.

  7. The mobile thylakoid phosphoprotein TSP9 interacts with the light-harvesting complex II and the peripheries of both photosystems.

    PubMed

    Hansson, Maria; Dupuis, Tiphaine; Strömquist, Ragna; Andersson, Bertil; Vener, Alexander V; Carlberg, Inger

    2007-06-01

    The localization of the plant-specific thylakoid-soluble phosphoprotein of 9 kDa, TSP9, within the chloroplast thylakoid membrane of spinach has been established by the combined use of fractionation, immunoblotting, cross-linking, and mass spectrometry. TSP9 was found to be exclusively confined to the thylakoid membranes, where it is enriched in the stacked grana membrane domains. After mild solubilization of the membranes, TSP9 migrated together with the major light-harvesting antenna (LHCII) of photosystem II (PSII) and with PSII-LHCII supercomplexes upon separation of the protein complexes by either native gel electrophoresis or sucrose gradient centrifugation. Studies with a cleavable cross-linking agent revealed the interaction of TSP9 with both major and minor LHCII proteins as identified by mass spectrometric sequencing. Cross-linked complexes that in addition to TSP9 contain the peripheral PSII subunits CP29, CP26, and PsbS, which form the interface between LHCII and the PSII core, were found. Our observations also clearly suggest an interaction of TSP9 with photosystem I (PSI) as shown by both immunodetection and mass spectrometry. Sequencing identified the peripheral PSI subunits PsaL, PsaF, and PsaE, originating from cross-linked protein complexes of around 30 kDa that also contained TSP9. The distribution of TSP9 among the cross-linked forms was found to be sensitive to conditions such as light exposure. An association of TSP9 with LHCII as well as the peripheries of the photosystems suggests its involvement in regulation of photosynthetic light harvesting. PMID:17400553

  8. Long range excitonic transport in a biomimetic system inspired by the bacterial light-harvesting apparatus

    NASA Astrophysics Data System (ADS)

    Harel, Elad

    2012-05-01

    Photosynthesis, the process by which energy from sunlight drives cellular metabolism, relies on a unique organization of light-harvesting and reaction center complexes. Recently, the organization of light-harvesting LH2 complexes and dimeric reaction center-light-harvesting I-PufX core complexes in membranes of purple non-sulfur bacteria was revealed by atomic force microscopy [S. Bahatyrova et al., Nature (London) 430, 1058 (2004)]. Here, we discuss optimal exciton transfer in a biomimetic system closely modeled on the structure of LH2 and its organization within the membrane using a Markovian quantum model with dissipation and trapping added phenomenologically. In a deliberate manner, we neglect the high level detail of the bacterial light-harvesting complex and its interaction with the phonon bath in order to elucidate a set of design principles that may be incorporated in artificial pigment-scaffold constructs in a supramolecular assembly. We show that our scheme reproduces many of the most salient features found in their natural counterpart and may be largely explained by simple electrostatic considerations. Most importantly, we show that quantum effects act primarily to enforce robustness with respect to spatial and spectral disorder between and within complexes. The implications of such an arrangement are discussed in the context of biomimetic photosynthetic analogs capable of transferring energy efficiently across tens to hundreds of nanometers.

  9. Long range excitonic transport in a biomimetic system inspired by the bacterial light-harvesting apparatus

    SciTech Connect

    Harel, Elad

    2012-05-07

    Photosynthesis, the process by which energy from sunlight drives cellular metabolism, relies on a unique organization of light-harvesting and reaction center complexes. Recently, the organization of light-harvesting LH2 complexes and dimeric reaction center-light-harvesting I-PufX core complexes in membranes of purple non-sulfur bacteria was revealed by atomic force microscopy [S. Bahatyrova et al., Nature (London) 430, 1058 (2004)]. Here, we discuss optimal exciton transfer in a biomimetic system closely modeled on the structure of LH2 and its organization within the membrane using a Markovian quantum model with dissipation and trapping added phenomenologically. In a deliberate manner, we neglect the high level detail of the bacterial light-harvesting complex and its interaction with the phonon bath in order to elucidate a set of design principles that may be incorporated in artificial pigment-scaffold constructs in a supramolecular assembly. We show that our scheme reproduces many of the most salient features found in their natural counterpart and may be largely explained by simple electrostatic considerations. Most importantly, we show that quantum effects act primarily to enforce robustness with respect to spatial and spectral disorder between and within complexes. The implications of such an arrangement are discussed in the context of biomimetic photosynthetic analogs capable of transferring energy efficiently across tens to hundreds of nanometers.

  10. Multiscale model of light harvesting by photosystem II in plants.

    PubMed

    Amarnath, Kapil; Bennett, Doran I G; Schneider, Anna R; Fleming, Graham R

    2016-02-01

    The first step of photosynthesis in plants is the absorption of sunlight by pigments in the antenna complexes of photosystem II (PSII), followed by transfer of the nascent excitation energy to the reaction centers, where long-term storage as chemical energy is initiated. Quantum mechanical mechanisms must be invoked to explain the transport of excitation within individual antenna. However, it is unclear how these mechanisms influence transfer across assemblies of antenna and thus the photochemical yield at reaction centers in the functional thylakoid membrane. Here, we model light harvesting at the several-hundred-nanometer scale of the PSII membrane, while preserving the dominant quantum effects previously observed in individual complexes. We show that excitation moves diffusively through the antenna with a diffusion length of 50 nm until it reaches a reaction center, where charge separation serves as an energetic trap. The diffusion length is a single parameter that incorporates the enhancing effect of excited state delocalization on individual rates of energy transfer as well as the complex kinetics that arise due to energy transfer and loss by decay to the ground state. The diffusion length determines PSII's high quantum efficiency in ideal conditions, as well as how it is altered by the membrane morphology and the closure of reaction centers. We anticipate that the model will be useful in resolving the nonphotochemical quenching mechanisms that PSII employs in conditions of high light stress.

  11. Multiscale model of light harvesting by photosystem II in plants

    PubMed Central

    Amarnath, Kapil; Bennett, Doran I. G.; Schneider, Anna R.; Fleming, Graham R.

    2016-01-01

    The first step of photosynthesis in plants is the absorption of sunlight by pigments in the antenna complexes of photosystem II (PSII), followed by transfer of the nascent excitation energy to the reaction centers, where long-term storage as chemical energy is initiated. Quantum mechanical mechanisms must be invoked to explain the transport of excitation within individual antenna. However, it is unclear how these mechanisms influence transfer across assemblies of antenna and thus the photochemical yield at reaction centers in the functional thylakoid membrane. Here, we model light harvesting at the several-hundred-nanometer scale of the PSII membrane, while preserving the dominant quantum effects previously observed in individual complexes. We show that excitation moves diffusively through the antenna with a diffusion length of 50 nm until it reaches a reaction center, where charge separation serves as an energetic trap. The diffusion length is a single parameter that incorporates the enhancing effect of excited state delocalization on individual rates of energy transfer as well as the complex kinetics that arise due to energy transfer and loss by decay to the ground state. The diffusion length determines PSII’s high quantum efficiency in ideal conditions, as well as how it is altered by the membrane morphology and the closure of reaction centers. We anticipate that the model will be useful in resolving the nonphotochemical quenching mechanisms that PSII employs in conditions of high light stress. PMID:26787911

  12. Tracking Membrane Protein Association in Model Membranes

    PubMed Central

    Reffay, Myriam; Gambin, Yann; Benabdelhak, Houssain; Phan, Gilles; Taulier, Nicolas; Ducruix, Arnaud; Hodges, Robert S.; Urbach, Wladimir

    2009-01-01

    Membrane proteins are essential in the exchange processes of cells. In spite of great breakthrough in soluble proteins studies, membrane proteins structures, functions and interactions are still a challenge because of the difficulties related to their hydrophobic properties. Most of the experiments are performed with detergent-solubilized membrane proteins. However widely used micellar systems are far from the biological two-dimensions membrane. The development of new biomimetic membrane systems is fundamental to tackle this issue. We present an original approach that combines the Fluorescence Recovery After fringe Pattern Photobleaching technique and the use of a versatile sponge phase that makes it possible to extract crucial informations about interactions between membrane proteins embedded in the bilayers of a sponge phase. The clear advantage lies in the ability to adjust at will the spacing between two adjacent bilayers. When the membranes are far apart, the only possible interactions occur laterally between proteins embedded within the same bilayer, whereas when membranes get closer to each other, interactions between proteins embedded in facing membranes may occur as well. After validating our approach on the streptavidin-biotinylated peptide complex, we study the interactions between two membrane proteins, MexA and OprM, from a Pseudomonas aeruginosa efflux pump. The mode of interaction, the size of the protein complex and its potential stoichiometry are determined. In particular, we demonstrate that: MexA is effectively embedded in the bilayer; MexA and OprM do not interact laterally but can form a complex if they are embedded in opposite bilayers; the population of bound proteins is at its maximum for bilayers separated by a distance of about 200 Å, which is the periplasmic thickness of Pseudomonas aeruginosa. We also show that the MexA-OprM association is enhanced when the position and orientation of the protein is restricted by the bilayers. We

  13. Proteins causing membrane fouling in membrane bioreactors.

    PubMed

    Miyoshi, Taro; Nagai, Yuhei; Aizawa, Tomoyasu; Kimura, Katsuki; Watanabe, Yoshimasa

    2015-01-01

    In this study, the details of proteins causing membrane fouling in membrane bioreactors (MBRs) treating real municipal wastewater were investigated. Two separate pilot-scale MBRs were continuously operated under significantly different operating conditions; one MBR was a submerged type whereas the other was a side-stream type. The submerged and side-stream MBRs were operated for 20 and 10 days, respectively. At the end of continuous operation, the foulants were extracted from the fouled membranes. The proteins contained in the extracted foulants were enriched by using the combination of crude concentration with an ultrafiltration membrane and trichloroacetic acid precipitation, and then separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). The N-terminal amino acid sequencing analysis of the proteins which formed intensive spots on the 2D-PAGE gels allowed us to partially identify one protein (OmpA family protein originated from genus Brevundimonas or Riemerella anatipestifer) from the foulant obtained from the submerged MBR, and two proteins (OprD and OprF originated from genus Pseudomonas) from that obtained from the side-stream MBR. Despite the significant difference in operating conditions of the two MBRs, all proteins identified in this study belong to β-barrel protein. These findings strongly suggest the importance of β-barrel proteins in developing membrane fouling in MBRs.

  14. Proteins causing membrane fouling in membrane bioreactors.

    PubMed

    Miyoshi, Taro; Nagai, Yuhei; Aizawa, Tomoyasu; Kimura, Katsuki; Watanabe, Yoshimasa

    2015-01-01

    In this study, the details of proteins causing membrane fouling in membrane bioreactors (MBRs) treating real municipal wastewater were investigated. Two separate pilot-scale MBRs were continuously operated under significantly different operating conditions; one MBR was a submerged type whereas the other was a side-stream type. The submerged and side-stream MBRs were operated for 20 and 10 days, respectively. At the end of continuous operation, the foulants were extracted from the fouled membranes. The proteins contained in the extracted foulants were enriched by using the combination of crude concentration with an ultrafiltration membrane and trichloroacetic acid precipitation, and then separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). The N-terminal amino acid sequencing analysis of the proteins which formed intensive spots on the 2D-PAGE gels allowed us to partially identify one protein (OmpA family protein originated from genus Brevundimonas or Riemerella anatipestifer) from the foulant obtained from the submerged MBR, and two proteins (OprD and OprF originated from genus Pseudomonas) from that obtained from the side-stream MBR. Despite the significant difference in operating conditions of the two MBRs, all proteins identified in this study belong to β-barrel protein. These findings strongly suggest the importance of β-barrel proteins in developing membrane fouling in MBRs. PMID:26360742

  15. High-resolution AFM topographs of Rubrivivax gelatinosus light-harvesting complex LH2

    PubMed Central

    Scheuring, Simon; Reiss-Husson, Francoise; Engel, Andreas; Rigaud, Jean-Louis; Ranck, Jean-Luc

    2001-01-01

    Light-harvesting complexes 2 (LH2) are the accessory antenna proteins in the bacterial photosynthetic apparatus and are built up of αβ-heterodimers containing three bacteriochlorophylls and one carotenoid each. We have used atomic force microscopy (AFM) to investigate reconstituted LH2 from Rubrivivax gelatinosus, which has a C-terminal hydrophobic extension of 21 amino acids on the α-subunit. High-resolution topographs revealed a nonameric organization of the regularly packed cylindrical complexes incorporated into the membrane in both orientations. Native LH2 showed one surface which protruded by ∼6 Å and one that protruded by ∼14 Å from the membrane. Topographs of samples reconstituted with thermolysin-digested LH2 revealed a height reduction of the strongly protruding surface to ∼9 Å, and a change of its surface appearance. These results suggested that the α-subunit of R.gelatinosus comprises a single transmembrane helix and an extrinsic C-terminus, and allowed the periplasmic surface to be assigned. Occasionally, large rings (∼120 Å diameter) surrounded by LH2 rings were observed. Their diameter and appearance suggest the large rings to be LH1 complexes. PMID:11406579

  16. Polymer light harvesting composites for optoelectronic applications

    NASA Astrophysics Data System (ADS)

    Sun, Sam-Shajing; Wang, Dan

    2015-09-01

    Polymer based optoelectronic composites and thin film devices exhibit great potential in space applications due to their lightweight, flexible shape, high photon absorption coefficients, and robust radiation tolerance in space environment. Polymer/dye composites appear promising for optoelectronics applications due to potential enhancements in both light harvesting and charge separation. In this study, the optoelectronic properties of a series of molecular dyes paired with a conjugated polymer Poly(3-hexylthiophene-2,5-diyl) (P3HT) were investigated. Specifically, the solution PL quenching coefficients (Ksv) of dye/polymer follows a descending order from dyes of Chloro(protoporphyrinato)iron(III) (Hemin), Protoporphyrin, to meso-Tetra(4-carboxyphenyl)porphine (TCPP). In optoelectronic devices made of the P3HT/dye/PCBM composites, the short circuit current densities Jsc as well as the overall power conversion efficiencies (PCE) also follow a descending order from Hemin, Protoporphyrin, to TCPP, despite Hemin exhibits the intermediate polymer/dye LUMO (lowest unoccupied molecular orbital) offset and lowest absorption coefficient as compared to the other two dyes, i.e., the cell optoelectronic efficiency did not follow the LUMO offsets which are the key driving forces for the photo induced charge separations. This study reveals that too large LUMO offset or electron transfer driving force may result in smaller PL quenching and optoelectronic conversion efficiency, this could be another experimental evidence for the Marcus electron transfer model, particularly for the Marcus `inverted region'. It appears an optimum electron transfer driving force or strong PL quenching appears more critical than absorption coefficient for optoelectronic conversion devices.

  17. Modulation of the light-harvesting chlorophyll antenna size in Chlamydomonas reinhardtii by TLA1 gene over-expression and RNA interference

    PubMed Central

    Mitra, Mautusi; Kirst, Henning; Dewez, David; Melis, Anastasios

    2012-01-01

    Truncated light-harvesting antenna 1 (TLA1) is a nuclear gene proposed to regulate the chlorophyll (Chl) antenna size in Chlamydomonas reinhardtii. The Chl antenna size of the photosystems and the chloroplast ultrastructure were manipulated upon TLA1 gene over-expression and RNAi downregulation. The TLA1 over-expressing lines possessed a larger chlorophyll antenna size for both photosystems and contained greater levels of Chl b per cell relative to the wild type. Conversely, TLA1 RNAi transformants had a smaller Chl antenna size for both photosystems and lower levels of Chl b per cell. Western blot analyses of the TLA1 over-expressing and RNAi transformants showed that modulation of TLA1 gene expression was paralleled by modulation in the expression of light-harvesting protein, reaction centre D1 and D2, and VIPP1 genes. Transmission electron microscopy showed that modulation of TLA1 gene expression impacts the organization of thylakoid membranes in the chloroplast. Over-expressing lines showed well-defined grana, whereas RNAi transformants possessed loosely held together and more stroma-exposed thylakoids. Cell fractionation suggested localization of the TLA1 protein in the inner chloroplast envelope and potentially in association with nascent thylakoid membranes, indicating a role in Chl antenna assembly and thylakoid membrane biogenesis. The results provide a mechanistic understanding of the Chl antenna size regulation by the TLA1 gene. PMID:23148270

  18. Ab inito study on triplet excitation energy transfer in photosynthetic light-harvesting complexes.

    PubMed

    You, Zhi-Qiang; Hsu, Chao-Ping

    2011-04-28

    We have studied the triplet energy transfer (TET) for photosynthetic light-harvesting complexes, the bacterial light-harvesting complex II (LH2) of Rhodospirillum molischianum and Rhodopseudomonas acidophila, and the peridinin-chlorophyll a protein (PCP) from Amphidinium carterae. The electronic coupling factor was calculated with the recently developed fragment spin difference scheme (You and Hsu, J. Chem. Phys. 2010, 133, 074105), which is a general computational scheme that yields the overall coupling under the Hamiltonian employed. The TET rates were estimated based on the couplings obtained. For all light-harvesting complexes studied, there exist nanosecond triplet energy transfer from the chlorophylls to the carotenoids. This result supports a direct triplet quenching mechanism for the photoprotection function of carotenoids. The TET rates are similar for a broad range of carotenoid triplet state energy, which implies a general and robust TET quenching role for carotenoids in photosynthesis. This result is also consistent with the weak dependence of TET kinetics on the type or the number of π conjugation lengths in the carotenoids and their analogues reported in the literature. We have also explored the possibility of forming triplet excitons in these complexes. In B850 of LH2 or the peridinin cluster in PCP, it is unlikely to have triplet exciton since the energy differences of any two neighboring molecules are likely to be much larger than their TET couplings. Our results provide theoretical limits to the possible photophysics in the light-harvesting complexes.

  19. Microtechnologies for membrane protein studies

    PubMed Central

    Suzuki, Hiroaki

    2008-01-01

    Despite the rapid and enormous progress in biotechnologies, the biochemical analysis of membrane proteins is still a difficult task. The presence of the large hydrophobic region buried in the lipid bilayer membrane (transmembrane domain) makes it difficult to analyze membrane proteins in standard assays developed for water-soluble proteins. To handle membrane proteins, the lipid bilayer membrane may be used as a platform to sustain their functionalities. Relatively slow progress in developing micro total analysis systems (μTAS) for membrane protein analysis directly reflects the difficulty of handling lipid membranes, which is a common problem in bulk measurement technologies. Nonetheless, researchers are continuing to develop efficient and sensitive analytical microsystems for the study of membrane proteins. Here, we review the latest developments, which enable detection of events caused by membrane proteins, such as ion channel current, membrane transport, and receptor/ligand interaction, by utilizing microfabricated structures. High-throughput and highly sensitive detection systems for membrane proteins are now becoming a realistic goal. Although most of these systems are still in the early stages of development, we believe this field will become one of the most important applications of μTAS for pharmaceutical and clinical screenings as well as for basic biochemical research. PMID:18335213

  20. A femtosecond visible/visible and visible/mid-infrared transient absorption study of the light harvesting complex II.

    PubMed

    Stahl, Andreas D; Di Donato, Mariangela; van Stokkum, Ivo; van Grondelle, Rienk; Groot, Marie Louise

    2009-12-16

    Light harvesting complex II (LHCII) is the most abundant protein in the thylakoid membrane of higher plants and green algae. LHCII acts to collect solar radiation, transferring this energy mainly toward photosystem II, with a smaller amount going to photosystem I; it is then converted into a chemical, storable form. We performed time-resolved femtosecond visible pump/mid-infrared probe and visible pump/visible probe absorption difference spectroscopy on purified LHCII to gain insight into the energy transfer in this complex occurring in the femto-picosecond time regime. We find that information derived from mid-infrared spectra, together with structural and modeling information, provides a unique visualization of the flow of energy via the bottleneck pigment chlorophyll a604.

  1. Understanding photosynthetic light-harvesting: a bottom up theoretical approach.

    PubMed

    Renger, Thomas; Müh, Frank

    2013-03-14

    We discuss a bottom up approach for modeling photosynthetic light-harvesting. Methods are reviewed for a full structure-based parameterization of the Hamiltonian of pigment-protein complexes (PPCs). These parameters comprise (i) the local transition energies of the pigments in their binding sites in the protein, the site energies; (ii) the couplings between optical transitions of the pigments, the excitonic couplings; and (iii) the spectral density characterizing the dynamic modulation of pigment transition energies and excitonic couplings by protein vibrations. Starting with quantum mechanics perturbation theory, we provide a microscopic foundation for the standard PPC Hamiltonian and relate the expressions obtained for its matrix elements to quantities that can be calculated with classical molecular mechanics/electrostatics approaches including the whole PPC in atomic detail and using charge and transition densities obtained with quantum chemical calculations on the isolated building blocks of the PPC. In the second part of this perspective, the Hamiltonian is utilized to describe the quantum dynamics of excitons. Situations are discussed that differ in the relative strength of excitonic and exciton-vibrational coupling. The predictive power of the approaches is demonstrated in application to different PPCs, and challenges for future work are outlined.

  2. Proteins interacting with Membranes: Protein Sorting and Membrane Shaping

    NASA Astrophysics Data System (ADS)

    Callan-Jones, Andrew

    2015-03-01

    Membrane-bound transport in cells requires generating membrane curvature. In addition, transport is selective, in order to establish spatial gradients of membrane components in the cell. The mechanisms underlying cell membrane shaping by proteins and the influence of curvature on membrane composition are active areas of study in cell biophysics. In vitro approaches using Giant Unilamellar Vesicles (GUVs) are a useful tool to identify the physical mechanisms that drive sorting of membrane components and membrane shape change by proteins. I will present recent work on the curvature sensing and generation of IRSp53, a protein belonging to the BAR family, whose members, sharing a banana-shaped backbone, are involved in endocytosis. Pulling membrane tubes with 10-100 nm radii from GUVs containing encapsulated IRSp53 have, unexpectedly, revealed a non-monotonic dependence of the protein concentration on the tube as a function of curvature. Experiments also show that bound proteins alter the tube mechanics and that protein phase separation along the tube occurs at low tensions. I will present accompanying theoretical work that can explain these findings based on the competition between the protein's intrinsic curvature and the effective rigidity of a membrane-protein patch.

  3. Functionalized dye encapsulated polymer nanoparticles attached with a BSA scaffold as efficient antenna materials for artificial light harvesting.

    PubMed

    Jana, Bikash; Bhattacharyya, Santanu; Patra, Amitava

    2016-09-21

    A potential strategy for a new generation light harvesting system is multi-chromophoric donor-acceptor pairs where light energy is absorbed by an antenna complex and subsequently transfers its energy to the acceptor via energy transfer. Here, we design a system of a functionalized polymer nanoparticle-protein scaffold for efficient light harvesting and white light generation where a dye doped polymer nanoparticle acts as a donor and a dye encapsulated BSA protein acts as an acceptor. Analysis reveals that 91.3% energy transfer occurs from the dye doped polymer nanoparticle to the dye encapsulated BSA protein. The antenna effect of this light harvesting system is found to be 31 at a donor to acceptor ratio of 0.82 : 1 which is unprecedented. The enhanced effective molar extinction coefficient of the acceptor dye is potential for the light harvesting system. Bright white light emission with a quantum yield of 14% under single wavelength excitation is obtained by changing the ratio of donor to acceptor. Analysis reveals that the efficient energy transfer in this polymer-protein assembly may open up new possibilities in designing artificial light harvesting systems for future applications.

  4. Functionalized dye encapsulated polymer nanoparticles attached with a BSA scaffold as efficient antenna materials for artificial light harvesting.

    PubMed

    Jana, Bikash; Bhattacharyya, Santanu; Patra, Amitava

    2016-09-21

    A potential strategy for a new generation light harvesting system is multi-chromophoric donor-acceptor pairs where light energy is absorbed by an antenna complex and subsequently transfers its energy to the acceptor via energy transfer. Here, we design a system of a functionalized polymer nanoparticle-protein scaffold for efficient light harvesting and white light generation where a dye doped polymer nanoparticle acts as a donor and a dye encapsulated BSA protein acts as an acceptor. Analysis reveals that 91.3% energy transfer occurs from the dye doped polymer nanoparticle to the dye encapsulated BSA protein. The antenna effect of this light harvesting system is found to be 31 at a donor to acceptor ratio of 0.82 : 1 which is unprecedented. The enhanced effective molar extinction coefficient of the acceptor dye is potential for the light harvesting system. Bright white light emission with a quantum yield of 14% under single wavelength excitation is obtained by changing the ratio of donor to acceptor. Analysis reveals that the efficient energy transfer in this polymer-protein assembly may open up new possibilities in designing artificial light harvesting systems for future applications. PMID:27546792

  5. LINEAR AND CIRCULAR DICHROISM OF MEMBRANES FROM RHODOPSEUDOMONAS CAPSULATA

    SciTech Connect

    Bolt, John D.; Sauer, Kenneth; Shiozawa, Judith A.; Drews, Gerhart

    1980-10-01

    Absorption, linear dichroism and circular dichroism spectra of Rhodopseudomonas capsulate (wild-type - St. Louis strain, mutant Y5 and mutant A1a{sup +}) are particularly sensitive to the nature of the light-harvesting bacteriochlorophyll-carotenoid-protein complexes. Evidence for exciton-type interactions is seen near 855 nm in the membranes from the wild-type and from mutant Y5, as well as in an isolated B800+850 light-harvesting complex from mutant Y5. The strong circular dichroism that reflects these interactions is attenuated more than 10-fold in membranes from the A1a+ mutant, which lacks both B800+850 and colored carotenoids and contains only the B875 light-harvesting complex. These results lead to the conclusion that these two light-harvesting complexes have significantly different chromophore arrangements or local environments.

  6. How some proteins tubulate membranes

    NASA Astrophysics Data System (ADS)

    Bassereau, Patricia

    2009-03-01

    Endocytosis, exocytosis, membrane transport between intracellular compartments, virus or toxin entry or exit out of the cell, all imply to deform membrane. Membrane deformation mechanisms of cell membranes by proteins are currently actively studied. Giant vesicles (GUV) are interesting model membrane systems because they are composed of a very limited number of components compared to cellular membranes. The deformations induced by the interaction with a specific protein or any other additional components to the system, can then be directly monitored and the deformation mechanism eventually understood. In this talk, we will focus on different tubular structures induced by proteins. We will show that the B-subunits of Shiga toxin or Cholera Toxin, binding to their lipid receptors, Gb3 or GM1 respectively, incorporated in GUV membrane, induce negative membrane curvature and form tubular invaginations, in absence of any other cellular machinery. Tubular structures can also be obtained when molecular motors walking along microtubules exert a pulling force on the membrane of GUV. The helicoidal assembly of dynamin, a protein involved in vivo in membrane fission can also produce tubular structures. This assembly has been reconstituted around membrane nanotubes of controlled diameter; we will show that the initial tube diameter strongly influences dynamin polymerisation. In each case, a physical framework for understanding deformation mechanism will be presented

  7. Anti-Correlated Pigment Fluctuations of Allophycocyanin for Highly Efficient Photosynthetic Light Harvesting in Cyanobacteria

    NASA Astrophysics Data System (ADS)

    Moran, Andrew; Nome, Rene; Scherer, Norbert

    2008-03-01

    The phycobiliprotein, allophycocyanin (APC), is an excellent model system for the study of light harvesting pigment interactions with a protein bath. This work investigates the relaxation of electronic excitations in APC with electric field-resolved transient grating and photon echo spectroscopies. Transient grating experiments observe a 35 fs internal conversion process between single exciton levels. Most importantly, our analysis shows that anti-correlated phycocyanobilin pigment energy level fluctuations cause the anti-diagonal orientation of the node in the measured dispersive photon echo spectrum. We believe this novel observation to reflect concerted protein bath fluctuations over the 2 nm length scale that separates the pigments. Consideration of the Forster energy transfer rate theory suggests that APC has evolved with this property to enhance its photosynthetic light harvesting efficiency.

  8. Plasmon-controlled light-harvesting: design rules for biohybrid devices via multiscale modeling.

    PubMed

    Andreussi, Oliviero; Biancardi, Alessandro; Corni, Stefano; Mennucci, Benedetta

    2013-09-11

    Photosynthesis is triggered by the absorption of light by light-harvesting (LH) pigment-protein complexes followed by excitation energy transfer to the reaction center(s). A promising strategy to achieve control on and to improve light harvesting is to complement the LH complexes with plasmonic particles. Here a recently developed QM/MM/continuum approach is used to investigate the LH process of the peridinin-chlorophyll-protein (PCP) complex on a silver island film. The simulations not only reproduce and interpret the experiments but they also suggest general rules to design novel biohybrid devices; hot-spot configurations in which the LH complex is sandwiched between couples of metal aggregates are found to produce the largest amplifications. Indications about the best distances and orientations are also reported together with illumination and emission geometries of the PCP-NP system necessary to achieve the maximum enhancement.

  9. Solid-state NMR applied to photosynthetic light-harvesting complexes.

    PubMed

    Pandit, Anjali; de Groot, Huub J M

    2012-03-01

    This short review describes how solid-state NMR has provided a mechanistic and electronic picture of pigment-protein and pigment-pigment interactions in photosynthetic antenna complexes. NMR results on purple bacterial antenna complexes show how the packing of the protein and the pigments inside the light-harvesting oligomers induces mutual conformational stress. The protein scaffold produces deformation and electrostatic polarization of the BChl macrocycles and leads to a partial electronic charge transfer between the BChls and their coordinating histidines, which can tune the light-harvesting function. In chlorosome antennae assemblies, the NMR template structure reveals how the chromophores can direct their self-assembly into higher macrostructures which, in turn, tune the light-harvesting properties of the individual molecules by controlling their disorder, structural deformation, and electronic polarization without the need for a protein scaffold. These results pave the way for addressing the next challenge, which is to resolve the functional conformational dynamics of the lhc antennae of oxygenic species that allows them to switch between light-emitting and light-energy dissipating states.

  10. Light-harvesting regulation from leaf to molecule with the emphasis on rapid changes in antenna size.

    PubMed

    Xu, Da-Quan; Chen, Yue; Chen, Gen-Yun

    2015-05-01

    In the sunlight-fluctuating environment, plants often encounter both light-deficiency and light-excess cases. Therefore, regulation of light harvesting is absolutely essential for photosynthesis in order to maximize light utilization at low light and avoid photodamage of the photosynthetic apparatus at high light. Plants have developed a series of strategies of light-harvesting regulation during evolution. These strategies include rapid responses such as leaf movement and chloroplast movement, state transitions, and reversible dissociation of some light-harvesting complex of the photosystem II (LHCIIs) from PSII core complexes, and slow acclimation strategies such as changes in the protein abundance of light-harvesting antenna and modifications of leaf morphology, structure, and compositions. This review discusses successively these strategies and focuses on the rapid change in antenna size, namely reversible dissociation of some peripheral light-harvesting antennas (LHCIIs) from PSII core complex. It is involved in protective role and species dependence of the dissociation, differences between the dissociation and state transitions, relationship between the dissociation and thylakoid protein phosphorylation, and possible mechanism for thermal dissipation by the dissociated LHCIIs. PMID:25773873

  11. Single-molecule exploration of photoprotective mechanisms in light-harvesting complexes

    NASA Astrophysics Data System (ADS)

    Yang, Hsiang-Yu; Schlau-Cohen, Gabriela S.; Gwizdala, Michal; Krüger, Tjaart; Xu, Pengqi; Croce, Roberta; van Grondelle, Rienk; Moerner, W. E.

    2015-03-01

    Plants harvest sunlight by converting light energy to electron flow through the primary events in photosynthesis. One important question is how the light harvesting machinery adapts to fluctuating sunlight intensity. As a result of various regulatory processes, efficient light harvesting and photoprotection are balanced. Some of the biological steps in the photoprotective processes have been extensively studied and physiological regulatory factors have been identified. For example, the effect of lumen pH in changing carotenoid composition has been explored. However, the importance of photophysical dynamics in the initial light-harvesting steps and its relation to photoprotection remain poorly understood. Conformational and excited-state dynamics of multi-chromophore pigment-protein complexes are often difficult to study and limited information can be extracted from ensemble-averaged measurements. To address the problem, we use the Anti-Brownian ELectrokinetic (ABEL) trap to investigate the fluorescence from individual copies of light-harvesting complex II (LHCII), the primary antenna protein in higher plants, in a solution-phase environment. Perturbative surface immobilization or encapsulation schemes are avoided, and therefore the intrinsic dynamics and heterogeneity in the fluorescence of individual proteins are revealed. We perform simultaneous measurements of fluorescence intensity (brightness), excited-state lifetime, and emission spectrum of single trapped proteins. By analyzing the correlated changes between these observables, we identify forms of LHCII with different fluorescence intensities and excited-state lifetimes. The distinct forms may be associated with different energy dissipation mechanisms in the energy transfer chain. Changes of relative populations in response to pH and carotenoid composition are observed, which may extend our understanding of the molecular mechanisms of photoprotection.

  12. Predicting the structure of the light-harvesting complex II of Rhodospirillum molischianum.

    PubMed Central

    Hu, X.; Xu, D.; Hamer, K.; Schulten, K.; Koepke, J.; Michel, H.

    1995-01-01

    We attempted to predict through computer modeling the structure of the light-harvesting complex II (LH-II) of Rhodospirillum molischianum, before the impending publication of the structure of a homologous protein solved by means of X-ray diffraction. The protein studied is an integral membrane protein of 16 independent polypeptides, 8 alpha-apoproteins and 8 beta-apoproteins, which aggregate and bind to 24 bacteriochlorophyll-a's and 12 lycopenes. Available diffraction data of a crystal of the protein, which could not be phased due to a lack of heavy metal derivatives, served to test the predicted structure, guiding the search. In order to determine the secondary structure, hydropathy analysis was performed to identify the putative transmembrane segments and multiple sequence alignment propensity analyses were used to pinpoint the exact sites of the 20-residue-long transmembrane segment and the 4-residue-long terminal sequence at both ends, which were independently verified and improved by homology modeling. A consensus assignment for the secondary structure was derived from a combination of all the prediction methods used. Three-dimensional structures for the alpha- and the beta-apoprotein were built by comparative modeling. The resulting tertiary structures are combined, using X-PLOR, into an alpha beta dimer pair with bacteriochlorophyll-a's attached under constraints provided by site-directed mutagenesis and spectral data. The alpha beta dimer pairs were then aggregated into a quaternary structure through further molecular dynamics simulations and energy minimization. The structure of LH-II so determined is an octamer of alpha beta heterodimers forming a ring with a diameter of 70 A. PMID:8528066

  13. Early folding events during light harvesting complex II assembly in vitro monitored by pulsed electron paramagnetic resonance.

    PubMed

    Fehr, Niklas; García-Rubio, Inés; Jeschke, Gunnar; Paulsen, Harald

    2016-06-01

    Efficient energy transfer in the major light harvesting complex II (LHCII) of green plants is facilitated by the precise alignment of pigments due to the protein matrix they are bound to. Much is known about the import of the LHCII apoprotein into the chloroplast via the TOC/TIC system and its targeting to the thylakoid membrane but information is sparse about when and where the pigments are bound and how this is coordinated with protein folding. In vitro, the LHCII apoprotein spontaneously folds and binds its pigments if the detergent-solubilized protein is combined with a mixture of chlorophylls a and b and carotenoids. In the present work, we employed this approach to study apoprotein folding and pigment binding in a time-resolved manner by using pulsed electron paramagnetic resonance (EPR). Intra-molecular distances were measured before folding, after 255 ms and 40 s folding time in the absence of cryoprotectant, and in the fully folded and assembled LHCII. In accordance with earlier results, the most of the folding of the three membrane-spanning alpha helices precedes their apposition into the final tertiary structure. However, their formation follows different kinetics, partially extending into the final phase of LHCII formation during which much of the condensation of the pigment-protein structure occurs, presumably governed by the binding of chlorophyll b. A rough timetable is proposed to sort partial events into the LHCII formation process. PMID:27063475

  14. Design principles of natural light-harvesting as revealed by single molecule spectroscopy

    NASA Astrophysics Data System (ADS)

    Krüger, T. P. J.; van Grondelle, R.

    2016-01-01

    Biology offers a boundless source of adaptation, innovation, and inspiration. A wide range of photosynthetic organisms exist that are capable of harvesting solar light in an exceptionally efficient way, using abundant and low-cost materials. These natural light-harvesting complexes consist of proteins that strongly bind a high density of chromophores to capture solar photons and rapidly transfer the excitation energy to the photochemical reaction centre. The amount of harvested light is also delicately tuned to the level of solar radiation to maintain a constant energy throughput at the reaction centre and avoid the accumulation of the products of charge separation. In this Review, recent developments in the understanding of light-harvesting by plants will be discussed, based on results obtained from single molecule spectroscopy studies. Three design principles of the main light-harvesting antenna of plants will be highlighted: (a) fine, photoactive control over the intrinsic protein disorder to efficiently use intrinsically available thermal energy dissipation mechanisms; (b) the design of the protein microenvironment of a low-energy chromophore dimer to control the amount of shade absorption; (c) the design of the exciton manifold to ensure efficient funneling of the harvested light to the terminal emitter cluster.

  15. Protein kinase and phosphatase activities of thylakoid membranes

    SciTech Connect

    Michel, H.; Shaw, E.K.; Bennett, J.

    1987-01-01

    Dephosphorylation of the 25 and 27 kDa light-harvesting Chl a/b proteins (LHCII) of the thylakoid membranes is catalyzed by a phosphatase which differs from previously reported thylakoid-bound phosphatases in having an alkaline pH optimum (9.0) and a requirement for Mg/sup 2 +/ ions. Dephosphorylation of the 8.3 kDa psb H gene product requires a Mg/sup 2 +/ ion concentration more than 200 fold higher than that for dephosphorylation of LHC II. The 8.3 kDa and 27 kDa proteins appear to be phosphorylated by two distinct kinases, which differ in substrate specificity and sensitivity to inhibitors. The plastoquinone antagonist 2,5-dibromo-3-methyl-6-isopropyl-benzoquinone (DBMIB) inhibits phosphorylation of the 27 kDa LHC II much more readily than phosphorylation of the 8.3 kDa protein. A similar pattern of inhibition is seen for two synthetic oligopeptides (MRKSATTKKAVC and ATQTLESSSRC) which are analogs of the phosphorylation sites of the two proteins. Possible modes of action of DBMIB are discussed. 45 refs., 7 figs., 3 tabs.

  16. Convergent synthesis of multiporphyrin light-harvesting rods

    DOEpatents

    Lindsey, Jonathan S.; Loewe, Robert S.

    2003-08-05

    The present invention provides a convergent method for the synthesis of light harvesting rods. The rods are oligomers of the formula A.sup.1 (A.sup.b+1).sub.b, wherein b is at least 1, A.sup.1 through A.sup.b+1 are covalently coupled rod segments, and each rod segment A.sup.1 through A.sup.1+b comprises a compound of the formula X.sup.1 (X.sup.m+1).sub.m wherein m is at least 1 and X.sup.1 through X.sup.m+1 are covalently coupled porphyrinic macrocycles. Light harvesting arrays and solar cells containing such light harvesting rods are also described, along with intermediates useful in such methods and rods produced by such methods.

  17. Quantum coherent energy transfer over varying pathways in single light-harvesting complexes.

    PubMed

    Hildner, Richard; Brinks, Daan; Nieder, Jana B; Cogdell, Richard J; van Hulst, Niek F

    2013-06-21

    The initial steps of photosynthesis comprise the absorption of sunlight by pigment-protein antenna complexes followed by rapid and highly efficient funneling of excitation energy to a reaction center. In these transport processes, signatures of unexpectedly long-lived coherences have emerged in two-dimensional ensemble spectra of various light-harvesting complexes. Here, we demonstrate ultrafast quantum coherent energy transfer within individual antenna complexes of a purple bacterium under physiological conditions. We find that quantum coherences between electronically coupled energy eigenstates persist at least 400 femtoseconds and that distinct energy-transfer pathways that change with time can be identified in each complex. Our data suggest that long-lived quantum coherence renders energy transfer in photosynthetic systems robust in the presence of disorder, which is a prerequisite for efficient light harvesting.

  18. Metal-Enhanced Fluorescence of Chlorophylls in Light-Harvesting Complexes Coupled to Silver Nanowires

    PubMed Central

    Kowalska, Dorota; Krajnik, Bartosz; Olejnik, Maria; Czechowski, Nikodem; Mackowski, Sebastian

    2013-01-01

    We investigate metal-enhanced fluorescence of peridinin-chlorophyll protein coupled to silver nanowires using optical microscopy combined with spectrally and time-resolved fluorescence techniques. In particular we study two different sample geometries: first, in which the light-harvesting complexes are deposited onto silver nanowires, and second, where solution of both nanostructures are mixed prior deposition on a substrate. The results indicate that for the peridinin-chlorophyll complexes placed in the vicinity of the silver nanowires we observe higher intensities of fluorescence emission as compared to the reference sample, where no nanowires are present. Enhancement factors estimated for the sample where the light-harvesting complexes are mixed together with the silver nanowires prior deposition on a substrate are generally larger in comparison to the other geometry of a hybrid nanostructure. While fluorescence spectra are identical both in terms of overall shape and maximum wavelength for peridinin-chlorophyll-protein complexes both isolated and coupled to metallic nanostructures, we conclude that interaction with plasmon excitations in the latter remains neutral to the functionality of the biological system. Fluorescence transients measured for the PCP complexes coupled to the silver nanowires indicate shortening of the fluorescence lifetime pointing towards modifications of radiative rate due to plasmonic interactions. Our results can be applied for developing ways to plasmonically control the light-harvesting capability of photosynthetic complexes. PMID:23533354

  19. Metal-enhanced fluorescence of chlorophylls in light-harvesting complexes coupled to silver nanowires.

    PubMed

    Kowalska, Dorota; Krajnik, Bartosz; Olejnik, Maria; Twardowska, Magdalena; Czechowski, Nikodem; Hofmann, Eckhard; Mackowski, Sebastian

    2013-01-01

    We investigate metal-enhanced fluorescence of peridinin-chlorophyll protein coupled to silver nanowires using optical microscopy combined with spectrally and time-resolved fluorescence techniques. In particular we study two different sample geometries: first, in which the light-harvesting complexes are deposited onto silver nanowires, and second, where solution of both nanostructures are mixed prior deposition on a substrate. The results indicate that for the peridinin-chlorophyll complexes placed in the vicinity of the silver nanowires we observe higher intensities of fluorescence emission as compared to the reference sample, where no nanowires are present. Enhancement factors estimated for the sample where the light-harvesting complexes are mixed together with the silver nanowires prior deposition on a substrate are generally larger in comparison to the other geometry of a hybrid nanostructure. While fluorescence spectra are identical both in terms of overall shape and maximum wavelength for peridinin-chlorophyll-protein complexes both isolated and coupled to metallic nanostructures, we conclude that interaction with plasmon excitations in the latter remains neutral to the functionality of the biological system. Fluorescence transients measured for the PCP complexes coupled to the silver nanowires indicate shortening of the fluorescence lifetime pointing towards modifications of radiative rate due to plasmonic interactions. Our results can be applied for developing ways to plasmonically control the light-harvesting capability of photosynthetic complexes.

  20. Molecular dynamics of membrane proteins.

    SciTech Connect

    Woolf, Thomas B.; Crozier, Paul Stewart; Stevens, Mark Jackson

    2004-10-01

    Understanding the dynamics of the membrane protein rhodopsin will have broad implications for other membrane proteins and cellular signaling processes. Rhodopsin (Rho) is a light activated G-protein coupled receptor (GPCR). When activated by ligands, GPCRs bind and activate G-proteins residing within the cell and begin a signaling cascade that results in the cell's response to external stimuli. More than 50% of all current drugs are targeted toward G-proteins. Rho is the prototypical member of the class A GPCR superfamily. Understanding the activation of Rho and its interaction with its Gprotein can therefore lead to a wider understanding of the mechanisms of GPCR activation and G-protein activation. Understanding the dark to light transition of Rho is fully analogous to the general ligand binding and activation problem for GPCRs. This transition is dependent on the lipid environment. The effect of lipids on membrane protein activity in general has had little attention, but evidence is beginning to show a significant role for lipids in membrane protein activity. Using the LAMMPS program and simulation methods benchmarked under the IBIG program, we perform a variety of allatom molecular dynamics simulations of membrane proteins.

  1. Light-harvesting photocatalysis for water oxidation using mesoporous organosilica.

    PubMed

    Takeda, Hiroyuki; Ohashi, Masataka; Goto, Yasutomo; Ohsuna, Tetsu; Tani, Takao; Inagaki, Shinji

    2014-07-14

    An organic-based photocatalysis system for water oxidation, with visible-light harvesting antennae, was constructed using periodic mesoporous organosilica (PMO). PMO containing acridone groups in the framework (Acd-PMO), a visible-light harvesting antenna, was supported with [Ru(II)(bpy)3(2+)] complex (bpy = 2,2'-bipyridyl) coupled with iridium oxide (IrO(x)) particles in the mesochannels as photosensitizer and catalyst, respectively. Acd-PMO absorbed visible light and funneled the light energy into the Ru complex in the mesochannels through excitation energy transfer. The excited state of Ru complex is oxidatively quenched by a sacrificial oxidant (Na2S2O8) to form Ru(3+) species. The Ru(3+) species extracts an electron from IrO(x) to oxidize water for oxygen production. The reaction quantum yield was 0.34 %, which was improved to 0.68 or 1.2 % by the modifications of PMO. A unique sequence of reactions mimicking natural photosystem II, 1) light-harvesting, 2) charge separation, and 3) oxygen generation, were realized for the first time by using the light-harvesting PMO.

  2. Temperature and Ionic Strength Effects on the Chlorosome Light-Harvesting Antenna Complex

    SciTech Connect

    Tang, Kuo-Hsiang; Zhu, Liying; Urban, Volker S; Collins, Aaron M.; Biswas, Pratim; Blankenship, R. E.

    2011-03-15

    Chlorosomes, the peripheral light-harvesting antenna complex from green photosynthetic bacteria, are the largest and one of the most efficient light-harvesting antenna complexes found in nature. In contrast to other light-harvesting antennas, chlorosomes are constructed from more than 150,000 self-assembled bacteriochlorophylls (BChls) and contain relatively few proteins that play secondary roles. These unique properties have led to chlorosomes as an attractive candidate for developing biohybrid solar cell devices. In this article, we investigate the temperature and ionic strength effects on the viability of chlorosomes from the photosynthetic green bacterium Chloroflexus aurantiacus using small-angle neutron scattering and dynamic light scattering. Our studies indicate that chlorosomes remain intact up to 75 °C and that salt induces the formation of large aggregates of chlorosomes. No internal structural changes are observed for the aggregates. The salt-induced aggregation, which is a reversible process, is more efficient with divalent metal ions than with monovalent metal ions. Moreover, with treatment at 98 °C for 2 min, the bulk of the chlorosome pigments are undamaged, while the baseplate is destroyed. Chlorosomes without the baseplate remain rodlike in shape and are 30-40% smaller than with the baseplate attached. Further, chlorosomes are stable from pH 5.5 to 11.0. Together, this is the first time such a range of characterization tools have been used for chlorosomes, and this has enabled elucidation of properties that are not only important to understanding their functionality but also may be useful in biohybrid devices for effective light harvesting.

  3. Two Cyanobacterial Photoreceptors Regulate Photosynthetic Light Harvesting by Sensing Teal, Green, Yellow, and Red Light

    PubMed Central

    Wiltbank, Lisa B.

    2016-01-01

    ABSTRACT The genomes of many photosynthetic and nonphotosynthetic bacteria encode numerous phytochrome superfamily photoreceptors whose functions and interactions are largely unknown. Cyanobacterial genomes encode particularly large numbers of phytochrome superfamily members called cyanobacteriochromes. These have diverse light color-sensing abilities, and their functions and interactions are just beginning to be understood. One of the best characterized of these functions is the regulation of photosynthetic light-harvesting antenna composition in the cyanobacterium Fremyella diplosiphon by the cyanobacteriochrome RcaE in response to red and green light, a process known as chromatic acclimation. We have identified a new cyanobacteriochrome named DpxA that maximally senses teal (absorption maximum, 494 nm) and yellow (absorption maximum, 568 nm) light and represses the accumulation of a key light-harvesting protein called phycoerythrin, which is also regulated by RcaE during chromatic acclimation. Like RcaE, DpxA is a two-component system kinase, although these two photoreceptors can influence phycoerythrin expression through different signaling pathways. The peak responsiveness of DpxA to teal and yellow light provides highly refined color discrimination in the green spectral region, which provides important wavelengths for photosynthetic light harvesting in cyanobacteria. These results redefine chromatic acclimation in cyanobacteria and demonstrate that cyanobacteriochromes can coordinately impart sophisticated light color sensing across the visible spectrum to regulate important photosynthetic acclimation processes. PMID:26861023

  4. Cytochrome b 6 f function and localization, phosphorylation state of thylakoid membrane proteins and consequences on cyclic electron flow.

    PubMed

    Dumas, Louis; Chazaux, Marie; Peltier, Gilles; Johnson, Xenie; Alric, Jean

    2016-09-01

    Both the structure and the protein composition of thylakoid membranes have an impact on light harvesting and electron transfer in the photosynthetic chain. Thylakoid membranes form stacks and lamellae where photosystem II and photosystem I localize, respectively. Light-harvesting complexes II can be associated to either PSII or PSI depending on the redox state of the plastoquinone pool, and their distribution is governed by state transitions. Upon state transitions, the thylakoid ultrastructure and lateral distribution of proteins along the membrane are subject to significant rearrangements. In addition, quinone diffusion is limited to membrane microdomains and the cytochrome b 6 f complex localizes either to PSII-containing grana stacks or PSI-containing stroma lamellae. Here, we discuss possible similarities or differences between green algae and C3 plants on the functional consequences of such heterogeneities in the photosynthetic electron transport chain and propose a model in which quinones, accepting electrons either from PSII (linear flow) or NDH/PGR pathways (cyclic flow), represent a crucial control point. Our aim is to give an integrated description of these processes and discuss their potential roles in the balance between linear and cyclic electron flows. PMID:27534565

  5. Plant Plasma Membrane Proteins 1

    PubMed Central

    Grimes, Howard D.; Breidenbach, R. William

    1987-01-01

    A major 75 kD protein group from the tomato plasma membrane was semipurified on polyacrylamide gels and used to raise a rabbit antiserum. The resulting antiserum recognized a single 75 kilodalton band from phase partitioned tomato plasma membrane (from both suspension cells and mature, green fruit) after resolution on one-dimensional polyacrylamide gels. Two-dimensional polyacrylamide gel analysis of proteins from tomato plasma membrane showed that the 75 kilodalton antiserum recognized a group of proteins ranging from 63.1 to 88.2 kilodaltons (mean = 75.6 kilodaltons) and with isoelectric point values ranging from 5.7 to 6.3. No other spots were visible on the two-dimensional blots. This antiserum was shown to bind protoplast surface epitopes by indirect immunofluorescence. The presence of this protein group in both monocotyledonous and dicotyledonous plants was established by immunoblotting the tomato 75 kilodalton antiserum against proteins obtained from plasma membrane-enriched fractions from corn roots and soybean roots. The data suggest that this 75 kilodalton protein group is a major proteinaceous component of the plant plasma membrane. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:16665801

  6. The interactions of peripheral membrane proteins with biological membranes

    DOE PAGES

    Johs, Alexander; Whited, A. M.

    2015-01-01

    The interactions of peripheral proteins with membrane surfaces are critical to many biological processes, including signaling, recognition, membrane trafficking, cell division and cell structure. On a molecular level, peripheral membrane proteins can modulate lipid composition, membrane dynamics and protein-protein interactions. Biochemical and biophysical studies have shown that these interactions are in fact highly complex, dominated by several different types of interactions, and have an interdependent effect on both the protein and membrane. Here we examine three major mechanisms underlying the interactions between peripheral membrane proteins and membranes: electrostatic interactions, hydrophobic interactions, and fatty acid modification of proteins. While experimental approachesmore » continue to provide critical insights into specific interaction mechanisms, emerging bioinformatics resources and tools contribute to a systems-level picture of protein-lipid interactions. Through these recent advances, we begin to understand the pivotal role of protein-lipid interactions underlying complex biological functions at membrane interfaces.« less

  7. The interactions of peripheral membrane proteins with biological membranes.

    PubMed

    Whited, A M; Johs, A

    2015-11-01

    The interactions of peripheral proteins with membrane surfaces are critical to many biological processes, including signaling, recognition, membrane trafficking, cell division and cell structure. On a molecular level, peripheral membrane proteins can modulate lipid composition, membrane dynamics and protein-protein interactions. Biochemical and biophysical studies have shown that these interactions are in fact highly complex, dominated by several different types of interactions, and have an interdependent effect on both the protein and membrane. Here we examine three major mechanisms underlying the interactions between peripheral membrane proteins and membranes: electrostatic interactions, hydrophobic interactions, and fatty acid modification of proteins. While experimental approaches continue to provide critical insights into specific interaction mechanisms, emerging bioinformatics resources and tools contribute to a systems-level picture of protein-lipid interactions. Through these recent advances, we begin to understand the pivotal role of protein-lipid interactions underlying complex biological functions at membrane interfaces.

  8. The interactions of peripheral membrane proteins with biological membranes

    SciTech Connect

    Johs, Alexander; Whited, A. M.

    2015-01-01

    The interactions of peripheral proteins with membrane surfaces are critical to many biological processes, including signaling, recognition, membrane trafficking, cell division and cell structure. On a molecular level, peripheral membrane proteins can modulate lipid composition, membrane dynamics and protein-protein interactions. Biochemical and biophysical studies have shown that these interactions are in fact highly complex, dominated by several different types of interactions, and have an interdependent effect on both the protein and membrane. Here we examine three major mechanisms underlying the interactions between peripheral membrane proteins and membranes: electrostatic interactions, hydrophobic interactions, and fatty acid modification of proteins. While experimental approaches continue to provide critical insights into specific interaction mechanisms, emerging bioinformatics resources and tools contribute to a systems-level picture of protein-lipid interactions. Through these recent advances, we begin to understand the pivotal role of protein-lipid interactions underlying complex biological functions at membrane interfaces.

  9. Self-interaction chromatography as a tool for optimizing conditions for membrane protein crystallization.

    PubMed

    Gabrielsen, Mads; Nagy, Lisa A; DeLucas, Lawrence J; Cogdell, Richard J

    2010-01-01

    The second virial coefficient, or B value, is a measurement of how well a protein interacts with itself in solution. These interactions can lead to protein crystallization or precipitation, depending on their strength, with a narrow range of B values (the 'crystallization slot') being known to promote crystallization. A convenient method of determining the B value is by self-interaction chromatography. This paper describes how the light-harvesting complex 1-reaction centre core complex from Allochromatium vinosum yielded single straight-edged crystals after iterative cycles of self-interaction chromatography and crystallization. This process allowed the rapid screening of small molecules and detergents as crystallization additives. Here, a description is given of how self-interaction chromatography has been utilized to improve the crystallization conditions of a membrane protein.

  10. Conformational dynamics of a membrane protein chaperone enables spatially regulated substrate capture and release

    PubMed Central

    Liang, Fu-Cheng; Kroon, Gerard; McAvoy, Camille Z.; Chi, Chris; Wright, Peter E.; Shan, Shu-ou

    2016-01-01

    Membrane protein biogenesis poses enormous challenges to cellular protein homeostasis and requires effective molecular chaperones. Compared with chaperones that promote soluble protein folding, membrane protein chaperones require tight spatiotemporal coordination of their substrate binding and release cycles. Here we define the chaperone cycle for cpSRP43, which protects the largest family of membrane proteins, the light harvesting chlorophyll a/b-binding proteins (LHCPs), during their delivery. Biochemical and NMR analyses demonstrate that cpSRP43 samples three distinct conformations. The stromal factor cpSRP54 drives cpSRP43 to the active state, allowing it to tightly bind substrate in the aqueous compartment. Bidentate interactions with the Alb3 translocase drive cpSRP43 to a partially inactive state, triggering selective release of LHCP’s transmembrane domains in a productive unloading complex at the membrane. Our work demonstrates how the intrinsic conformational dynamics of a chaperone enables spatially coordinated substrate capture and release, which may be general to other ATP-independent chaperone systems. PMID:26951662

  11. Reconstitution of Gloeobacter violaceus Rhodopsin with a Light-Harvesting Carotenoid Antenna†

    PubMed Central

    Imasheva, Eleonora S.; Balashov, Sergei P.; Choi, Ah Reum; Jung, Kwang-Hwan; Lanyi, Janos K.

    2009-01-01

    We show that salinixanthin, the light-harvesting carotenoid antenna of xanthorhodopsin, can be reconstituted into the retinal protein from Gloeobacter violaceus expressed in E. coli. Reconstitution of gloeobacter rhodopsin with the carotenoid is accompanied by characteristic absorption changes and the appearance of CD bands similar to those observed for xanthorhodopsin that indicate immobilization and twist of the carotenoid in the binding site. As in xanthorhodopsin, the carotenoid functions as a light-harvesting antenna. The excitation spectrum for retinal fluorescence emission shows that ca. 36% of the energy absorbed by the carotenoid is transferred to the retinal. From excitation anisotropy, we calculate the angle between the two chromophores as ca. 50°, similar to that in xanthorhodopsin. The results indicate that gloeobacter rhodopsin binds salinixanthin in a similar way as xanthorhodopsin, and suggest that it might bind a carotenoid also in vivo. In the crystallographic structure of xanthorhodopsin, the conjugated chain of the carotenoid lies on the surface of helices E and F, and the 4-keto-ring is immersed in the protein at van der Waals distance from the ionone ring of the retinal. The 4-keto-ring is in the space occupied by a tryptophan in bacteriorhodopsin, which is replaced by the smaller glycine in xanthorhodopsin and gloeobacter rhodopsin. Specific binding of the carotenoid and its light-harvesting function are eliminated by a single mutation of the gloeobacter protein that replaces this glycine with a tryptophan. This indicates that the 4-keto-ring is critically involved in carotenoid binding, and suggests that a number of other recently identified retinal proteins, from a diverse group of organisms, could also contain carotenoid antenna since they carry the homologous glycine near the retinal. PMID:19842712

  12. Reconstitution of Gloeobacter violaceus rhodopsin with a light-harvesting carotenoid antenna.

    PubMed

    Imasheva, Eleonora S; Balashov, Sergei P; Choi, Ah Reum; Jung, Kwang-Hwan; Lanyi, Janos K

    2009-11-24

    We show that salinixanthin, the light-harvesting carotenoid antenna of xanthorhodopsin, can be reconstituted into the retinal protein from Gloeobacter violaceus expressed in Escherichia coli. Reconstitution of gloeobacter rhodopsin with the carotenoid is accompanied by characteristic absorption changes and the appearance of CD bands similar to those observed for xanthorhodopsin that indicate immobilization and twist of the carotenoid in the binding site. As in xanthorhodopsin, the carotenoid functions as a light-harvesting antenna. The excitation spectrum for retinal fluorescence emission shows that ca. 36% of the energy absorbed by the carotenoid is transferred to the retinal. From excitation anisotropy, we calculate the angle between the two chromophores as being ca. 50 degrees , similar to that in xanthorhodopsin. The results indicate that gloeobacter rhodopsin binds salinixanthin in a manner similar to that of xanthorhodopsin and suggest that it might bind a carotenoid also in vivo. In the crystallographic structure of xanthorhodopsin, the conjugated chain of the carotenoid lies on the surface of helices E and F, and the 4-keto ring is immersed in the protein at van der Waals distance from the ionone ring of the retinal. The 4-keto ring is in the space occupied by a tryptophan in bacteriorhodopsin, which is replaced by the smaller glycine in xanthorhodopsin and gloeobacter rhodopsin. Specific binding of the carotenoid and its light-harvesting function are eliminated by a single mutation of the gloeobacter protein that replaces this glycine with a tryptophan. This indicates that the 4-keto ring is critically involved in carotenoid binding and suggests that a number of other recently identified retinal proteins, from a diverse group of organisms, could also contain carotenoid antenna since they carry the homologous glycine near the retinal.

  13. Replication of Leaf Surface Structures for Light Harvesting

    PubMed Central

    Huang, Zhongjia; Yang, Sai; Zhang, Hui; Zhang, Meng; Cao, Wei

    2015-01-01

    As one of the most important hosts of natural light harvesting, foliage normally has complicated surface structures to capture solar radiances. Bio-mimicking leaf surface structures can provide novel designs of covers in photovoltaic systems. In this article, we reported on replicating leaf surface structures on poly-(methyl methacrylate) polymers to prompt harvesting efficiencies. Prepared via a double transfer process, the polymers were found to have high optical transparencies and transmission hazes, with both values exceeding 80% in some species. Benefiting from optical properties and wrinkled surfaces, the biomimetic polymers brought up to 17% gains to photovoltaic efficiencies. Through Monte-Carlo simulations of light transport, ultrahigh haze values and low reflections were attributed to lightwave guidance schemes lead by the nano- and micro-morphologies which are inherited from master leaves. Thus, leaf surface bio-mimicking can be considered as a strategic direction to design covers of light harvesting systems. PMID:26381702

  14. Ultraviolet-B photodestruction of a light-harvesting complex.

    PubMed Central

    Lao, K; Glazer, A N

    1996-01-01

    Cyanobacteria are important contributors to global photosynthesis in both marine and terrestrial environments. Quantitative data are presented on UV-B-induced damage to the major cyanobacterial photosynthetic light harvesting complex, the phycobilisome, and to each of its constituent phycobiliproteins. The photodestruction quantum yield, phi295 nm, for the phycobiliproteins is high (approximately 10(-3), as compared with approximately 10(-7) for visible light). Energy transfer on a picosecond time scale does not compete with photodestruction. Photodamage to phycobilisomes in vitro and in living cells is amplified by causing dissociation and loss of function of the complex. In photosynthetic organisms, UV-B damage to light-harvesting complexes may significantly exceed that to DNA. Images Fig. 1 PMID:8643563

  15. Light-harvesting materials: Soft support for energy conversion

    SciTech Connect

    Stolley, Ryan M.; Helm, Monte L.

    2014-11-10

    To convert solar energy into viable fuel sources, coupling light-harvesting materials to catalysts is a critical challenge. Now, coupling between an organic supramolecular hydrogel and a non precious metal catalyst has been demonstrated to be effective for photocatalytic H2 production. Ryan M. Stolley and Monte L. Helm are at Pacific Northwest National Laboratory (PNNL), Richland, WA, USA 99352. PNNL is operated by Battelle for the US Department of Energy. e-mail: Monte.Helm@pnnl.gov

  16. A light-harvesting array of synthetic porphyrins

    NASA Astrophysics Data System (ADS)

    Davila, Jorge; Harriman, Anthony; Milgrom, Lionel R.

    1987-05-01

    An array of five porphyrin molecules has been synthesized and used as a simple model of the light-harvesting complex found in natural photosynthesis. Efficient Förster energy transfer occurs from antenna zinc porphyrins to a central free-base porphyrin molecule. This central porphyrin retains long-lived singlet and triplet excited states that can be quenched by diffusional processes, Both electron and energy transfer quenching reactions can be observed.

  17. Programming Light-Harvesting Efficiency Using DNA Origami.

    PubMed

    Hemmig, Elisa A; Creatore, Celestino; Wünsch, Bettina; Hecker, Lisa; Mair, Philip; Parker, M Andy; Emmott, Stephen; Tinnefeld, Philip; Keyser, Ulrich F; Chin, Alex W

    2016-04-13

    The remarkable performance and quantum efficiency of biological light-harvesting complexes has prompted a multidisciplinary interest in engineering biologically inspired antenna systems as a possible route to novel solar cell technologies. Key to the effectiveness of biological "nanomachines" in light capture and energy transport is their highly ordered nanoscale architecture of photoactive molecules. Recently, DNA origami has emerged as a powerful tool for organizing multiple chromophores with base-pair accuracy and full geometric freedom. Here, we present a programmable antenna array on a DNA origami platform that enables the implementation of rationally designed antenna structures. We systematically analyze the light-harvesting efficiency with respect to number of donors and interdye distances of a ring-like antenna using ensemble and single-molecule fluorescence spectroscopy and detailed Förster modeling. This comprehensive study demonstrates exquisite and reliable structural control over multichromophoric geometries and points to DNA origami as highly versatile platform for testing design concepts in artificial light-harvesting networks. PMID:26906456

  18. Programming Light-Harvesting Efficiency Using DNA Origami

    PubMed Central

    2016-01-01

    The remarkable performance and quantum efficiency of biological light-harvesting complexes has prompted a multidisciplinary interest in engineering biologically inspired antenna systems as a possible route to novel solar cell technologies. Key to the effectiveness of biological “nanomachines” in light capture and energy transport is their highly ordered nanoscale architecture of photoactive molecules. Recently, DNA origami has emerged as a powerful tool for organizing multiple chromophores with base-pair accuracy and full geometric freedom. Here, we present a programmable antenna array on a DNA origami platform that enables the implementation of rationally designed antenna structures. We systematically analyze the light-harvesting efficiency with respect to number of donors and interdye distances of a ring-like antenna using ensemble and single-molecule fluorescence spectroscopy and detailed Förster modeling. This comprehensive study demonstrates exquisite and reliable structural control over multichromophoric geometries and points to DNA origami as highly versatile platform for testing design concepts in artificial light-harvesting networks. PMID:26906456

  19. Artificial photosynthetic reaction centers coupled to light-harvesting antennas.

    PubMed

    Ghosh, Pulak Kumar; Smirnov, Anatoly Yu; Nori, Franco

    2011-12-01

    We analyze a theoretical model for energy and electron transfer in an artificial photosynthetic system. The photosystem consists of a molecular triad (i.e., with a donor, a photosensitive unit, and an acceptor) coupled to four accessory light-harvesting-antenna pigments. The resonant energy transfer from the antennas to the artificial reaction center (the molecular triad) is described here by the Förster mechanism. We consider two different kinds of arrangements of the accessory light-harvesting pigments around the reaction center. The first arrangement allows direct excitation transfer to the reaction center from all the surrounding pigments. The second configuration transmits energy via a cascade mechanism along a chain of light-harvesting chromophores, where only one chromophore is connected to the reaction center. We show that the artificial photosynthetic system using the cascade energy transfer absorbs photons in a broader wavelength range and converts their energy into electricity with a higher efficiency than the system based on direct couplings between all the antenna chromophores and the reaction center.

  20. Thin films and assemblies of photosensitive membrane proteins and colloidal nanocrystals for engineering of hybrid materials with advanced properties.

    PubMed

    Zaitsev, Sergei Yu; Solovyeva, Daria O; Nabiev, Igor

    2012-11-15

    The development and study of nano-bio hybrid materials engineered from membrane proteins (the key functional elements of various biomembranes) and nanoheterostructures (inorganic colloidal nanoparticles, transparent electrodes, and films) is a rapidly growing field at the interface of materials and life sciences. The mainspring of the development of bioinspired materials and devices is the fact that biological evolution has solved many problems similar to those that humans are attempting to solve in the field of light-harvesting and energy-transferring inorganic compounds. Along this way, bioelectronics and biophotonics have shown considerable promise. A number of proteins have been explored in terms of bioelectronic device applications, but bacteriorhodopsin (bR, a photosensitive membrane protein from purple membranes of the bacterium Halobacterium salinarum) and bacterial photosynthetic reaction centres have received the most attention. The energy harvesting in plants has a maximum efficiency of 5%, whereas bR, in the absence of a specific light-harvesting system, allows bacteria to utilize only 0.1-0.5% of the solar light. Recent nano-bioengineering approaches employing colloidal semiconductor and metal nanoparticles conjugated with biosystems permit the enhancement of the light-harvesting capacity of photosensitive proteins, thus providing a strong impetus to protein-based device optimisation. Fabrication of ultrathin and highly oriented films from biological membranes and photosensitive proteins is the key task for prospective bioelectronic and biophotonic applications. In this review, the main advances in techniques of preparation of such films are analyzed. Comparison of the techniques for obtaining thin films leads to the conclusion that the homogeneity and orientation of biomembrane fragments or proteins in these films depend on the method of their fabrication and increase in the following order: electrophoretic sedimentation < Langmuir-Blodgett and

  1. Physiological and structural analysis of light-harvesting mutants of Rhodobacter sphaeroides.

    PubMed

    Kiley, P J; Varga, A; Kaplan, S

    1988-03-01

    Two mutants of Rhodobacter sphaeroides defective in formation of light-harvesting spectral complexes were examined in detail. Mutant RS103 lacked the B875 spectral complex despite the fact that substantial levels of the B875-alpha polypeptide (and presumably the beta polypeptide) were present. The B800-850 spectral complex was derepressed in RS103, even at high light intensities, and the growth rate was near normal at high light intensity but decreased relative to the wild type as the light intensity used for growth decreased. Mutant RS104 lacked colored carotenoids and the B800-850 spectral complex, as well as the cognate apoproteins. This strain grew normally at high light intensity and, as with RS103, the growth rate decreased as the light intensity used for growth decreased. At very low light intensities, however, RS104 would grow, whereas RS103 would not. Structural analysis of these mutants as well as others revealed that the morphology of the intracytoplasmic membrane invaginations is associated with the presence or absence of the B800-850 complex as well as of carotenoids. A low-molecular-weight intracytoplasmic membrane polypeptide, which may play a role in B800-850 complex formation, is described, as is a 62,000-dalton polypeptide whose abundance is directly related to light intensity as well as the absence of either of the light-harvesting spectral complexes. These data, obtained from studies of mutant strains and the wild type, are discussed in light of photosynthetic membrane formation and the abundance of spectral complexes per unit area of membrane. Finally, a method for the bulk preparation of the B875 complex from wild-type strain 2.4.1 is reported.

  2. The structural basis of light-harvesting in purple bacteria.

    PubMed

    Cogdell, Richard J; Isaacs, Neil W; Freer, Andrew A; Howard, Tina D; Gardiner, Alastair T; Prince, Steve M; Papiz, Miroslavr Z

    2003-11-27

    A typical purple bacterial photosynthetic unit consists of two types of light-harvesting complex (LH1 and LH2) together with a reaction centre. This short review presents a description of the structure of the LH2 complex from Rhodopseudomonas acidophila, which has recently been improved to a resolution of 2.0 A [Papiz et al., J. Mol. Biol. 326 (2003) 1523-1538]. We show how this structure has helped to reveal the details of the various excitation energy transfer events in which it is involved.

  3. Computational modeling of membrane proteins

    PubMed Central

    Leman, Julia Koehler; Ulmschneider, Martin B.; Gray, Jeffrey J.

    2014-01-01

    The determination of membrane protein (MP) structures has always trailed that of soluble proteins due to difficulties in their overexpression, reconstitution into membrane mimetics, and subsequent structure determination. The percentage of MP structures in the protein databank (PDB) has been at a constant 1-2% for the last decade. In contrast, over half of all drugs target MPs, only highlighting how little we understand about drug-specific effects in the human body. To reduce this gap, researchers have attempted to predict structural features of MPs even before the first structure was experimentally elucidated. In this review, we present current computational methods to predict MP structure, starting with secondary structure prediction, prediction of trans-membrane spans, and topology. Even though these methods generate reliable predictions, challenges such as predicting kinks or precise beginnings and ends of secondary structure elements are still waiting to be addressed. We describe recent developments in the prediction of 3D structures of both α-helical MPs as well as β-barrels using comparative modeling techniques, de novo methods, and molecular dynamics (MD) simulations. The increase of MP structures has (1) facilitated comparative modeling due to availability of more and better templates, and (2) improved the statistics for knowledge-based scoring functions. Moreover, de novo methods have benefitted from the use of correlated mutations as restraints. Finally, we outline current advances that will likely shape the field in the forthcoming decade. PMID:25355688

  4. Membrane proteins in senescent erythrocytes.

    PubMed Central

    Suzuki, T; Dale, G L

    1989-01-01

    The examination of erythrocyte senescence has been facilitated by recent advances in techniques for the isolation of aged red cells. One of these methods, which uses biotinylated rabbit erythrocytes, has been used to examine the state of membrane proteins in effete cells. These aged red cells were found to have normal ratios of alpha-spectrin and beta-spectrin as well as normal levels of ankyrin. The observation concerning ankyrin is particularly important due to the sensitivity of this protein to proteolysis and the postulated action of proteinases in the aging process. The senescent erythrocytes were also found to have an altered ratio of bands 4.1a and 4.1b without any apparent change in the total level of 4.1. In addition, the analysis of the aged cell membranes did not show any large-molecular-mass aggregated protein at the origin of the SDS/polyacrylamide gels, indicating a lack of transglutaminase activity in the senescence process for rabbit erythrocytes. These results indicate that aging of the rabbit erythrocyte is not accompanied by gross proteolytic degradation or transglutaminase-catalysed cross-linking of membrane components. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. PMID:2522000

  5. Integral Light-Harvesting Complex Expression In Symbiodinium Within The Coral Acropora aspera Under Thermal Stress

    PubMed Central

    Gierz, Sarah L.; Gordon, Benjamin R.; Leggat, William

    2016-01-01

    Coral reef success is largely dependent on the symbiosis between coral hosts and dinoflagellate symbionts belonging to the genus Symbiodinium. Elevated temperatures can result in the expulsion of Symbiodinium or loss of their photosynthetic pigments and is known as coral bleaching. It has been postulated that the expression of light-harvesting protein complexes (LHCs), which bind chlorophylls (chl) and carotenoids, are important in photobleaching. This study explored the effect a sixteen-day thermal stress (increasing daily from 25–34 °C) on integral LHC (chlorophyll a-chlorophyll c2-peridinin protein complex (acpPC)) gene expression in Symbiodinium within the coral Acropora aspera. Thermal stress leads to a decrease in Symbiodinium photosynthetic efficiency by day eight, while symbiont density was significantly lower on day sixteen. Over this time period, the gene expression of five Symbiodinium acpPC genes was quantified. Three acpPC genes exhibited up-regulated expression when corals were exposed to temperatures above 31.5 °C (acpPCSym_1:1, day sixteen; acpPCSym_15, day twelve; and acpPCSym_18, day ten and day sixteen). In contrast, the expression of acpPCSym_5:1 and acpPCSym_10:1 was unchanged throughout the experiment. Interestingly, the three acpPC genes with increased expression cluster together in a phylogenetic analysis of light-harvesting complexes. PMID:27117333

  6. Integral Light-Harvesting Complex Expression In Symbiodinium Within The Coral Acropora aspera Under Thermal Stress

    NASA Astrophysics Data System (ADS)

    Gierz, Sarah L.; Gordon, Benjamin R.; Leggat, William

    2016-04-01

    Coral reef success is largely dependent on the symbiosis between coral hosts and dinoflagellate symbionts belonging to the genus Symbiodinium. Elevated temperatures can result in the expulsion of Symbiodinium or loss of their photosynthetic pigments and is known as coral bleaching. It has been postulated that the expression of light-harvesting protein complexes (LHCs), which bind chlorophylls (chl) and carotenoids, are important in photobleaching. This study explored the effect a sixteen-day thermal stress (increasing daily from 25–34 °C) on integral LHC (chlorophyll a-chlorophyll c2-peridinin protein complex (acpPC)) gene expression in Symbiodinium within the coral Acropora aspera. Thermal stress leads to a decrease in Symbiodinium photosynthetic efficiency by day eight, while symbiont density was significantly lower on day sixteen. Over this time period, the gene expression of five Symbiodinium acpPC genes was quantified. Three acpPC genes exhibited up-regulated expression when corals were exposed to temperatures above 31.5 °C (acpPCSym_1:1, day sixteen; acpPCSym_15, day twelve; and acpPCSym_18, day ten and day sixteen). In contrast, the expression of acpPCSym_5:1 and acpPCSym_10:1 was unchanged throughout the experiment. Interestingly, the three acpPC genes with increased expression cluster together in a phylogenetic analysis of light-harvesting complexes.

  7. Integral Light-Harvesting Complex Expression In Symbiodinium Within The Coral Acropora aspera Under Thermal Stress.

    PubMed

    Gierz, Sarah L; Gordon, Benjamin R; Leggat, William

    2016-01-01

    Coral reef success is largely dependent on the symbiosis between coral hosts and dinoflagellate symbionts belonging to the genus Symbiodinium. Elevated temperatures can result in the expulsion of Symbiodinium or loss of their photosynthetic pigments and is known as coral bleaching. It has been postulated that the expression of light-harvesting protein complexes (LHCs), which bind chlorophylls (chl) and carotenoids, are important in photobleaching. This study explored the effect a sixteen-day thermal stress (increasing daily from 25-34 °C) on integral LHC (chlorophyll a-chlorophyll c2-peridinin protein complex (acpPC)) gene expression in Symbiodinium within the coral Acropora aspera. Thermal stress leads to a decrease in Symbiodinium photosynthetic efficiency by day eight, while symbiont density was significantly lower on day sixteen. Over this time period, the gene expression of five Symbiodinium acpPC genes was quantified. Three acpPC genes exhibited up-regulated expression when corals were exposed to temperatures above 31.5 °C (acpPCSym_1:1, day sixteen; acpPCSym_15, day twelve; and (acpPCSym_18), day ten and day sixteen). In contrast, the expression of acpPCSym_5:1 and acpPCSym_10:1 was unchanged throughout the experiment. Interestingly, the three acpPC genes with increased expression cluster together in a phylogenetic analysis of light-harvesting complexes. PMID:27117333

  8. Cell-free system for synthesizing membrane proteins cell free method for synthesizing membrane proteins

    DOEpatents

    Laible, Philip D; Hanson, Deborah K

    2013-06-04

    The invention provides an in vitro method for producing proteins, membrane proteins, membrane-associated proteins, and soluble proteins that interact with membrane-associated proteins for assembly into an oligomeric complex or that require association with a membrane for proper folding. The method comprises, supplying intracytoplasmic membranes from organisms; modifying protein composition of intracytoplasmic membranes from organism by modifying DNA to delete genes encoding functions of the organism not associated with the formation of the intracytoplasmic membranes; generating appropriate DNA or RNA templates that encode the target protein; and mixing the intracytoplasmic membranes with the template and a transcription/translation-competent cellular extract to cause simultaneous production of the membrane proteins and encapsulation of the membrane proteins within the intracytoplasmic membranes.

  9. The proteolysis adaptor, NblA, is essential for degradation of the core pigment of the cyanobacterial light-harvesting complex.

    PubMed

    Sendersky, Eleonora; Kozer, Noga; Levi, Mali; Moizik, Michael; Garini, Yuval; Shav-Tal, Yaron; Schwarz, Rakefet

    2015-09-01

    The cyanobacterial light-harvesting complex, the phycobilisome, is degraded under nutrient limitation, allowing the cell to adjust light absorbance to its metabolic capacity. This large light-harvesting antenna comprises a core complex of the pigment allophycocyanin, and rod-shaped pigment assemblies emanating from the core. NblA, a low-molecular-weight protein, is essential for degradation of the phycobilisome. NblA mutants exhibit high absorbance of rod pigments under conditions that generally elicit phycobilisome degradation, implicating NblA in degradation of these pigments. However, the vast abundance of rod pigments and the substantial overlap between the absorbance spectra of rod and core pigments has made it difficult to directly associate NblA with proteolysis of the phycobilisome core. Furthermore, lack of allophycocyanin degradation in an NblA mutant may reflect a requirement for rod degradation preceding core degradation, and does not prove direct involvement of NblA in proteolysis of the core pigment. Therefore, in this study, we used a mutant lacking phycocyanin, the rod pigment of Synechococcus elongatusPCC7942, to examine whether NblA is required for allophycocyanin degradation. We demonstrate that NblA is essential for degradation of the core complex of the phycobilisome. Furthermore, fluorescence lifetime imaging microscopy provided in situ evidence for the interaction of NblA with allophycocyanin, and indicated that NblA interacts with allophycocyanin complexes that are associated with the photosynthetic membranes. Based on these data, as well as previous observations indicating interaction of NblA with phycobilisomes attached to the photosynthetic membranes, we suggest a model for sequential phycobilisome disassembly by NblA. PMID:26173720

  10. Ultrafast energy relaxation in single light-harvesting complexes

    PubMed Central

    Malý, Pavel; Gruber, J. Michael; Cogdell, Richard J.; Mančal, Tomáš; van Grondelle, Rienk

    2016-01-01

    Energy relaxation in light-harvesting complexes has been extensively studied by various ultrafast spectroscopic techniques, the fastest processes being in the sub–100-fs range. At the same time, much slower dynamics have been observed in individual complexes by single-molecule fluorescence spectroscopy (SMS). In this work, we use a pump–probe-type SMS technique to observe the ultrafast energy relaxation in single light-harvesting complexes LH2 of purple bacteria. After excitation at 800 nm, the measured relaxation time distribution of multiple complexes has a peak at 95 fs and is asymmetric, with a tail at slower relaxation times. When tuning the excitation wavelength, the distribution changes in both its shape and position. The observed behavior agrees with what is to be expected from the LH2 excited states structure. As we show by a Redfield theory calculation of the relaxation times, the distribution shape corresponds to the expected effect of Gaussian disorder of the pigment transition energies. By repeatedly measuring few individual complexes for minutes, we find that complexes sample the relaxation time distribution on a timescale of seconds. Furthermore, by comparing the distribution from a single long-lived complex with the whole ensemble, we demonstrate that, regarding the relaxation times, the ensemble can be considered ergodic. Our findings thus agree with the commonly used notion of an ensemble of identical LH2 complexes experiencing slow random fluctuations. PMID:26903650

  11. Green grasses as light harvesters in dye sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Shanmugam, Vinoth; Manoharan, Subbaiah; Sharafali, A.; Anandan, Sambandam; Murugan, Ramaswamy

    2015-01-01

    Chlorophylls, the major pigments presented in plants are responsible for the process of photosynthesis. The working principle of dye sensitized solar cell (DSSC) is analogous to natural photosynthesis in light-harvesting and charge separation. In a similar way, natural dyes extracted from three types of grasses viz. Hierochloe Odorata (HO), Torulinium Odoratum (TO) and Dactyloctenium Aegyptium (DA) were used as light harvesters in dye sensitized solar cells (DSSCs). The UV-Vis absorption spectroscopy, Fourier transform infrared (FT-IR), and liquid chromatography-mass spectrometry (LC-MS) were used to characterize the dyes. The electron transport mechanism and internal resistance of the DSSCs were investigated by the electrochemical impedance spectroscopy (EIS). The performance of the cells fabricated with the grass extract shows comparable efficiencies with the reported natural dyes. Among the three types of grasses, the DSSC fabricated with the dye extracted from Hierochloe Odorata (HO) exhibited the maximum efficiency. LC-MS investigations indicated that the dominant pigment present in HO dye was pheophytin a (Pheo a).

  12. Ultrafast energy relaxation in single light-harvesting complexes.

    PubMed

    Malý, Pavel; Gruber, J Michael; Cogdell, Richard J; Mančal, Tomáš; van Grondelle, Rienk

    2016-03-15

    Energy relaxation in light-harvesting complexes has been extensively studied by various ultrafast spectroscopic techniques, the fastest processes being in the sub-100-fs range. At the same time, much slower dynamics have been observed in individual complexes by single-molecule fluorescence spectroscopy (SMS). In this work, we use a pump-probe-type SMS technique to observe the ultrafast energy relaxation in single light-harvesting complexes LH2 of purple bacteria. After excitation at 800 nm, the measured relaxation time distribution of multiple complexes has a peak at 95 fs and is asymmetric, with a tail at slower relaxation times. When tuning the excitation wavelength, the distribution changes in both its shape and position. The observed behavior agrees with what is to be expected from the LH2 excited states structure. As we show by a Redfield theory calculation of the relaxation times, the distribution shape corresponds to the expected effect of Gaussian disorder of the pigment transition energies. By repeatedly measuring few individual complexes for minutes, we find that complexes sample the relaxation time distribution on a timescale of seconds. Furthermore, by comparing the distribution from a single long-lived complex with the whole ensemble, we demonstrate that, regarding the relaxation times, the ensemble can be considered ergodic. Our findings thus agree with the commonly used notion of an ensemble of identical LH2 complexes experiencing slow random fluctuations. PMID:26903650

  13. Ultrafast energy relaxation in single light-harvesting complexes.

    PubMed

    Malý, Pavel; Gruber, J Michael; Cogdell, Richard J; Mančal, Tomáš; van Grondelle, Rienk

    2016-03-15

    Energy relaxation in light-harvesting complexes has been extensively studied by various ultrafast spectroscopic techniques, the fastest processes being in the sub-100-fs range. At the same time, much slower dynamics have been observed in individual complexes by single-molecule fluorescence spectroscopy (SMS). In this work, we use a pump-probe-type SMS technique to observe the ultrafast energy relaxation in single light-harvesting complexes LH2 of purple bacteria. After excitation at 800 nm, the measured relaxation time distribution of multiple complexes has a peak at 95 fs and is asymmetric, with a tail at slower relaxation times. When tuning the excitation wavelength, the distribution changes in both its shape and position. The observed behavior agrees with what is to be expected from the LH2 excited states structure. As we show by a Redfield theory calculation of the relaxation times, the distribution shape corresponds to the expected effect of Gaussian disorder of the pigment transition energies. By repeatedly measuring few individual complexes for minutes, we find that complexes sample the relaxation time distribution on a timescale of seconds. Furthermore, by comparing the distribution from a single long-lived complex with the whole ensemble, we demonstrate that, regarding the relaxation times, the ensemble can be considered ergodic. Our findings thus agree with the commonly used notion of an ensemble of identical LH2 complexes experiencing slow random fluctuations.

  14. Triplet Energy Transport in Platinum-Acetylide Light Harvesting Arrays.

    PubMed

    Chen, Zhuo; Hsu, Hsien-Yi; Arca, Mert; Schanze, Kirk S

    2015-06-18

    Light harvesting and triplet energy transport is investigated in chromophore-functionalized polystyrene polymers featuring light harvesting and energy acceptor chromophores (traps) at varying loading. The series of precision polymers was constructed via reversible addition-fragmentation transfer polymerization and functionalized with platinum acetylide triplet chromophores by using an azide-alkyne "click" reaction. The polymers have narrow polydispersity and degree of polymerization ∼60. The chromophores have the general structure, trans-[-R-C6H4-C≡C-Pt(PBu3)2-C≡C-Ar], where R is the attachment point to the polystyrene backbone and Ar is either -C6H4-C≡C-Ph or -pyrenyl (PE2-Pt and Py-Pt, respectively, with triplet energies of 2.35 and 1.88 eV). The polychromophores contain mainly the high-energy PE2-Pt units (light absorber and energy donor), with randomly distributed Py-Pt units (3-20% loading, energy acceptor). Photophysical methods are used to study the dynamics and efficiency of energy transport from the PE2-Pt to Py-Pt units in the polychromophores. The energy transfer efficiency is >90% for copolymers that contain 5% of the Py-Pt acceptor units. Time-resolved phosphorescence measurements combined with Monte Carlo exciton dynamics simulations suggest that the mechanism of exciton transport is exchange energy transfer hopping between PE2-Pt units.

  15. Enhanced membrane protein expression by engineering increased intracellular membrane production

    PubMed Central

    2013-01-01

    Background Membrane protein research is frequently hampered by the low natural abundance of these proteins in cells and typically relies on recombinant gene expression. Different expression systems, like mammalian cells, insect cells, bacteria and yeast are being used, but very few research efforts have been directed towards specific host cell customization for enhanced expression of membrane proteins. Here we show that by increasing the intracellular membrane production by interfering with a key enzymatic step of lipid synthesis, enhanced expression of membrane proteins in yeast is achieved. Results We engineered the oleotrophic yeast, Yarrowia lipolytica, by deleting the phosphatidic acid phosphatase, PAH1, which led to massive proliferation of endoplasmic reticulum (ER) membranes. For all eight tested representatives of different integral membrane protein families, we obtained enhanced protein accumulation levels and in some cases enhanced proteolytic integrity in the ∆pah1 strain. We analysed the adenosine A2AR G-protein coupled receptor case in more detail and found that concomitant induction of the unfolded protein response in the ∆pah1 strain enhanced the specific ligand binding activity of the receptor. These data indicate an improved quality control mechanism for membrane proteins accumulating in yeast cells with proliferated ER. Conclusions We conclude that redirecting the metabolic flux of fatty acids away from triacylglycerol- and sterylester-storage towards membrane phospholipid synthesis by PAH1 gene inactivation, provides a valuable approach to enhance eukaryotic membrane protein production. Complementary to this improvement in membrane protein quantity, UPR co-induction further enhances the quality of the membrane protein in terms of its proper folding and biological activity. Importantly, since these pathways are conserved in all eukaryotes, it will be of interest to investigate similar engineering approaches in other cell types of

  16. Dephosphorylation of Photosystem II Reaction Center Proteins in Plant Photosynthetic Membranes as an Immediate Response to Abrupt Elevation of Temperature1

    PubMed Central

    Rokka, Anne; Aro, Eva-Mari; Herrmann, Reinhold G.; Andersson, Bertil; Vener, Alexander V.

    2000-01-01

    Kinetic studies of protein dephosphorylation in photosynthetic thylakoid membranes revealed specifically accelerated dephosphorylation of photosystem II (PSII) core proteins at elevated temperatures. Raising the temperature from 22°C to 42°C resulted in a more than 10-fold increase in the dephosphorylation rates of the PSII reaction center proteins D1 and D2 and of the chlorophyll a binding protein CP43 in isolated spinach (Spinacia oleracea) thylakoids. In contrast the dephosphorylation rates of the light harvesting protein complex and the 9-kD protein of the PSII (PsbH) were accelerated only 2- to 3-fold. The use of a phospho-threonine antibody to measure in vivo phosphorylation levels in spinach leaves revealed a more than 20-fold acceleration in D1, D2, and CP43 dephosphorylation induced by abrupt elevation of temperature, but no increase in light harvesting protein complex dephosphorylation. This rapid dephosphorylation is catalyzed by a PSII-specific, intrinsic membrane protein phosphatase. Phosphatase assays, using intact thylakoids, solubilized membranes, and the isolated enzyme, revealed that the temperature-induced lateral migration of PSII to the stroma-exposed thylakoids only partially contributed to the rapid increase in the dephosphorylation rate. Significant activation of the phosphatase coincided with the temperature-induced release of TLP40 from the membrane into thylakoid lumen. TLP40 is a peptidyl-prolyl cis-trans isomerase, which acts as a regulatory subunit of the membrane phosphatase. Thus dissociation of TLP40 caused by an abrupt elevation in temperature and activation of the membrane protein phosphatase are suggested to trigger accelerated repair of photodamaged PSII and to operate as possible early signals initiating other heat shock responses in chloroplasts. PMID:10938368

  17. Membrane proteins: always an insoluble problem?

    PubMed Central

    Rawlings, Andrea E.

    2016-01-01

    Membrane proteins play crucial roles in cellular processes and are often important pharmacological drug targets. The hydrophobic properties of these proteins make full structural and functional characterization challenging because of the need to use detergents or other solubilizing agents when extracting them from their native lipid membranes. To aid membrane protein research, new methodologies are required to allow these proteins to be expressed and purified cheaply, easily, in high yield and to provide water soluble proteins for subsequent study. This mini review focuses on the relatively new area of water soluble membrane proteins and in particular two innovative approaches: the redesign of membrane proteins to yield water soluble variants and how adding solubilizing fusion proteins can help to overcome these challenges. This review also looks at naturally occurring membrane proteins, which are able to exist as stable, functional, water soluble assemblies with no alteration to their native sequence. PMID:27284043

  18. Post-transcriptional control of light-harvesting genes expression under light stress.

    PubMed

    Floris, Maïna; Bassi, Roberto; Robaglia, Christophe; Alboresi, Alessandro; Lanet, Elodie

    2013-05-01

    Plants have to deal with fluctuating light environment and the regulation of the photosynthetic apparatus is crucial for their survival. The large multigenic family of nuclear encoded chloroplastic proteins called light harvesting complex (LHC) is involved in both light harvesting and photoprotection. Changes in light intensity induce a complex set of molecular events within both the chloroplast and the cytoplasmic compartments of the cell leading to reorganization of the photosynthetic apparatus in order to optimize photosynthesis to the new conditions. In this study we have investigated the occurrence of translational regulations during light stress in Arabidopsis thaliana by using polysomes profiling. We have observed a strong effect of light on global translation activity of the cell. We show that individual LHC genes are translationally regulated in response to light conditions by changing the ratio between polysomal versus total messenger RNA. In addition, we found that cytoplasmic translational regulation can precede nuclear transcriptional regulation. Thus translational control appears as an important component of the crosstalk between chloroplast and the nucleus in plant cells.

  19. Membrane tension and peripheral protein density mediate membrane shape transitions

    NASA Astrophysics Data System (ADS)

    Shi, Zheng; Baumgart, Tobias

    2015-01-01

    Endocytosis is a ubiquitous eukaryotic membrane budding, vesiculation and internalization process fulfilling numerous roles including compensation of membrane area increase after bursts of exocytosis. The mechanism of the coupling between these two processes to enable homeostasis is not well understood. Recently, an ultrafast endocytosis (UFE) pathway was revealed with a speed significantly exceeding classical clathrin-mediated endocytosis (CME). Membrane tension reduction is a potential mechanism by which endocytosis can be rapidly activated at remote sites. Here, we provide experimental evidence for a mechanism whereby membrane tension reduction initiates membrane budding and tubulation mediated by endocytic proteins, such as endophilin A1. We find that shape instabilities occur at well-defined membrane tensions and surface densities of endophilin. From our data, we obtain a membrane shape stability diagram that shows remarkable consistency with a quantitative model. This model applies to all laterally diffusive curvature-coupling proteins and therefore a wide range of endocytic proteins.

  20. Higher plant photosystem II light-harvesting antenna, not the reaction center, determines the excited-state lifetime-both the maximum and the nonphotochemically quenched.

    PubMed

    Belgio, Erica; Johnson, Matthew P; Jurić, Snježana; Ruban, Alexander V

    2012-06-20

    The maximum chlorophyll fluorescence lifetime in isolated photosystem II (PSII) light-harvesting complex (LHCII) antenna is 4 ns; however, it is quenched to 2 ns in intact thylakoid membranes when PSII reaction centers (RCIIs) are closed (Fm). It has been proposed that the closed state of RCIIs is responsible for the quenching. We investigated this proposal using a new, to our knowledge, model system in which the concentration of RCIIs was highly reduced within the thylakoid membrane. The system was developed in Arabidopsis thaliana plants under long-term treatment with lincomycin, a chloroplast protein synthesis inhibitor. The treatment led to 1), a decreased concentration of RCIIs to 10% of the control level and, interestingly, an increased antenna component; 2), an average reduction in the yield of photochemistry to 0.2; and 3), an increased nonphotochemical chlorophyll fluorescence quenching (NPQ). Despite these changes, the average fluorescence lifetimes measured in Fm and Fm' (with NPQ) states were nearly identical to those obtained from the control. A 77 K fluorescence spectrum analysis of treated PSII membranes showed the typical features of preaggregation of LHCII, indicating that the state of LHCII antenna in the dark-adapted photosynthetic membrane is sufficient to determine the 2 ns Fm lifetime. Therefore, we conclude that the closed RCs do not cause quenching of excitation in the PSII antenna, and play no role in the formation of NPQ.

  1. How membrane surface affects protein structure.

    PubMed

    Bychkova, V E; Basova, L V; Balobanov, V A

    2014-12-01

    The immediate environment of the negatively charged membrane surface is characterized by decreased dielectric constant and pH value. These conditions can be modeled by water-alcohol mixtures at moderately low pH. Several globular proteins were investigated under these conditions, and their conformational behavior in the presence of phospholipid membranes was determined, as well as under conditions modeling the immediate environment of the membrane surface. These proteins underwent conformational transitions from the native to a molten globule-like state. Increased flexibility of the protein structure facilitated protein functioning. Our experimental data allow understanding forces that affect the structure of a protein functioning near the membrane surface (in other words, in the membrane field). Similar conformational states are widely reported in the literature. This indicates that the negatively charged membrane surface can serve as a moderately denaturing agent in the cell. We conclude that the effect of the membrane field on the protein structure must be taken into account.

  2. From light-harvesting to photoprotection: structural basis of the dynamic switch of the major antenna complex of plants (LHCII).

    PubMed

    Liguori, Nicoletta; Periole, Xavier; Marrink, Siewert J; Croce, Roberta

    2015-10-23

    Light-Harvesting Complex II (LHCII) is largely responsible for light absorption and excitation energy transfer in plants in light-limiting conditions, while in high-light it participates in photoprotection. It is generally believed that LHCII can change its function by switching between different conformations. However, the underlying molecular picture has not been elucidated yet. The available crystal structures represent the quenched form of the complex, while solubilized LHCII has the properties of the unquenched state. To determine the structural changes involved in the switch and to identify potential quenching sites, we have explored the structural dynamics of LHCII, by performing a series of microsecond Molecular Dynamics simulations. We show that LHCII in the membrane differs substantially from the crystal and has the signatures that were experimentally associated with the light-harvesting state. Local conformational changes at the N-terminus and at the xanthophyll neoxanthin are found to strongly correlate with changes in the interactions energies of two putative quenching sites. In particular conformational disorder is observed at the terminal emitter resulting in large variations of the excitonic coupling strength of this chlorophyll pair. Our results strongly support the hypothesis that light-harvesting regulation in LHCII is coupled with structural changes.

  3. From light-harvesting to photoprotection: structural basis of the dynamic switch of the major antenna complex of plants (LHCII)

    PubMed Central

    Liguori, Nicoletta; Periole, Xavier; Marrink, Siewert J.; Croce, Roberta

    2015-01-01

    Light-Harvesting Complex II (LHCII) is largely responsible for light absorption and excitation energy transfer in plants in light-limiting conditions, while in high-light it participates in photoprotection. It is generally believed that LHCII can change its function by switching between different conformations. However, the underlying molecular picture has not been elucidated yet. The available crystal structures represent the quenched form of the complex, while solubilized LHCII has the properties of the unquenched state. To determine the structural changes involved in the switch and to identify potential quenching sites, we have explored the structural dynamics of LHCII, by performing a series of microsecond Molecular Dynamics simulations. We show that LHCII in the membrane differs substantially from the crystal and has the signatures that were experimentally associated with the light-harvesting state. Local conformational changes at the N-terminus and at the xanthophyll neoxanthin are found to strongly correlate with changes in the interactions energies of two putative quenching sites. In particular conformational disorder is observed at the terminal emitter resulting in large variations of the excitonic coupling strength of this chlorophyll pair. Our results strongly support the hypothesis that light-harvesting regulation in LHCII is coupled with structural changes. PMID:26493782

  4. The phycobilisome, a light-harvesting complex responsive to environmental conditions.

    PubMed Central

    Grossman, A R; Schaefer, M R; Chiang, G G; Collier, J L

    1993-01-01

    Photosynthetic organisms can acclimate to their environment by changing many cellular processes, including the biosynthesis of the photosynthetic apparatus. In this article we discuss the phycobilisome, the light-harvesting apparatus of cyanobacteria and red algae. Unlike most light-harvesting antenna complexes, the phycobilisome is not an integral membrane complex but is attached to the surface of the photosynthetic membranes. It is composed of both the pigmented phycobiliproteins and the nonpigmented linker polypeptides; the former are important for absorbing light energy, while the latter are important for stability and assembly of the complex. The composition of the phycobilisome is very sensitive to a number of different environmental factors. Some of the filamentous cyanobacteria can alter the composition of the phycobilisome in response to the prevalent wavelengths of light in the environment. This process, called complementary chromatic adaptation, allows these organisms to efficiently utilize available light energy to drive photosynthetic electron transport and CO2 fixation. Under conditions of macronutrient limitation, many cyanobacteria degrade their phycobilisomes in a rapid and orderly fashion. Since the phycobilisome is an abundant component of the cell, its degradation may provide a substantial amount of nitrogen to nitrogen-limited cells. Furthermore, degradation of the phycobilisome during nutrient-limited growth may prevent photodamage that would occur if the cells were to absorb light under conditions of metabolic arrest. The interplay of various environmental parameters in determining the number of phycobilisomes and their structural characteristics and the ways in which these parameters control phycobilisome biosynthesis are fertile areas for investigation. PMID:8246846

  5. Modeling coherent excitation energy transfer in photosynthetic light harvesting systems

    NASA Astrophysics Data System (ADS)

    Huo, Pengfei

    2011-12-01

    Recent non-linear spectroscopy experiments suggest the excitation energy transfer in some biological light harvesting systems initially occurs coherently. Treating such processes brings significant challenge for conventional theoretical tools that usually involve different approximations. In this dissertation, the recently developed Iterative Linearized Density Matrix (ILDM) propagation scheme, which is non-perturbative and non-Markovian is extended to study coherent excitation energy transfer in various light harvesting complexes. It is demonstrated that the ILDM approach can successfully describe the coherent beating of the site populations on model systems and gives quantitative agreement with both experimental results and the results of other theoretical methods have been developed recently to going beyond the usual approximations, thus providing a new reliable theoretical tool to study this phenomenon. This approach is used to investigate the excited energy transfer dynamics in various experimentally studied bacteria light harvesting complexes, such as Fenna-Matthews-Olsen (FMO) complex, Phycocyanin 645 (PC645). In these model calculations, quantitative agreement is found between computed de-coherence times and quantum beating pattens observed in the non-linear spectroscopy. As a result of these studies, it is concluded that the stochastic resonance behavior is important in determining the optimal throughput. To begin addressing possible mechanics for observed long de-coherence time, various models which include correlation between site energy fluctuations as well as correlation between site energy and inter-site coupling are developed. The influence of both types of correlation on the coherence and transfer rate is explored using with a two state system-bath hamiltonian parametrized to model the reaction center of Rhodobacter sphaeroides bacteria. To overcome the disadvantages of a fully reduced approach or a full propagation method, a brownian dynamics

  6. Structural Implications of Hydrogen-Bond Energetics in Membrane Proteins Revealed by High-Pressure Spectroscopy

    PubMed Central

    Freiberg, Arvi; Kangur, Liina; Olsen, John D.; Hunter, C. Neil

    2012-01-01

    The light-harvesting 1 (LH1) integral membrane complex of Rhodobacter sphaeroides provides a convenient model system in which to examine the poorly understood role of hydrogen bonds (H-bonds) as stabilizing factors in membrane protein complexes. We used noncovalently bound arrays of bacteriochlorophyll chromophores within native and genetically modified variants of LH1 complexes to monitor local changes in the chromophore binding sites induced by externally applied hydrostatic pressure. Whereas membrane-bound complexes demonstrated very high resilience to pressures reaching 2.1 GPa, characteristic discontinuous shifts and broadenings of the absorption spectra were observed around 1 GPa for detergent-solubilized proteins, in similarity to those observed when specific (α or β) H-bonds between the chromophores and the surrounding protein were selectively removed by mutagenesis. These pressure effects, which were reversible upon decompression, allowed us to estimate the rupture energies of H-bonds to the chromophores in LH1 complexes. A quasi-independent, additive role of H-bonds in the α- and β-sublattices in reinforcing the wild-type LH1 complex was established. A comparison of a reaction-center-deficient LH1 complex with complexes containing reaction centers also demonstrated a stabilizing effect of the reaction center. This study thus provides important insights into the design principles of natural photosynthetic complexes. PMID:23283234

  7. Protein-Induced Membrane Curvature Alters Local Membrane Tension

    PubMed Central

    Rangamani, Padmini; Mandadap, Kranthi K.; Oster, George

    2014-01-01

    Adsorption of proteins onto membranes can alter the local membrane curvature. This phenomenon has been observed in biological processes such as endocytosis, tubulation, and vesiculation. However, it is not clear how the local surface properties of the membrane, such as membrane tension, change in response to protein adsorption. In this article, we show that the partial differential equations arising from classical elastic model of lipid membranes, which account for simultaneous changes in shape and membrane tension due to protein adsorption in a local region, cannot be solved for nonaxisymmetric geometries using straightforward numerical techniques; instead, a viscous-elastic formulation is necessary to fully describe the system. Therefore, we develop a viscous-elastic model for inhomogeneous membranes of the Helfrich type. Using the newly available viscous-elastic model, we find that the lipids flow to accommodate changes in membrane curvature during protein adsorption. We show that, at the end of protein adsorption process, the system sustains a residual local tension to balance the difference between the actual mean curvature and the imposed spontaneous curvature. We also show that this change in membrane tension can have a functional impact such as altered response to pulling forces in the presence of proteins. PMID:25099814

  8. Malate synthase a membrane protein

    SciTech Connect

    Chapman, K.D.; Turley, R.B.; Hermerath, C.A.; Carrapico, F.; Trelease, R.N.

    1987-04-01

    Malate synthase (MS) is generally regarded as a peripheral membrane protein, and believed by some to be ontogenetically associated with ER. However, immuno- and cyto-chemical in situ localizations show MS throughout the matrix of cotton (and cucumber) glyoxysomes, not specifically near their boundary membranes, nor in ER. Only a maximum of 50% MS can be solubilized from cotton glyoxysomes with 1% Triton X-100, 2mM Zwittergen 14, or 10mM DOC +/- salts. Cotton MS does not incorporate /sup 3/H-glucosamine in vivo, nor does it react with Con A on columns or blots. Cotton MS banded with ER in sucrose gradients (20-40%) in Tricine after 3h, but not after 22h in Tricine or Hepes, or after 3h in Hepes or K-phosphate. Collectively the authors data are inconsistent with physiologically meaningful MS-membrane associations in ER or glyoxysomes. It appears that experimentally-induced aggregates of MS migrate in ER gradients and occur in isolated glyoxysomes. These data indicate that ER is not involved in synthesis or modification of cottonseed MS prior to its import into the glyoxysomal matrix.

  9. Artificial light-harvesting arrays for solar energy conversion.

    PubMed

    Harriman, Anthony

    2015-07-28

    Solar fuel production, the process whereby an energy-rich substance is produced using electrons provided by water under exposure to sunlight, requires the cooperative accumulation of multiple numbers of photons. Identifying the optimum reagents is a difficult challenge, even without imposing the restriction that these same materials must function as both sensitiser and catalyst. The blockade caused by an inadequate supply of photons at the catalytic sites might be resolved by making use of an artificial light-harvesting array whose sole purpose is to funnel photons of appropriate frequency to the active catalyst, which can now be a dark reagent. Here we consider several types of artificial photon collectors built from fluorescent modules interconnected via electronic energy transfer. Emphasis is placed on the materials aspects and on establishing the basic operating principles.

  10. Dendrimer light-harvesting: intramolecular electrodynamics and mechanisms.

    PubMed

    Andrews, David L; Bradshaw, David S; Jenkins, Robert D; Rodríguez, Justo

    2009-12-01

    In the development of highly efficient materials for harvesting solar energy, there is an increasing focus on purpose-built dendrimers and allied multi-chromophore systems. A proliferation of antenna chromophores is not the only factor determining the sought light-harvesting efficiency; the internal geometry and photophysics of these molecules are also crucially important. In particular, the mechanisms by means of which radiant energy is ultimately trapped depends on an intricate interplay of electronic, structural, energetic and symmetry properties. To better understand these processes a sound theoretical representation of the intramolecular electrodynamics is required. A suitable formalism, based on quantum electrodynamics, readily delivers physical insights into the necessary excitation channelling processes, and it affords a rigorous basis for modelling the intramolecular flow of energy.

  11. Perspective: Detecting and measuring exciton delocalization in photosynthetic light harvesting

    SciTech Connect

    Scholes, Gregory D. Smyth, Cathal

    2014-03-21

    Photosynthetic units perform energy transfer remarkably well under a diverse range of demanding conditions. However, the mechanism of energy transfer, from excitation to conversion, is still not fully understood. Of particular interest is the possible role that coherence plays in this process. In this perspective, we overview photosynthetic light harvesting and discuss consequences of excitons for energy transfer and how delocalization can be assessed. We focus on challenges such as decoherence and nuclear-coordinate dependent delocalization. These approaches complement conventional spectroscopy and delocalization measurement techniques. New broadband transient absorption data may help uncover the difference between electronic and vibrational coherences present in two-dimensional electronic spectroscopy data. We describe how multipartite entanglement from quantum information theory allows us to formulate measures that elucidate the delocalization length of excitation and the details of that delocalization even from highly averaged information such as the density matrix.

  12. Functionalizing Microporous Membranes for Protein Purification and Protein Digestion

    NASA Astrophysics Data System (ADS)

    Dong, Jinlan; Bruening, Merlin L.

    2015-07-01

    This review examines advances in the functionalization of microporous membranes for protein purification and the development of protease-containing membranes for controlled protein digestion prior to mass spectrometry analysis. Recent studies confirm that membranes are superior to bead-based columns for rapid protein capture, presumably because convective mass transport in membrane pores rapidly brings proteins to binding sites. Modification of porous membranes with functional polymeric films or TiO2 nanoparticles yields materials that selectively capture species ranging from phosphopeptides to His-tagged proteins, and protein-binding capacities often exceed those of commercial beads. Thin membranes also provide a convenient framework for creating enzyme-containing reactors that afford control over residence times. With millisecond residence times, reactors with immobilized proteases limit protein digestion to increase sequence coverage in mass spectrometry analysis and facilitate elucidation of protein structures. This review emphasizes the advantages of membrane-based techniques and concludes with some challenges for their practical application.

  13. Artificial membranes for membrane protein purification, functionality and structure studies.

    PubMed

    Parmar, Mayuriben J; Lousa, Carine De Marcos; Muench, Stephen P; Goldman, Adrian; Postis, Vincent L G

    2016-06-15

    Membrane proteins represent one of the most important targets for pharmaceutical companies. Unfortunately, technical limitations have long been a major hindrance in our understanding of the function and structure of such proteins. Recent years have seen the refinement of classical approaches and the emergence of new technologies that have resulted in a significant step forward in the field of membrane protein research. This review summarizes some of the current techniques used for studying membrane proteins, with overall advantages and drawbacks for each method. PMID:27284055

  14. Tuning microbial hosts for membrane protein production

    PubMed Central

    2009-01-01

    The last four years have brought exciting progress in membrane protein research. Finally those many efforts that have been put into expression of eukaryotic membrane proteins are coming to fruition and enable to solve an ever-growing number of high resolution structures. In the past, many skilful optimization steps were required to achieve sufficient expression of functional membrane proteins. Optimization was performed individually for every membrane protein, but provided insight about commonly encountered bottlenecks and, more importantly, general guidelines how to alleviate cellular limitations during microbial membrane protein expression. Lately, system-wide analyses are emerging as powerful means to decipher cellular bottlenecks during heterologous protein production and their use in microbial membrane protein expression has grown in popularity during the past months. This review covers the most prominent solutions and pitfalls in expression of eukaryotic membrane proteins using microbial hosts (prokaryotes, yeasts), highlights skilful applications of our basic understanding to improve membrane protein production. Omics technologies provide new concepts to engineer microbial hosts for membrane protein production. PMID:20040113

  15. Crystal Dehydration in Membrane Protein Crystallography.

    PubMed

    Sanchez-Weatherby, Juan; Moraes, Isabel

    2016-01-01

    Crystal dehydration has been successfully implemented to facilitate the structural solution of a number of soluble and membrane protein structures over the years. This chapter will present the currently available tools to undertake controlled crystal dehydration, focusing on some successful membrane protein cases. Also discussed here will be some practical considerations regarding membrane protein crystals and the relationship between different techniques in order to help researchers to select the most suitable technique for their projects. PMID:27553236

  16. Serial Femtosecond Crystallography of Membrane Proteins.

    PubMed

    Zhu, Lan; Weierstall, Uwe; Cherezov, Vadim; Liu, Wei

    2016-01-01

    Membrane proteins, including G protein-coupled receptors (GPCRs), constitute the most important drug targets. The increasing number of targets requires new structural information, which has proven tremendously challenging due to the difficulties in growing diffraction-quality crystals. Recent developments of serial femtosecond crystallography at X-ray free electron lasers combined with the use of membrane-mimetic gel-like matrix of lipidic cubic phase (LCP-SFX) for crystal growth and delivery hold significant promise to accelerate structural studies of membrane proteins. This chapter describes the development and current status of the LCP-SFX technology and elaborates its future role in structural biology of membrane proteins. PMID:27553241

  17. Detection of proteins on blot transfer membranes.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2008-11-01

    Staining of blot transfer membranes permits visualization of proteins and allows the extent of transfer to be monitored. In the protocols described in this unit, proteins are stained after electroblotting from one-dimensional or two-dimensional polyacrylamide gels to blot membranes such as polyvinylidene difluoride (PVDF), nitrocellulose, or nylon membranes. Protocols are provided for the use of six general protein stains: Amido black, Coomassie blue, Ponceau S, colloidal gold, colloidal silver, and India ink. In addition, the fluorescent stains fluorescamine and IAEDANS, which covalently react with bound proteins, are described. Approximate detection limits for each nonfluorescent stain are indicated along with membrane compatibilities.

  18. Detection of proteins on blot membranes.

    PubMed

    Harper, S; Speicher, D W

    2001-05-01

    Staining of blot transfer membranes permits visualization of proteins and allows the extent of transfer to be monitored. In the protocols described in this unit, proteins are stained after electroblotting from one-dimensional or two-dimensional polyacrylamide gels to blot membranes such as polyvinylidene difluoride (PVDF), nitrocellulose, or nylon membranes. Protocols are provided for the use of six general protein stains: amido black, Coomassie blue, Ponceau S, colloidal gold, colloidal silver, and India ink. In addition, the fluorescent stains fluorescamine and IAEDANS, which covalently react with bound proteins, are described. Approximate detection limits for each nonfluorescent stain are indicated along with membrane compatibilities. PMID:18429099

  19. Detection of proteins on blot transfer membranes.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2008-11-01

    Staining of blot transfer membranes permits visualization of proteins and allows the extent of transfer to be monitored. In the protocols described in this unit, proteins are stained after electroblotting from one-dimensional or two-dimensional polyacrylamide gels to blot membranes such as polyvinylidene difluoride (PVDF), nitrocellulose, or nylon membranes. Protocols are provided for the use of six general protein stains: Amido black, Coomassie blue, Ponceau S, colloidal gold, colloidal silver, and India ink. In addition, the fluorescent stains fluorescamine and IAEDANS, which covalently react with bound proteins, are described. Approximate detection limits for each nonfluorescent stain are indicated along with membrane compatibilities. PMID:19016450

  20. Mapping membrane protein structure with fluorescence

    PubMed Central

    Taraska, Justin W.

    2012-01-01

    Membrane proteins regulate many cellular processes including signaling cascades, ion transport, membrane fusion, and cell-to-cell communications. Understanding the architecture and conformational fluctuations of these proteins is critical to understanding their regulation and functions. Fluorescence methods including intensity mapping, fluorescence resonance energy transfer, and photo-induced electron transfer, allow for targeted measurements of domains within membrane proteins. These methods can reveal how a protein is structured and how it transitions between different conformational states. Here, I will review recent work done using fluorescence to map the structures of membrane proteins, focusing on how each of these methods can be applied to understanding the dynamic nature of individual membrane proteins and protein complexes. PMID:22445227

  1. Influence of phospholipid composition on self-assembly and energy-transfer efficiency in networks of light-harvesting 2 complexes.

    PubMed

    Sumino, Ayumi; Dewa, Takehisa; Noji, Tomoyasu; Nakano, Yuki; Watanabe, Natsuko; Hildner, Richard; Bösch, Nils; Köhler, Jürgen; Nango, Mamoru

    2013-09-12

    In the photosynthetic membrane of purple bacteria networks of light-harvesting 2 (LH2) complexes capture the sunlight and transfer the excitation energy. In order to investigate the mutual relationship between the supramolecular organization of the pigment-protein complexes and their biological function, the LH2 complexes were reconstituted into three types of phospholipid membranes, consisting of L-α-phosphatidylglycerol (PG), L-α-phosphatidylcholine (PC), and L-α-phosphatidylethanolamine (PE)/PG/cardiolipin (CL). Atomic force microscopy (AFM) revealed that the type of phospholipids had a crucial influence on the clustering tendency of the LH2 complexes increased from PG over PC to PE/PG/CL, where the LH2 complexes formed large, densely packed clusters. Time-resolved spectroscopy uncovered a strong quenching of the LH2 fluorescence that is ascribed to singlet-singlet and singlet-triplet annihilation by an efficient energy transfer between the LH2 complexes in the artificial membrane systems. Quantitative analysis reveals that the intercomplex energy transfer efficiency varies strongly as a function of the morphology of the nanostructure, namely in the order PE/PG/CL > PC > PG, which is in line with the clustering tendency of LH2 observed by AFM. These results suggest a strong influence of the phospholipids on the self-assembly of LH2 complexes into networks and concomitantly on the intercomplex energy transfer efficiency.

  2. Membrane Protein Insertion at the Endoplasmic Reticulum

    PubMed Central

    Shao, Sichen; Hegde, Ramanujan S.

    2014-01-01

    Integral membrane proteins of the cell surface and most intracellular compartments of eukaryotic cells are assembled at the endoplasmic reticulum. Two highly conserved and parallel pathways mediate membrane protein targeting to and insertion into this organelle. The classical cotranslational pathway, utilized by most membrane proteins, involves targeting by the signal recognition particle followed by insertion via the Sec61 translocon. A more specialized posttranslational pathway, employed by many tail-anchored membrane proteins, is composed of entirely different factors centered around a cytosolic ATPase termed TRC40 or Get3. Both of these pathways overcome the same biophysical challenges of ferrying hydrophobic cargo through an aqueous milieu, selectively delivering it to one among several intracellular membranes and asymmetrically integrating its transmembrane domain(s) into the lipid bilayer. Here, we review the conceptual and mechanistic themes underlying these core membrane protein insertion pathways, the complexities that challenge our understanding, and future directions to over-come these obstacles. PMID:21801011

  3. Shape matters in protein mobility within membranes

    PubMed Central

    Quemeneur, François; Sigurdsson, Jon K.; Renner, Marianne; Atzberger, Paul J.; Bassereau, Patricia; Lacoste, David

    2014-01-01

    The lateral mobility of proteins within cell membranes is usually thought to be dependent on their size and modulated by local heterogeneities of the membrane. Experiments using single-particle tracking on reconstituted membranes demonstrate that protein diffusion is significantly influenced by the interplay of membrane curvature, membrane tension, and protein shape. We find that the curvature-coupled voltage-gated potassium channel (KvAP) undergoes a significant increase in protein mobility under tension, whereas the mobility of the curvature-neutral water channel aquaporin 0 (AQP0) is insensitive to it. Such observations are well explained in terms of an effective friction coefficient of the protein induced by the local membrane deformation. PMID:24706877

  4. On improving the performance of nonphotochemical quenching in CP29 light-harvesting antenna complex

    NASA Astrophysics Data System (ADS)

    Berman, Gennady P.; Nesterov, Alexander I.; Sayre, Richard T.; Still, Susanne

    2016-03-01

    We model and simulate the performance of charge-transfer in nonphotochemical quenching (NPQ) in the CP29 light-harvesting antenna-complex associated with photosystem II (PSII). The model consists of five discrete excitonic energy states and two sinks, responsible for the potentially damaging processes and charge-transfer channels, respectively. We demonstrate that by varying (i) the parameters of the chlorophyll-based dimer, (ii) the resonant properties of the protein-solvent environment interaction, and (iii) the energy transfer rates to the sinks, one can significantly improve the performance of the NPQ. Our analysis suggests strategies for improving the performance of the NPQ in response to environmental changes, and may stimulate experimental verification.

  5. Modeling Electronic-Nuclear Interactions for Excitation Energy Transfer Processes in Light-Harvesting Complexes.

    PubMed

    Lee, Mi Kyung; Coker, David F

    2016-08-18

    An accurate approach for computing intermolecular and intrachromophore contributions to spectral densities to describe the electronic-nuclear interactions relevant for modeling excitation energy transfer processes in light harvesting systems is presented. The approach is based on molecular dynamics (MD) calculations of classical correlation functions of long-range contributions to excitation energy fluctuations and a separate harmonic analysis and single-point gradient quantum calculations for electron-intrachromophore vibrational couplings. A simple model is also presented that enables detailed analysis of the shortcomings of standard MD-based excitation energy fluctuation correlation function approaches. The method introduced here avoids these problems, and its reliability is demonstrated in accurate predictions for bacteriochlorophyll molecules in the Fenna-Matthews-Olson pigment-protein complex, where excellent agreement with experimental spectral densities is found. This efficient approach can provide instantaneous spectral densities for treating the influence of fluctuations in environmental dissipation on fast electronic relaxation.

  6. Modeling Electronic-Nuclear Interactions for Excitation Energy Transfer Processes in Light-Harvesting Complexes.

    PubMed

    Lee, Mi Kyung; Coker, David F

    2016-08-18

    An accurate approach for computing intermolecular and intrachromophore contributions to spectral densities to describe the electronic-nuclear interactions relevant for modeling excitation energy transfer processes in light harvesting systems is presented. The approach is based on molecular dynamics (MD) calculations of classical correlation functions of long-range contributions to excitation energy fluctuations and a separate harmonic analysis and single-point gradient quantum calculations for electron-intrachromophore vibrational couplings. A simple model is also presented that enables detailed analysis of the shortcomings of standard MD-based excitation energy fluctuation correlation function approaches. The method introduced here avoids these problems, and its reliability is demonstrated in accurate predictions for bacteriochlorophyll molecules in the Fenna-Matthews-Olson pigment-protein complex, where excellent agreement with experimental spectral densities is found. This efficient approach can provide instantaneous spectral densities for treating the influence of fluctuations in environmental dissipation on fast electronic relaxation. PMID:27472379

  7. Sensing Membrane Stresses by Protein Insertions

    PubMed Central

    Campelo, Felix; Kozlov, Michael M.

    2014-01-01

    Protein domains shallowly inserting into the membrane matrix are ubiquitous in peripheral membrane proteins involved in various processes of intracellular membrane shaping and remodeling. It has been suggested that these domains sense membrane curvature through their preferable binding to strongly curved membranes, the binding mechanism being mediated by lipid packing defects. Here we make an alternative statement that shallow protein insertions are universal sensors of the intra-membrane stresses existing in the region of the insertion embedding rather than sensors of the curvature per se. We substantiate this proposal computationally by considering different independent ways of the membrane stress generation among which some include changes of the membrane curvature whereas others do not alter the membrane shape. Our computations show that the membrane-binding coefficient of shallow protein insertions is determined by the resultant stress independently of the way this stress has been produced. By contrast, consideration of the correlation between the insertion binding and the membrane curvature demonstrates that the binding coefficient either increases or decreases with curvature depending on the factors leading to the curvature generation. To validate our computational model, we treat quantitatively the experimental results on membrane binding by ALPS1 and ALPS2 motifs of ArfGAP1. PMID:24722359

  8. Self-Assembly Strategies for Integrating Light Harvesting and Charge Separation in Artificial Photosynthetic Systems

    SciTech Connect

    Wasielewski, Michael R.

    2011-09-28

    In natural photosynthesis, organisms optimize solar energy conversion through organized assemblies of photofunctional chromophores and catalysts within proteins that provide specifically tailored environments for chemical reactions. As with their natural counterparts, artificial photosynthetic systems for practical solar fuels production must collect light energy, separate charge, and transport charge to catalytic sites where multielectron redox processes will occur. While encouraging progress has been made on each aspect of this complex problem, researchers have not yet developed self-ordering and self-assembling components and the tailored environments necessary to realize a fully-functional artificial system. Previously researchers have used complex, covalent molecular systems comprised of chromophores, electron donors, and electron acceptors to mimic both the light-harvesting and the charge separation functions of photosynthetic proteins. These systems allow for study of the dependencies of electron transfer rate constants on donor?acceptor distance and orientation, electronic interaction, and the free energy of the reaction. The most useful and informative systems are those in which structural constraints control both the distance and the orientation between the electron donors and acceptors. Self-assembly provides a facile means for organizing large numbers of molecules into supramolecular structures that can bridge length scales from nanometers to macroscopic dimensions. The resulting structures must provide pathways for migration of light excitation energy among antenna chromophores, and from antennas to reaction centers. They also must incorporate charge conduits, that is, molecular 'wires' that can efficiently move electrons and holes between reaction centers and catalytic sites. The central scientific challenge is to develop small, functional building blocks with a minimum number of covalent linkages, which also have the appropriate molecular recognition

  9. Circular dichroism spectroscopy of membrane proteins.

    PubMed

    Miles, A J; Wallace, B A

    2016-09-21

    Circular dichroism (CD) spectroscopy is a well-established technique for studying the secondary structures, dynamics, folding pathways, and interactions of soluble proteins, and is complementary to the high resolution but generally static structures produced by X-ray crystallography, NMR spectroscopy, and cryo electron microscopy. CD spectroscopy has special relevance for the study of membrane proteins, which are difficult to crystallise and largely ignored in structural genomics projects. However, the requirement for membrane proteins to be embedded in amphipathic environments such as membranes, lipid vesicles, detergent micelles, bicelles, oriented bilayers, or nanodiscs, in order for them to be soluble or dispersed in solution whilst maintaining their structure and function, necessitates the use of different experimental and analytical approaches than those employed for soluble proteins. This review discusses specialised methods for collecting and analysing membrane protein CD data, highlighting where protocols for soluble and membrane proteins diverge.

  10. Circular dichroism spectroscopy of membrane proteins.

    PubMed

    Miles, A J; Wallace, B A

    2016-09-21

    Circular dichroism (CD) spectroscopy is a well-established technique for studying the secondary structures, dynamics, folding pathways, and interactions of soluble proteins, and is complementary to the high resolution but generally static structures produced by X-ray crystallography, NMR spectroscopy, and cryo electron microscopy. CD spectroscopy has special relevance for the study of membrane proteins, which are difficult to crystallise and largely ignored in structural genomics projects. However, the requirement for membrane proteins to be embedded in amphipathic environments such as membranes, lipid vesicles, detergent micelles, bicelles, oriented bilayers, or nanodiscs, in order for them to be soluble or dispersed in solution whilst maintaining their structure and function, necessitates the use of different experimental and analytical approaches than those employed for soluble proteins. This review discusses specialised methods for collecting and analysing membrane protein CD data, highlighting where protocols for soluble and membrane proteins diverge. PMID:27347568

  11. Phosphoinositide Control of Membrane Protein Function

    PubMed Central

    Logothetis, Diomedes E.; Petrou, Vasileios I.; Zhang, Miao; Mahajan, Rahul; Meng, Xuan-Yu; Adney, Scott K.; Cui, Meng; Baki, Lia

    2015-01-01

    Anionic phospholipids are critical constituents of the inner leaflet of the plasma membrane, ensuring appropriate membrane topology of transmembrane proteins. Additionally, in eukaryotes, the negatively charged phosphoinositides serve as key signals not only through their hydrolysis products but also through direct control of transmembrane protein function. Direct phosphoinositide control of the activity of ion channels and transporters has been the most convincing case of the critical importance of phospholipid-protein interactions in the functional control of membrane proteins. Furthermore, second messengers, such as [Ca2+]i, or posttranslational modifications, such as phosphorylation, can directly or allosterically fine-tune phospholipid-protein interactions and modulate activity. Recent advances in structure determination of membrane proteins have allowed investigators to obtain complexes of ion channels with phosphoinositides and to use computational and experimental approaches to probe the dynamic mechanisms by which lipid-protein interactions control active and inactive protein states. PMID:25293526

  12. Integral Membrane Proteins and Bilayer Proteomics

    PubMed Central

    Whitelegge, Julian P.

    2013-01-01

    Integral membrane proteins reside within the bilayer membranes that surround cells and organelles, playing critical roles in movement of molecules across them and the transduction of energy and signals. While their extreme amphipathicity presents technical challenges, biological mass spectrometry has been applied to all aspects of membrane protein chemistry and biology, including analysis of primary, secondary, tertiary and quaternary structure, as well as the dynamics that accompany functional cycles and catalysis. PMID:23301778

  13. 2-D Pigment Langmuir Monolayer Assemblies for Light Harvesting Applications.

    NASA Astrophysics Data System (ADS)

    Gregory, Brian W.; Vaknin, David; Cotton, Therese M.; Struve, Walter S.

    1996-03-01

    The use of Coulombic forces to isolate charged, water-soluble macrocycles at the air/water interface (through their interactions with the oppositely charged headgroups of a phospholipid Langmuir monolayer) is currently being exploited in this laboratory as a means to create two-dimensional arrays of pigments for light-harvesting purposes. Significant differences have been observed in the surface pressure-molecular area (π-A) isotherms of dihexadecyl phosphate on subphases containing either tetra-(N-methylaza)- phthalocyanine (i.e., tetra-(N-methyl)-2,3-pyridinoporphyrazine) or tetra- (N-methylpyridyl)-porphyrin, both of which are cationic. In situ x-ray specular reflectivity has been employed to determine interfacial organization in these systems and to elucidate the origin of their different phase behavior at the air/water interface. In addition, electronic absorption spectra and electronic linear dichroism have been utilized to determine average pigment orientation in transferred films. * Ames Laboratory is operated by Iowa State University for the U.S. Department of Energy under Cotract No. W-4705-Eng-82.

  14. Protein Solvation in Membranes and at Water-Membrane Interfaces

    NASA Technical Reports Server (NTRS)

    Pohorille, Andrew; Chipot, Christophe; Wilson, Michael A.

    2002-01-01

    Different salvation properties of water and membranes mediate a host of biologically important processes, such as folding, insertion into a lipid bilayer, associations and functions of membrane proteins. These processes will be discussed in several examples involving synthetic and natural peptides. In particular, a mechanism by which a helical peptide becomes inserted into a model membrane will be described. Further, the molecular mechanism of recognition and association of protein helical segments in membranes will be discussed. These processes are crucial for proper functioning of a cell. A membrane-spanning domain of glycophorin A, which exists as a helical dimer, serves as the model system. For this system, the free energy of dissociation of the helices is being determined for both the wild type and a mutant, in which dimerization is disrupted.

  15. Crystallization of Membrane protein under Microgravity

    NASA Astrophysics Data System (ADS)

    Henning, C.; Frank, J.; Laubender, G.; Fromme, P.

    2002-01-01

    Proteins are biological molecules which catalyse all essential reactions of cells. The knowledge on the structure of these molecular machines is necessary for the understanding of their function. Many diseases are caused by defects of membrane proteins. In order to develop new medical therapies the construction principle of the proteins must be known. The main difficulty in the determination of the structure of these membrane protein complexes is the crystallisation. Membrane proteins are normally not soluble in water and have therefore to be solubilised from the membranes by use of detergents. The whole protein-detergent micelle must be crystallised to maintain the functional integrity of the protein complexes. These difficulties are the reasons for the fact that crystals of membrane proteins are difficult to grow and most of them are badly ordered, being not appropriate for X-ray structure analysis. The crystallisation of proteins under microgravity leads to the growth of better-ordered crystals by reduction of nucleation rate and the undisturbed growth of the hovering seeds by the absence of sedimentation and convection. The successful crystallistation of a membrane protein under microgravity has been performed during the space shuttle missions USML2 and STS95 in the Space Shuttle with Photosystem I as model protein. Photosystem I is a large membrane protein complex which catalyses one of the first and fundamental steps in oxygen photosynthesis. The crystals of Photosystem I, grown under microgravity were twenty times larger than all Photosystem I crystals which have been grown on earth. They were the basis for the determination of an improved X-ray structure of Photo- system I. These experiments opened the way for the structure enlightenment of more membrane proteins on the basis of microgravity experiments. On board of the International Space Station ideal conditions for the crystallisation of proteins under zero gravity are existing.

  16. Distinct "assisted" and "spontaneous" mechanisms for the insertion of polytopic chlorophyll-binding proteins into the thylakoid membrane.

    PubMed

    Kim, S J; Jansson, S; Hoffman, N E; Robinson, C; Mant, A

    1999-02-19

    The biogenesis of several bacterial polytopic membrane proteins has been shown to require signal recognition particle (SRP) and protein transport machinery, and one such protein, the major light-harvesting chlorophyll-binding protein (LHCP) exhibits these requirements in chloroplasts. In this report we have used in vitro insertion assays to analyze four additional members of the chlorophyll-a/b-binding protein family. We show that two members, Lhca1 and Lhcb5, display an absolute requirement for stroma, nucleoside triphosphates, and protein transport apparatus, indicating an "assisted" pathway that probably resembles that of LHCP. Two other members, however, namely an early light-inducible protein 2 (Elip2) and photosystem II subunit S (PsbS), can insert efficiently in the complete absence of SRP, SecA activity, nucleoside triphosphates, or a functional Sec system. The data suggest a possibly spontaneous insertion mechanism that, to date, has been characterized only for simple single-span proteins. Of the membrane proteins whose insertion into thylakoids has been analyzed, five have now been shown to insert by a SRP/Sec-independent mechanism, suggesting that this is a mainstream form of targeting pathway. We also show that PsbS and Elip2 molecules are capable of following either "unassisted" or assisted pathways, and we discuss the implications for the mechanism and role of SRP in chloroplasts.

  17. PHOTOSYSTEM II PROTEIN33, a Protein Conserved in the Plastid Lineage, Is Associated with the Chloroplast Thylakoid Membrane and Provides Stability to Photosystem II Supercomplexes in Arabidopsis1[OPEN

    PubMed Central

    Fristedt, Rikard; Herdean, Andrei; Blaby-Haas, Crysten E.; Mamedov, Fikret; Lundin, Björn

    2015-01-01

    Photosystem II (PSII) is a multiprotein complex that catalyzes the light-driven water-splitting reactions of oxygenic photosynthesis. Light absorption by PSII leads to the production of excited states and reactive oxygen species that can cause damage to this complex. Here, we describe Arabidopsis (Arabidopsis thaliana) At1g71500, which encodes a previously uncharacterized protein that is a PSII auxiliary core protein and hence is named PHOTOSYSTEM II PROTEIN33 (PSB33). We present evidence that PSB33 functions in the maintenance of PSII-light-harvesting complex II (LHCII) supercomplex organization. PSB33 encodes a protein with a chloroplast transit peptide and one transmembrane segment. In silico analysis of PSB33 revealed a light-harvesting complex-binding motif within the transmembrane segment and a large surface-exposed head domain. Biochemical analysis of PSII complexes further indicates that PSB33 is an integral membrane protein located in the vicinity of LHCII and the PSII CP43 reaction center protein. Phenotypic characterization of mutants lacking PSB33 revealed reduced amounts of PSII-LHCII supercomplexes, very low state transition, and a lower capacity for nonphotochemical quenching, leading to increased photosensitivity in the mutant plants under light stress. Taken together, these results suggest a role for PSB33 in regulating and optimizing photosynthesis in response to changing light levels. PMID:25511433

  18. Outer membrane protein profiles of Haemophilus pleuropneumoniae.

    PubMed Central

    Rapp, V J; Munson, R S; Ross, R F

    1986-01-01

    Outer membrane protein profiles of Haemophilus pleuropneumoniae were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cells were disrupted by sonication, and outer membrane-enriched fractions were prepared by differential centrifugation and selective solubilization of the inner membrane with sodium N-lauroyl sarcosinate. Colony type, growth medium, time of harvest, and in vitro or in vivo passage had no appreciable effect on the protein profiles of the strains examined. Seven patterns were distinguished among the reference strains of the nine capsular serotypes. These patterns were based on the mobility of the major outer membrane proteins migrating in the 39,000- to 44,000-molecular-weight region of the gel, a 16K to 16.5K protein, and a heat-modifiable 29K protein. Strains of serotypes 1 and 9 had identical outer membrane protein profiles, as did strains of serotypes 2 and 6. The reference strains of the remaining five serotypes each had a distinct pattern. The outer membrane protein profiles of 95 field isolates belonging to serotypes 1, 5, 7, and 9 from swine in the midwestern United States were determined and compared with the reference patterns. The results indicate that the population of H. pleuropneumoniae is clonal, with three predominant clones distinguished by both serotype and outer membrane protein profile responsible for the majority of H. pleuropneumoniae disease occurring in swine in the United States. Images PMID:3699889

  19. Genome-wide Membrane Protein Structure Prediction

    PubMed Central

    Piccoli, Stefano; Suku, Eda; Garonzi, Marianna; Giorgetti, Alejandro

    2013-01-01

    Transmembrane proteins allow cells to extensively communicate with the external world in a very accurate and specific way. They form principal nodes in several signaling pathways and attract large interest in therapeutic intervention, as the majority pharmaceutical compounds target membrane proteins. Thus, according to the current genome annotation methods, a detailed structural/functional characterization at the protein level of each of the elements codified in the genome is also required. The extreme difficulty in obtaining high-resolution three-dimensional structures, calls for computational approaches. Here we review to which extent the efforts made in the last few years, combining the structural characterization of membrane proteins with protein bioinformatics techniques, could help describing membrane proteins at a genome-wide scale. In particular we analyze the use of comparative modeling techniques as a way of overcoming the lack of high-resolution three-dimensional structures in the human membrane proteome. PMID:24403851

  20. Surface-Cross-Linked Micelles as Multifunctionalized Organic Nanoparticles for Controlled Release, Light Harvesting, and Catalysis

    PubMed Central

    2016-01-01

    Surfactant micelles are dynamic entities with a rapid exchange of monomers. By “clicking” tripropargylammonium-containing surfactants with diazide cross-linkers, we obtained surface-cross-linked micelles (SCMs) that could be multifunctionalized for different applications. They triggered membrane fusion through tunable electrostatic interactions with lipid bilayers. Antenna chromophores could be installed on them to create artificial light-harvesting complexes with efficient energy migration among tens to hundreds of chromophores. When cleavable cross-linkers were used, the SCMs could break apart in response to redox or pH signals, ejecting entrapped contents quickly as a result of built-in electrostatic stress. They served as caged surfactants whose surface activity was turned on by environmental stimuli. They crossed cell membranes readily. Encapsulated fluorophores showed enhanced photophysical properties including improved quantum yields and greatly expanded Stokes shifts. Catalytic groups could be installed on the surface or in the interior, covalently attached or physically entrapped. As enzyme mimics, the SCMs enabled rational engineering of the microenvironment around the catalysts to afford activity and selectivity not possible with conventional catalysts. PMID:27181610

  1. Surface-Cross-Linked Micelles as Multifunctionalized Organic Nanoparticles for Controlled Release, Light Harvesting, and Catalysis.

    PubMed

    Zhao, Yan

    2016-06-14

    Surfactant micelles are dynamic entities with a rapid exchange of monomers. By "clicking" tripropargylammonium-containing surfactants with diazide cross-linkers, we obtained surface-cross-linked micelles (SCMs) that could be multifunctionalized for different applications. They triggered membrane fusion through tunable electrostatic interactions with lipid bilayers. Antenna chromophores could be installed on them to create artificial light-harvesting complexes with efficient energy migration among tens to hundreds of chromophores. When cleavable cross-linkers were used, the SCMs could break apart in response to redox or pH signals, ejecting entrapped contents quickly as a result of built-in electrostatic stress. They served as caged surfactants whose surface activity was turned on by environmental stimuli. They crossed cell membranes readily. Encapsulated fluorophores showed enhanced photophysical properties including improved quantum yields and greatly expanded Stokes shifts. Catalytic groups could be installed on the surface or in the interior, covalently attached or physically entrapped. As enzyme mimics, the SCMs enabled rational engineering of the microenvironment around the catalysts to afford activity and selectivity not possible with conventional catalysts. PMID:27181610

  2. Evolutionary changes in chlorophyllide a oxygenase (CAO) structure contribute to the acquisition of a new light-harvesting complex in micromonas.

    PubMed

    Kunugi, Motoshi; Takabayashi, Atsushi; Tanaka, Ayumi

    2013-07-01

    Chlorophyll b is found in photosynthetic prokaryotes and primary and secondary endosymbionts, although their light-harvesting systems are quite different. Chlorophyll b is synthesized from chlorophyll a by chlorophyllide a oxygenase (CAO), which is a Rieske-mononuclear iron oxygenase. Comparison of the amino acid sequences of CAO among photosynthetic organisms elucidated changes in the domain structures of CAO during evolution. However, the evolutionary relationship between the light-harvesting system and the domain structure of CAO remains unclear. To elucidate this relationship, we investigated the CAO structure and the pigment composition of chlorophyll-protein complexes in the prasinophyte Micromonas. The Micromonas CAO is composed of two genes, MpCAO1 and MpCAO2, that possess Rieske and mononuclear iron-binding motifs, respectively. Only when both genes were introduced into the chlorophyll b-less Arabidopsis mutant (ch1-1) was chlorophyll b accumulated, indicating that cooperation between the two subunits is required to synthesize chlorophyll b. Although Micromonas has a characteristic light-harvesting system in which chlorophyll b is incorporated into the core antennas of reaction centers, chlorophyll b was also incorporated into the core antennas of reaction centers of the Arabidopsis transformants that contained the two Micromonas CAO proteins. Based on these results, we discuss the evolutionary relationship between the structures of CAO and light-harvesting systems.

  3. Active Nuclear Import of Membrane Proteins Revisited.

    PubMed

    Laba, Justyna K; Steen, Anton; Popken, Petra; Chernova, Alina; Poolman, Bert; Veenhoff, Liesbeth M

    2015-01-01

    It is poorly understood how membrane proteins destined for the inner nuclear membrane pass the crowded environment of the Nuclear Pore Complex (NPC). For the Saccharomyces cerevisiae proteins Src1/Heh1 and Heh2, a transport mechanism was proposed where the transmembrane domains diffuse through the membrane while the extralumenal domains encoding a nuclear localization signal (NLS) and intrinsically disordered linker (L) are accompanied by transport factors and travel through the NPC. Here, we validate the proposed mechanism and explore and discuss alternative interpretations of the data. First, to disprove an interpretation where the membrane proteins become membrane embedded only after nuclear import, we present biochemical and localization data to support that the previously used, as well as newly designed reporter proteins are membrane-embedded irrespective of the presence of the sorting signals, the specific transmembrane domain (multipass or tail anchored), independent of GET, and also under conditions that the proteins are trapped in the NPC. Second, using the recently established size limit for passive diffusion of membrane proteins in yeast, and using an improved assay, we confirm active import of polytopic membrane protein with extralumenal soluble domains larger than those that can pass by diffusion on similar timescales. This reinforces that NLS-L dependent active transport is distinct from passive diffusion. Thirdly, we revisit the proposed route through the center of the NPC and conclude that the previously used trapping assay is, unfortunately, poorly suited to address the route through the NPC, and the route thus remains unresolved. Apart from the uncertainty about the route through the NPC, the data confirm active, transport factor dependent, nuclear transport of membrane-embedded mono- and polytopic membrane proteins in baker's yeast. PMID:26473931

  4. Active Nuclear Import of Membrane Proteins Revisited

    PubMed Central

    Laba, Justyna K.; Steen, Anton; Popken, Petra; Chernova, Alina; Poolman, Bert; Veenhoff, Liesbeth M.

    2015-01-01

    It is poorly understood how membrane proteins destined for the inner nuclear membrane pass the crowded environment of the Nuclear Pore Complex (NPC). For the Saccharomyces cerevisiae proteins Src1/Heh1 and Heh2, a transport mechanism was proposed where the transmembrane domains diffuse through the membrane while the extralumenal domains encoding a nuclear localization signal (NLS) and intrinsically disordered linker (L) are accompanied by transport factors and travel through the NPC. Here, we validate the proposed mechanism and explore and discuss alternative interpretations of the data. First, to disprove an interpretation where the membrane proteins become membrane embedded only after nuclear import, we present biochemical and localization data to support that the previously used, as well as newly designed reporter proteins are membrane-embedded irrespective of the presence of the sorting signals, the specific transmembrane domain (multipass or tail anchored), independent of GET, and also under conditions that the proteins are trapped in the NPC. Second, using the recently established size limit for passive diffusion of membrane proteins in yeast, and using an improved assay, we confirm active import of polytopic membrane protein with extralumenal soluble domains larger than those that can pass by diffusion on similar timescales. This reinforces that NLS-L dependent active transport is distinct from passive diffusion. Thirdly, we revisit the proposed route through the center of the NPC and conclude that the previously used trapping assay is, unfortunately, poorly suited to address the route through the NPC, and the route thus remains unresolved. Apart from the uncertainty about the route through the NPC, the data confirm active, transport factor dependent, nuclear transport of membrane-embedded mono- and polytopic membrane proteins in baker’s yeast. PMID:26473931

  5. AplA, a member of a new class of phycobiliproteins lacking a traditional role in photosynthetic light harvesting.

    PubMed

    Montgomery, Beronda L; Casey, Elena Silva; Grossman, Arthur R; Kehoe, David M

    2004-11-01

    All known phycobiliproteins have light-harvesting roles during photosynthesis and are found in water-soluble phycobilisomes, the light-harvesting complexes of cyanobacteria, cyanelles, and red algae. Phycobiliproteins are chromophore-bearing proteins that exist as heterodimers of alpha and beta subunits, possess a number of highly conserved amino acid residues important for dimerization and chromophore binding, and are invariably 160 to 180 amino acids long. A new and unusual group of proteins that is most closely related to the allophycocyanin members of the phycobiliprotein superfamily has been identified. Each of these proteins, which have been named allophycocyanin-like (Apl) proteins, apparently contains a 28-amino-acid extension at its amino terminus relative to allophycocyanins. Apl family members possess the residues critical for chromophore interactions, but substitutions are present at positions implicated in maintaining the proper alpha-beta subunit interactions and tertiary structure of phycobiliproteins, suggesting that Apl proteins are able to bind chromophores but fail to adopt typical allophycocyanin conformations. AplA isolated from the cyanobacterium Fremyella diplosiphon contained a covalently attached chromophore and, although present in the cell under a number of conditions, was not detected in phycobilisomes. Thus, Apl proteins are a new class of photoreceptors with a different cellular location and structure than any previously described members of the phycobiliprotein superfamily.

  6. Light stress photodynamics of chlorophyll-binding proteins in Arabidopsis thaliana thylakoid membranes revealed by high-resolution mass spectrometric studies.

    PubMed

    Galetskiy, D N; Lohscheider, J N; Kononikhin, A S; Kharybin, O N; Popov, I A; Adamska, I; Nikolaev, E N

    2011-01-01

    In higher plants the light energy is captured by the photosynthetic pigments that are bound to photosystem I and II and their light-harvesting complex (LHC) subunits. In this study, we examined the photodynamic changes within chlorophyll-protein complexes in the thylakoid membrane of Arabidopsis thaliana leaves adapted to low light and subsequently exposed to light stress. Chlorophyll-protein complexes were isolated using sucrose density gradient centrifugation and blue-native polyacrylamid gel electrophoresis (BN-PAGE). Proteome analysis was performed using SDS-PAGE, HPLC and high resolution mass spectrometry. We identified several rarely expressed and stress-induced chlorophyll-binding proteins, showed changes in localization of early light-induced protein family and LHC protein family members between different photosynthetic complexes and assembled/disassembled subcomplexes under light stress conditions and discuss their role in a variety of light stress-related processes.

  7. Light stress photodynamics of chlorophyll-binding proteins in Arabidopsis thaliana thylakoid membranes revealed by high-resolution mass spectrometric studies.

    PubMed

    Galetskiy, D N; Lohscheider, J N; Kononikhin, A S; Kharybin, O N; Popov, I A; Adamska, I; Nikolaev, E N

    2011-01-01

    In higher plants the light energy is captured by the photosynthetic pigments that are bound to photosystem I and II and their light-harvesting complex (LHC) subunits. In this study, we examined the photodynamic changes within chlorophyll-protein complexes in the thylakoid membrane of Arabidopsis thaliana leaves adapted to low light and subsequently exposed to light stress. Chlorophyll-protein complexes were isolated using sucrose density gradient centrifugation and blue-native polyacrylamid gel electrophoresis (BN-PAGE). Proteome analysis was performed using SDS-PAGE, HPLC and high resolution mass spectrometry. We identified several rarely expressed and stress-induced chlorophyll-binding proteins, showed changes in localization of early light-induced protein family and LHC protein family members between different photosynthetic complexes and assembled/disassembled subcomplexes under light stress conditions and discuss their role in a variety of light stress-related processes. PMID:21460887

  8. Helical Membrane Protein Conformations and their Environment

    PubMed Central

    Cross, Timothy A.; Murray, Dylan T.; Watts, Anthony

    2013-01-01

    Evidence that membrane proteins respond conformationally and functionally to their environment is gaining pace. Structural models, by necessity, have been characterized in preparations where the protein has been removed from its native environment. Different structural methods have used various membrane mimetics that have recently included lipid bilayers as a more native-like environment. Structural tools applied to lipid bilayer-embedded integral proteins are informing us about important generic characteristics of how membrane proteins respond to the lipid environment as compared with their response to other non-lipid environments. Here, we review the current status of the field, with specific reference to observations of some well-studied α-helical membrane proteins, as a starting point to aid the development of possible generic principals for model refinement. PMID:23996195

  9. Membrane protein expression in Lactococcus lactis.

    PubMed

    King, Martin S; Boes, Christoph; Kunji, Edmund R S

    2015-01-01

    The Gram-positive bacterium Lactococcus lactis has many properties that are ideal for the overproduction of membrane proteins in a functional form. Growth of lactococci is rapid, proceeds to high cell densities, and does not require aeration, which facilitates large-scale fermentation. The available promoter systems are strong and tightly regulated, allowing expression of toxic gene products in a controlled manner. Expressed membrane proteins are targeted exclusively to the cytoplasmic membrane, allowing the use of ionophores, ligands, and inhibitors to study activity of the membrane protein in whole cells. Constructed plasmids are stable and expression levels are highly reproducible. The relatively small genome size of the organism causes little redundancy, which facilitates complementation studies and allows for easier purification. The produced membrane proteins are often stable, as the organism has limited proteolytic capability, and they are readily solubilized from the membrane with mild detergents. Lactococci are multiple amino acid auxotrophs, allowing the incorporation of labels, such as selenomethionine. Among the few disadvantages are the low transformation frequency, AT-rich codon usage, and resistance to lysis by mechanical means, but these problems can be overcome fairly easily. We will describe in detail the protocols used to express membrane proteins in L. lactis, from cloning of the target gene to the isolation of membrane vesicles for the determination of expression levels. PMID:25857778

  10. Biophysical EPR Studies Applied to Membrane Proteins

    PubMed Central

    Sahu, Indra D; Lorigan, Gary A

    2015-01-01

    Membrane proteins are very important in controlling bioenergetics, functional activity, and initializing signal pathways in a wide variety of complicated biological systems. They also represent approximately 50% of the potential drug targets. EPR spectroscopy is a very popular and powerful biophysical tool that is used to study the structural and dynamic properties of membrane proteins. In this article, a basic overview of the most commonly used EPR techniques and examples of recent applications to answer pertinent structural and dynamic related questions on membrane protein systems will be presented. PMID:26855825

  11. Strong antenna-enhanced fluorescence of a single light-harvesting complex shows photon antibunching

    PubMed Central

    Wientjes, Emilie; Renger, Jan; Curto, Alberto G.; Cogdell, Richard; van Hulst, Niek F.

    2014-01-01

    The nature of the highly efficient energy transfer in photosynthetic light-harvesting complexes is a subject of intense research. Unfortunately, the low fluorescence efficiency and limited photostability hampers the study of individual light-harvesting complexes at ambient conditions. Here we demonstrate an over 500-fold fluorescence enhancement of light-harvesting complex 2 (LH2) at the single-molecule level by coupling to a gold nanoantenna. The resonant antenna produces an excitation enhancement of circa 100 times and a fluorescence lifetime shortening to ~\

  12. Membrane Protein Structure Determination Using Crystallography and Lipidic Mesophases - Recent Advances and Successes

    PubMed Central

    Caffrey, Martin; Li, Dianfan; Dukkipati, Abhiram

    2012-01-01

    The crystal structure of the β2-adrenergic receptor in complex with an agonist and its cognate G protein has just recently been solved. It is now possible to explore in molecular detail the means by which this paradigmatic transmembrane receptor binds agonist, communicates the impulse or signalling event across the membrane and sets in motion a series of G protein-directed intracellular responses. The structure was determined using crystals of the ternary complex grown in a rationally designed lipidic mesophase by the so-called in meso method. The method is proving to be particularly useful in the G protein-coupled receptor field where the structures of thirteen distinct receptor types have been solved in the past five years. In addition to receptors, the method has proven useful with a wide variety of integral membrane protein classes that include bacterial and eukaryotic rhodopsins, a light harvesting complex II (LHII), photosynthetic reaction centers, cytochrome oxidases, β-barrels, an exchanger, and an integral membrane peptide. This attests to the versatility and range of the method and supports the view that the in meso method should be included in the arsenal of the serious membrane structural biologist. For this to happen however, the reluctance in adopting it attributable, in part, to the anticipated difficulties associated with handling the sticky, viscous cubic mesophase in which crystals grow must be overcome. Harvesting and collecting diffraction data with the mesophase-grown crystals is also viewed with some trepidation. It is acknowledged that there are challenges associated with the method. Over the years, we have endeavored to establish how the method works at a molecular level and to make it user-friendly. To these ends, tools for handling the mesophase in the pico- to nano-liter volume range have been developed for highly efficient crystallization screening in manual and robotic modes. Methods have been implemented for evaluating the functional

  13. Flagellar membrane proteins in kinetoplastid parasites.

    PubMed

    Landfear, Scott M; Tran, Khoa D; Sanchez, Marco A

    2015-09-01

    All kinetoplastid parasites, including protozoa such as Leishmania species, Trypanosoma brucei, and Trypanosoma cruzi that cause devastating diseases in humans and animals, are flagellated throughout their life cycles. Although flagella were originally thought of primarily as motility organelles, flagellar functions in other critical processes, especially in sensing and signal transduction, have become more fully appreciated in the recent past. The flagellar membrane is a highly specialized subdomain of the surface membrane, and flagellar membrane proteins are likely to be critical components for all the biologically important roles of flagella. In this review, we summarize recent discoveries relevant to flagellar membrane proteins in these parasites, including the identification of such proteins, investigation of their biological functions, and mechanisms of selective trafficking to the flagellar membrane. Prospects for future investigations and current unsolved problems are highlighted.

  14. Membrane proteins: is the future disc shaped?

    PubMed

    Lee, Sarah C; Pollock, Naomi L

    2016-08-15

    The use of styrene maleic acid lipid particles (SMALPs) for the purification of membrane proteins (MPs) is a rapidly developing technology. The amphiphilic copolymer of styrene and maleic acid (SMA) disrupts biological membranes and can extract membrane proteins in nanodiscs of approximately 10 nm diameter. These discs contain SMA, protein and membrane lipids. There is evidence that MPs in SMALPs retain their native structures and functions, in some cases with enhanced thermal stability. In addition, the method is compatible with biological buffers and a wide variety of biophysical and structural analysis techniques. The use of SMALPs to solubilize and stabilize MPs offers a new approach in our attempts to understand, and influence, the structure and function of MPs and biological membranes. In this review, we critically assess progress with this method, address some of the associated technical challenges, and discuss opportunities for exploiting SMA and SMALPs to expand our understanding of MP biology. PMID:27528746

  15. Flagellar membrane proteins in kinetoplastid parasites.

    PubMed

    Landfear, Scott M; Tran, Khoa D; Sanchez, Marco A

    2015-09-01

    All kinetoplastid parasites, including protozoa such as Leishmania species, Trypanosoma brucei, and Trypanosoma cruzi that cause devastating diseases in humans and animals, are flagellated throughout their life cycles. Although flagella were originally thought of primarily as motility organelles, flagellar functions in other critical processes, especially in sensing and signal transduction, have become more fully appreciated in the recent past. The flagellar membrane is a highly specialized subdomain of the surface membrane, and flagellar membrane proteins are likely to be critical components for all the biologically important roles of flagella. In this review, we summarize recent discoveries relevant to flagellar membrane proteins in these parasites, including the identification of such proteins, investigation of their biological functions, and mechanisms of selective trafficking to the flagellar membrane. Prospects for future investigations and current unsolved problems are highlighted. PMID:26599841

  16. Flagellar Membrane Proteins in Kinetoplastid Parasites

    PubMed Central

    Landfear, Scott M.; Tran, Khoa D.; Sanchez, Marco A.

    2015-01-01

    All kinetoplastid parasites, including protozoa such as Leishmania species, Trypanosoma brucei, and Trypanosoma cruzi that cause devastating diseases in humans and animals, are flagellated throughout their life cycles. While flagella were originally thought of primarily as motility organelles, flagellar functions in other critical processes, especially in sensing and signal transduction, have become more fully appreciated in the recent past. The flagellar membrane is a highly specialized subdomain of the surface membrane, and flagellar membrane proteins are likely to be critical components for all the biologically important roles of flagella. In this review, we summarize recent discoveries relevant to flagellar membrane proteins in these parasites including the identification of such proteins, investigation of their biological functions, and mechanisms of selective trafficking to the flagellar membrane. Prospects for future investigations and current unsolved problems are highlighted. PMID:26599841

  17. Conformational switching explains the intrinsic multifunctionality of plant light-harvesting complexes.

    PubMed

    Krüger, Tjaart P J; Wientjes, Emilie; Croce, Roberta; van Grondelle, Rienk

    2011-08-16

    The light-harvesting complexes of photosystem I and II (Lhcas and Lhcbs) of plants display a high structural homology and similar pigment content and organization. Yet, the spectroscopic properties of these complexes, and accordingly their functionality, differ substantially. This difference is primarily due to the charge-transfer (CT) character of a chlorophyll dimer in all Lhcas, which mixes with the excitonic states of these complexes, whereas this CT character is generally absent in Lhcbs. By means of single-molecule spectroscopy near room temperature, we demonstrate that the presence or absence of such a CT state in Lhcas and Lhcbs can occasionally be reversed; i.e., these complexes are able to interconvert conformationally to quasi-stable spectral states that resemble the Lhcs of the other photosystem. The high structural similarity of all the Lhca and Lhcb proteins suggests that the stable conformational states that give rise to the mixed CT-excitonic state are similar for all these proteins, and similarly for the conformations that involve no CT state. This indicates that the specific functions related to Lhca and Lhcb complexes are realized by different stable conformations of a single generic protein structure. We propose that this functionality is modulated and controlled by the protein environment.

  18. Bacterial Light-Harvesting Complexes Showing Giant Second-Order Nonlinear Optical Response as Revealed by Hyper-Rayleigh Light Scattering.

    PubMed

    Ma, Fei; Yu, Long-Jiang; Ma, Xiao-Hua; Wang, Peng; Wang-Otomo, Zheng-Yu; Zhang, Jian-Ping

    2016-09-01

    The second-order nonlinear optical (NLO) properties of light-harvesting complexes (LHs) from the purple photosynthetic bacteria Thermochromatium (Tch.) tepidum were investigated for the first time by means of hyper-Rayleigh scattering (HRS). The carotenoid (Car) molecules bound to the isolated LH1 and LH2 proteins gave rise to second-harmonic scattering; however, they showed an opposite effect of the collective contribution from Car, that is, the first hyperpolarizability (β) reduced substantially from (10 510 ± 370) × 10(-30) esu for LH1 to (360 ± 120) × 10(-30) esu for LH2. Chromatophores of Tch. tepidum also showed a giant hyperpolarizability of (11 640 ± 630) × 10(-30) esu. On the basis of the structural information on bacterial LHs, it is found that the effective β of an LH is governed by the microenvironment and orientational correlation among the Car chromophores, which is concluded to be coherently enhanced for LH1. For LH2, however, additional destructive effects between different Car molecules may account for the small β value. This work demonstrates that LH1 and native membranes of purple bacteria can be potent NLO materials and that HRS is a promising spectroscopic means for investigating structural information of pigment-protein supramolecules. PMID:27505442

  19. STAY-GREEN and Chlorophyll Catabolic Enzymes Interact at Light-Harvesting Complex II for Chlorophyll Detoxification during Leaf Senescence in Arabidopsis[C][W

    PubMed Central

    Sakuraba, Yasuhito; Schelbert, Silvia; Park, So-Yon; Han, Su-Hyun; Lee, Byoung-Doo; Andrès, Céline Besagni; Kessler, Felix; Hörtensteiner, Stefan; Paek, Nam-Chon

    2012-01-01

    During leaf senescence, plants degrade chlorophyll to colorless linear tetrapyrroles that are stored in the vacuole of senescing cells. The early steps of chlorophyll breakdown occur in plastids. To date, five chlorophyll catabolic enzymes (CCEs), NONYELLOW COLORING1 (NYC1), NYC1-LIKE, pheophytinase, pheophorbide a oxygenase (PAO), and red chlorophyll catabolite reductase, have been identified; these enzymes catalyze the stepwise degradation of chlorophyll to a fluorescent intermediate, pFCC, which is then exported from the plastid. In addition, STAY-GREEN (SGR), Mendel’s green cotyledon gene encoding a chloroplast protein, is required for the initiation of chlorophyll breakdown in plastids. Senescence-induced SGR binds to light-harvesting complex II (LHCII), but its exact role remains elusive. Here, we show that all five CCEs also specifically interact with LHCII. In addition, SGR and CCEs interact directly or indirectly with each other at LHCII, and SGR is essential for recruiting CCEs in senescing chloroplasts. PAO, which had been attributed to the inner envelope, is found to localize in the thylakoid membrane. These data indicate a predominant role for the SGR-CCE-LHCII protein interaction in the breakdown of LHCII-located chlorophyll, likely to allow metabolic channeling of phototoxic chlorophyll breakdown intermediates upstream of nontoxic pFCC. PMID:22366162

  20. Detergents in Membrane Protein Purification and Crystallisation.

    PubMed

    Anandan, Anandhi; Vrielink, Alice

    2016-01-01

    Detergents play a significant role in structural and functional characterisation of integral membrane proteins (IMPs). IMPs reside in the biological membranes and exhibit a great variation in their structural and physical properties. For in vitro biophysical studies, structural and functional analyses, IMPs need to be extracted from the membrane lipid bilayer environment in which they are found and purified to homogeneity while maintaining a folded and functionally active state. Detergents are capable of successfully solubilising and extracting the IMPs from the membrane bilayers. A number of detergents with varying structure and physicochemical properties are commercially available and can be applied for this purpose. Nevertheless, it is important to choose a detergent that is not only able to extract the membrane protein but also provide an optimal environment while retaining the correct structural and physical properties of the protein molecule. Choosing the best detergent for this task can be made possible by understanding the physical and chemical properties of the different detergents and their interaction with the IMPs. In addition, understanding the mechanism of membrane solubilisation and protein extraction along with crystallisation requirements, if crystallographic studies are going to be undertaken, can help in choosing the best detergent for the purpose. This chapter aims to present the fundamental properties of detergents and highlight information relevant to IMP crystallisation. The first section of the chapter reviews the physicochemical properties of detergents and parameters essential for predicting their behaviour in solution. The second section covers the interaction of detergents with the biologic membranes and proteins followed by their role in membrane protein crystallisation. The last section will briefly cover the types of detergent and their properties focusing on custom designed detergents for membrane protein studies.

  1. Detergents in Membrane Protein Purification and Crystallisation.

    PubMed

    Anandan, Anandhi; Vrielink, Alice

    2016-01-01

    Detergents play a significant role in structural and functional characterisation of integral membrane proteins (IMPs). IMPs reside in the biological membranes and exhibit a great variation in their structural and physical properties. For in vitro biophysical studies, structural and functional analyses, IMPs need to be extracted from the membrane lipid bilayer environment in which they are found and purified to homogeneity while maintaining a folded and functionally active state. Detergents are capable of successfully solubilising and extracting the IMPs from the membrane bilayers. A number of detergents with varying structure and physicochemical properties are commercially available and can be applied for this purpose. Nevertheless, it is important to choose a detergent that is not only able to extract the membrane protein but also provide an optimal environment while retaining the correct structural and physical properties of the protein molecule. Choosing the best detergent for this task can be made possible by understanding the physical and chemical properties of the different detergents and their interaction with the IMPs. In addition, understanding the mechanism of membrane solubilisation and protein extraction along with crystallisation requirements, if crystallographic studies are going to be undertaken, can help in choosing the best detergent for the purpose. This chapter aims to present the fundamental properties of detergents and highlight information relevant to IMP crystallisation. The first section of the chapter reviews the physicochemical properties of detergents and parameters essential for predicting their behaviour in solution. The second section covers the interaction of detergents with the biologic membranes and proteins followed by their role in membrane protein crystallisation. The last section will briefly cover the types of detergent and their properties focusing on custom designed detergents for membrane protein studies. PMID:27553232

  2. Mass Spectrometry of Intact Membrane Protein Complexes

    PubMed Central

    Laganowsky, Arthur; Reading, Eamonn; Hopper, Jonathan T.S.; Robinson, Carol V.

    2014-01-01

    Mass spectrometry of intact soluble protein complexes has emerged as a powerful technique to study the stoichiometry, structure-function and dynamics of protein assemblies. Recent developments have extended this technique to the study of membrane protein complexes where it has already revealed subunit stoichiometries and specific phospholipid interactions. Here, we describe a protocol for mass spectrometry of membrane protein complexes. The protocol begins with preparation of the membrane protein complex enabling not only the direct assessment of stoichiometry, delipidation, and quality of the target complex, but also evaluation of the purification strategy. A detailed list of compatible non-ionic detergents is included, along with a protocol for screening detergents to find an optimal one for mass spectrometry, biochemical and structural studies. This protocol also covers the preparation of lipids for protein-lipid binding studies and includes detailed settings for a Q-ToF mass spectrometer after introduction of complexes from gold-coated nanoflow capillaries. PMID:23471109

  3. Detergent-Free Membrane Protein Purification.

    PubMed

    Rothnie, Alice J

    2016-01-01

    Membrane proteins are localized within a lipid bilayer; in order to purify them for functional and structural studies the first step must involve solubilizing or extracting the protein from these lipids. To date this has been achieved using detergents which disrupt the bilayer and bind to the protein in the transmembrane region. However finding conditions for optimal extraction, without destabilizing protein structure, is time consuming and expensive. Here we present a recently-developed method using a styrene-maleic acid (SMA) co-polymer instead of detergents. The SMA co-polymer extracts membrane proteins in a small disc of lipid bilayer which can be used for affinity chromatography purification, thus enabling the purification of membrane proteins while maintaining their native lipid bilayer environment. PMID:27485341

  4. Protein-Centric N-Glycoproteomics Analysis of Membrane and Plasma Membrane Proteins

    PubMed Central

    2015-01-01

    The advent of proteomics technology has transformed our understanding of biological membranes. The challenges for studying membrane proteins have inspired the development of many analytical and bioanalytical tools, and the techniques of glycoproteomics have emerged as an effective means to enrich and characterize membrane and plasma-membrane proteomes. This Review summarizes the development of various glycoproteomics techniques to overcome the hurdles formed by the unique structures and behaviors of membrane proteins with a focus on N-glycoproteomics. Example contributions of N-glycoproteomics to the understanding of membrane biology are provided, and the areas that require future technical breakthroughs are discussed. PMID:24754784

  5. Dielectrophoretic sorting of membrane protein nanocrystals.

    PubMed

    Abdallah, Bahige G; Chao, Tzu-Chiao; Kupitz, Christopher; Fromme, Petra; Ros, Alexandra

    2013-10-22

    Structure elucidation of large membrane protein complexes is still a considerable challenge, yet is a key factor in drug development and disease combat. Femtosecond nanocrystallography is an emerging technique with which structural information of membrane proteins is obtained without the need to grow large crystals, thus overcoming the experimental riddle faced in traditional crystallography methods. Here, we demonstrate for the first time a microfluidic device capable of sorting membrane protein crystals based on size using dielectrophoresis. We demonstrate the excellent sorting power of this new approach with numerical simulations of selected submicrometer beads in excellent agreement with experimental observations. Crystals from batch crystallization broths of the huge membrane protein complex photosystem I were sorted without further treatment, resulting in a high degree of monodispersity and crystallinity in the ~100 nm size range. Microfluidic integration, continuous sorting, and nanometer-sized crystal fractions make this method ideal for direct coupling to femtosecond nanocrystallography.

  6. Efficient preparation and analysis of membrane and membrane protein systems.

    PubMed

    Javanainen, Matti; Martinez-Seara, Hector

    2016-10-01

    Molecular dynamics (MD) simulations have become a highly important technique to consider lipid membrane systems, and quite often they provide considerable added value to laboratory experiments. Rapid development of both software and hardware has enabled the increase of time and size scales reachable by MD simulations to match those attainable by several accurate experimental techniques. However, until recently, the quality and maturity of software tools available for building membrane models for simulations as well as analyzing the results of these simulations have seriously lagged behind. Here, we discuss the recent developments of such tools from the end-users' point of view. In particular, we review the software that can be employed to build lipid bilayers and other related structures with or without embedded membrane proteins to be employed in MD simulations. Additionally, we provide a brief critical insight into force fields and MD packages commonly used for membrane and membrane protein simulations. Finally, we list analysis tools that can be used to study the properties of membrane and membrane protein systems. In all these points we comment on the respective compatibility of the covered tools. We also share our opinion on the current state of the available software. We briefly discuss the most commonly employed tools and platforms on which new software can be built. We conclude the review by providing a few ideas and guidelines on how the development of tools can be further boosted to catch up with the rapid pace at which the field of membrane simulation progresses. This includes improving the compatibility between software tools and promoting the openness of the codes on which these applications rely. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Róg. PMID:26947184

  7. Structure of a photosystem II supercomplex isolated from Prochloron didemni retaining its chlorophyll a/b light-harvesting system

    PubMed Central

    Bibby, Thomas S.; Nield, Jon; Chen, Min; Larkum, Anthony W. D.; Barber, James

    2003-01-01

    Prochlorophytes are a class of cyanobacteria that do not use phycobiliproteins as light-harvesting systems, but contain chlorophyll (Chl) a/b-binding Pcb proteins. Recently it was shown that Pcb proteins form an 18-subunit light-harvesting antenna ring around the photosystem I (PSI) trimeric reaction center complex of the prochlorophyte Prochlorococcus marinus SS120. Here we have investigated whether the symbiotic prochlorophyte Prochloron didemni also contains the same supermolecular complex. Using cells isolated directly from its ascidian host, we found no evidence for the presence of the Pcb–PSI supercomplex. Instead we have identified and characterized a supercomplex composed of photosystem II (PSII) and Pcb proteins. We show that 10-Pcb subunits associate with the PSII dimeric reaction center core to form a giant complex having an estimated Mr of 1,500 kDa with dimensions of 210 × 290 Å. Five-Pcb subunits flank each long side of the dimer and assuming each binds 13 Chl molecules, increase the antenna size of PSII by ≈200%. Fluorescence emission studies indicate that energy transfer occurs efficiently from the Pcb antenna. Modeling using the x-ray structure of cyanobacterial PSII suggests that energy transfer to the PSII reaction center is via the Chls bound to the CP47 and CP43 proteins. PMID:12837938

  8. Membrane protein structure determination - the next generation.

    PubMed

    Moraes, Isabel; Evans, Gwyndaf; Sanchez-Weatherby, Juan; Newstead, Simon; Stewart, Patrick D Shaw

    2014-01-01

    The field of Membrane Protein Structural Biology has grown significantly since its first landmark in 1985 with the first three-dimensional atomic resolution structure of a membrane protein. Nearly twenty-six years later, the crystal structure of the beta2 adrenergic receptor in complex with G protein has contributed to another landmark in the field leading to the 2012 Nobel Prize in Chemistry. At present, more than 350 unique membrane protein structures solved by X-ray crystallography (http://blanco.biomol.uci.edu/mpstruc/exp/list, Stephen White Lab at UC Irvine) are available in the Protein Data Bank. The advent of genomics and proteomics initiatives combined with high-throughput technologies, such as automation, miniaturization, integration and third-generation synchrotrons, has enhanced membrane protein structure determination rate. X-ray crystallography is still the only method capable of providing detailed information on how ligands, cofactors, and ions interact with proteins, and is therefore a powerful tool in biochemistry and drug discovery. Yet the growth of membrane protein crystals suitable for X-ray diffraction studies amazingly remains a fine art and a major bottleneck in the field. It is often necessary to apply as many innovative approaches as possible. In this review we draw attention to the latest methods and strategies for the production of suitable crystals for membrane protein structure determination. In addition we also highlight the impact that third-generation synchrotron radiation has made in the field, summarizing the latest strategies used at synchrotron beamlines for screening and data collection from such demanding crystals. This article is part of a Special Issue entitled: Structural and biophysical characterisation of membrane protein-ligand binding.

  9. NMR of Membrane Proteins: Beyond Crystals.

    PubMed

    Rajesh, Sundaresan; Overduin, Michael; Bonev, Boyan B

    2016-01-01

    Membrane proteins are essential for the flow of signals, nutrients and energy between cells and between compartments of the cell. Their mechanisms can only be fully understood once the precise structures, dynamics and interactions involved are defined at atomic resolution. Through advances in solution and solid state NMR spectroscopy, this information is now available, as demonstrated by recent studies of stable peripheral and transmembrane proteins. Here we highlight recent cases of G-protein coupled receptors, outer membrane proteins, such as VDAC, phosphoinositide sensors, such as the FAPP-1 pleckstrin homology domain, and enzymes including the metalloproteinase MMP-12. The studies highlighted have resulted in the determination of the 3D structures, dynamical properties and interaction surfaces for membrane-associated proteins using advanced isotope labelling strategies, solubilisation systems and NMR experiments designed for very high field magnets. Solid state NMR offers further insights into the structure and multimeric assembly of membrane proteins in lipid bilayers, as well as into interactions with ligands and targets. Remaining challenges for wider application of NMR to membrane structural biology include the need for overexpression and purification systems for the production of isotope-labelled proteins with fragile folds, and the availability of only a few expensive perdeuterated detergents.Step changes that may transform the field include polymers, such as styrene maleic acid, which obviate the need for detergent altogether, and allow direct high yield purification from cells or membranes. Broader demand for NMR may be facilitated by MODA software, which instantly predicts membrane interactive residues that can subsequently be validated by NMR. In addition, recent developments in dynamic nuclear polarization NMR instrumentation offer a remarkable sensitivity enhancement from low molarity samples and cell surfaces. These advances illustrate the current

  10. NMR of Membrane Proteins: Beyond Crystals.

    PubMed

    Rajesh, Sundaresan; Overduin, Michael; Bonev, Boyan B

    2016-01-01

    Membrane proteins are essential for the flow of signals, nutrients and energy between cells and between compartments of the cell. Their mechanisms can only be fully understood once the precise structures, dynamics and interactions involved are defined at atomic resolution. Through advances in solution and solid state NMR spectroscopy, this information is now available, as demonstrated by recent studies of stable peripheral and transmembrane proteins. Here we highlight recent cases of G-protein coupled receptors, outer membrane proteins, such as VDAC, phosphoinositide sensors, such as the FAPP-1 pleckstrin homology domain, and enzymes including the metalloproteinase MMP-12. The studies highlighted have resulted in the determination of the 3D structures, dynamical properties and interaction surfaces for membrane-associated proteins using advanced isotope labelling strategies, solubilisation systems and NMR experiments designed for very high field magnets. Solid state NMR offers further insights into the structure and multimeric assembly of membrane proteins in lipid bilayers, as well as into interactions with ligands and targets. Remaining challenges for wider application of NMR to membrane structural biology include the need for overexpression and purification systems for the production of isotope-labelled proteins with fragile folds, and the availability of only a few expensive perdeuterated detergents.Step changes that may transform the field include polymers, such as styrene maleic acid, which obviate the need for detergent altogether, and allow direct high yield purification from cells or membranes. Broader demand for NMR may be facilitated by MODA software, which instantly predicts membrane interactive residues that can subsequently be validated by NMR. In addition, recent developments in dynamic nuclear polarization NMR instrumentation offer a remarkable sensitivity enhancement from low molarity samples and cell surfaces. These advances illustrate the current

  11. Enhancing light-harvesting power with coherent vibrational interactions: A quantum heat engine picture

    SciTech Connect

    Killoran, N.; Huelga, S. F.; Plenio, M. B.

    2015-10-21

    Recent evidence suggests that quantum effects may have functional importance in biological light-harvesting systems. Along with delocalized electronic excitations, it is now suspected that quantum coherent interactions with certain near-resonant vibrations may contribute to light-harvesting performance. However, the actual quantum advantage offered by such coherent vibrational interactions has not yet been established. We investigate a quantum design principle, whereby coherent exchange of single energy quanta between electronic and vibrational degrees of freedom can enhance a light-harvesting system’s power above what is possible by thermal mechanisms alone. We present a prototype quantum heat engine which cleanly illustrates this quantum design principle and quantifies its quantum advantage using thermodynamic measures of performance. We also demonstrate the principle’s relevance in parameter regimes connected to natural light-harvesting structures.

  12. Expression, purification, and crystallisationof membrane proteins

    NASA Astrophysics Data System (ADS)

    Byrne, Bernadette

    Approximately, 29,000 protein structures are deposited in the Protein Databank (January 2005), but only about 90 of which are independent membrane protein structures. This represents a significant increase in knowledge compared with a matter of only 5 years ago when a mere handful of membrane protein structures were available. Despite the advances, our understanding of the structure-function relationships and mechanism of action of many membrane proteins is still lacking. This is particularly true of many of the more clinically relevant membrane proteins, such as the G-protein-coupled receptors (GPCRs). The GPCRs regulate cellular responses to a wide range of biologically active molecules including hormones and drugs and are thus important targets for therapeutic intervention in a number of disease states. However, the increasing number of membrane protein structures has provided a critical mass of information which has yielded a more rational approach to the process of obtaining diffraction quality crystals. It is the different stages of this process; expression, solubilisation, purification, and crystallisation that will be covered in this lecture.

  13. Tetrastyryl-BODIPY-based dendritic light harvester and estimation of energy transfer efficiency.

    PubMed

    Kostereli, Ziya; Ozdemir, Tugba; Buyukcakir, Onur; Akkaya, Engin U

    2012-07-20

    Versatile BODIPY dyes can be transformed into bright near-IR-emitting fluorophores by quadruple styryl substitutions. When clickable functionalities on the styryl moieties are inserted, an efficient synthesis of a light harvester is possible. In addition, clear spectral evidence is presented showing that, in dendritic light harvesters, calculations commonly based on quantum yield or emission lifetime changes of the donor are bound to yield large overestimations of energy transfer efficiency.

  14. Supermolecular organization of photosystem II and its associated light-harvesting antenna in Arabidopsis thaliana.

    PubMed

    Yakushevska, A E; Jensen, P E; Keegstra, W; van Roon, H; Scheller, H V; Boekema, E J; Dekker, J P

    2001-12-01

    The organization of Arabidopsis thaliana photosystem II (PSII) and its associated light-harvesting antenna (LHCII) was studied in isolated PSII-LHCII supercomplexes and native membrane-bound crystals by transmission electron microscopy and image analysis. Over 4000 single-particle projections of PSII-LHCII supercomplexes were analyzed. In comparison to spinach supercomplexes [Boekema, E.J., van Roon, H., van Breemen, J.F.L. & Dekker, J.P. (1999) Eur. J. Biochem. 266, 444-452] some striking differences were revealed: a much larger number of supercomplexes from Arabidopsis contain copies of M-type LHCII trimers. M-type trimers can also bind in the absence of the more common S-type trimers. No binding of l-type trimers could be detected. Analysis of native membrane-bound PSII crystals revealed a novel type of crystal with a unit cell of 25.6 x 21.4 nm (angle 77 degrees ), which is larger than any of the PSII lattices observed before. The data show that the unit cell is built up from C2S2M2 supercomplexes, rather than from C2S2M supercomplexes observed in native membrane crystals from spinach [Boekema, E.J., Van Breemen, J.F.L., Van Roon, H. & Dekker, J.P. (2000) J. Mol. Biol. 301, 1123-1133]. It is concluded from both the single particle analysis and the crystal analysis that the M-type trimers bind more strongly to PSII core complexes in Arabidopsis than in spinach.

  15. Overexpression of membrane proteins using Pichia pastoris.

    PubMed

    Bornert, Olivier; Alkhalfioui, Fatima; Logez, Christel; Wagner, Renaud

    2012-02-01

    Among the small number of expression systems validated for the mass production of eukaryotic membrane proteins (EMPs), the methylotrophic yeast Pichia pastoris stands as one of the most efficient hosts. This system has been used to produce crystallization-grade proteins for a variety of EMPs, from which high-resolution 3D structures have been determined. This unit describes a set of guidelines and instructions to overexpress membrane proteins using the P. pastoris system. Using a G protein-coupled receptor (GPCR) as a model EMP, these protocols illustrate the necessary steps, starting with the design of the DNA sequence to be expressed, through the preparation and analysis of samples containing the corresponding membrane protein of interest. In addition, recommendations are given on a series of experimental parameters that can be optimized to substantially improve the amount and/or the functionality of the expressed EMPs.

  16. Differential Synthesis of Photosystem Cores and Light-Harvesting Antenna during Proplastid to Chloroplast Development in Spirodela oligorrhiza.

    PubMed

    McCormac, D J; Greenberg, B M

    1992-03-01

    Proplastids and etioplasts are common starting points for monitoring chloroplast development in higher plants. Although proplastids are the primary precursor of chloroplasts, most proplastid to chloroplast systems are cumbersome to study temporally. Conversely, the etioplast to chloroplast transition is initiated by light and is readily examined as a function of time. Etioplasts, however, are found mostly in plants germinated in the dark and are not an obligatory step in chloroplast development. We have chosen to study chloroplast ontogeny in Spirodela oligorrhiza (Kurtz) Hegelm (a C(3)-monocot) because of its unique ability to grow indefinitely in the dark. Ultrastructural, physiological, and molecular evidence is presented in support of a temporal, light-triggered proplastid to chloroplast transition in Spirodela. The dark-grown plants are devoid of chlorophyll, and upon illumination synchronously green over a 3- to 5-day period. Synthesis of chloroplast proteins involved in photosynthesis is coincident with thylakoid assembly, chlorophyll accumulation, and appearance of CO(2) fixation activity. Interestingly, the developmental sequence in Spirodela was slow enough to reveal that biosynthesis of the D1 photosystem II reaction center protein precedes biosynthesis of the major light-harvesting antenna proteins. This, coupled with the high chlorophyll a/b ratio observed early in development, indicated that reaction center assembly occurred prior to accumulation of the light-harvesting complexes. Thus, with Spirodela one can study proplastid to chloroplast conversions temporally in higher plants and follow the process on a time scale that enables a detailed dissection of plastid maturation processes.

  17. Electronic Structure and Dynamics of Higher-Lying Excited States in Light Harvesting Complex 1 from Rhodobacter sphaeroides.

    PubMed

    Dahlberg, Peter D; Ting, Po-Chieh; Massey, Sara C; Martin, Elizabeth C; Hunter, C Neil; Engel, Gregory S

    2016-06-23

    Light harvesting in photosynthetic organisms involves efficient transfer of energy from peripheral antenna complexes to core antenna complexes, and ultimately to the reaction center where charge separation drives downstream photosynthetic processes. Antenna complexes contain many strongly coupled chromophores, which complicates analysis of their electronic structure. Two-dimensional electronic spectroscopy (2DES) provides information on energetic coupling and ultrafast energy transfer dynamics, making the technique well suited for the study of photosynthetic antennae. Here, we present 2DES results on excited state properties and dynamics of a core antenna complex, light harvesting complex 1 (LH1), embedded in the photosynthetic membrane of Rhodobacter sphaeroides. The experiment reveals weakly allowed higher-lying excited states in LH1 at 770 nm, which transfer energy to the strongly allowed states at 875 nm with a lifetime of 40 fs. The presence of higher-lying excited states is in agreement with effective Hamiltonians constructed using parameters from crystal structures and atomic force microscopy (AFM) studies. The energy transfer dynamics between the higher- and lower-lying excited states agree with Redfield theory calculations.

  18. Insight into the Structure of Light Harvesting Complex II and its Stabilization in Detergent Solution

    SciTech Connect

    Cardoso, Mateus B; Smolensky, Dmitriy; Heller, William T; O'Neill, Hugh Michael

    2009-01-01

    The structure of spinach light-harvesting complex II (LHC II), stabilized in a solution of the detergent n-octyl-{beta}-d-glucoside (BOG), was investigated by small-angle neutron scattering (SANS). Physicochemical characterization of the isolated complex indicated that it was pure (>95%) and also in its native trimeric state. SANS with contrast variation was used to investigate the properties of the protein-detergent complex at three different H{sub 2}O/D{sub 2}O contrast match points, enabling the scattering properties of the protein and detergent to be investigated independently. The topological shape of LHC II, determined using ab initio shape restoration methods from the SANS data at the contrast match point of BOG, was consistent with the X-ray crystallographic structure of LHC II (Liu et al. Nature 2004 428, 287-292). The interactions of the protein and detergent were investigated at the contrast match point for the protein and also in 100% D{sub 2}O. The data suggested that BOG micelle structure was altered by its interaction with LHC II, but large aggregate structures were not formed. Indirect Fourier transform analysis of the LHC II/BOG scattering curves showed that the increase in the maximum dimension of the protein-detergent complex was consistent with the presence of a monolayer of detergent surrounding the protein. A model of the LHC II/BOG complex was generated to interpret the measurements made in 100% D{sub 2}O. This model adequately reproduced the overall size of the LHC II/BOG complex, but demonstrated that the detergent does not have a highly regular shape that surrounds the hydrophobic periphery of LHC II. In addition to demonstrating that natively structured LHC II can be produced for functional characterization and for use in artificial solar energy applications, the analysis and modeling approaches described here can be used for characterizing detergent-associated {alpha}-helical transmembrane proteins.

  19. Protein Stains to Detect Antigen on Membranes.

    PubMed

    Dsouza, Anil; Scofield, R Hal

    2015-01-01

    Western blotting (protein blotting/electroblotting) is the gold standard in the analysis of complex protein mixtures. Electroblotting drives protein molecules from a polyacrylamide (or less commonly, of an agarose) gel to the surface of a binding membrane, thereby facilitating an increased availability of the sites with affinity for both general and specific protein reagents. The analysis of these complex protein mixtures is achieved by the detection of specific protein bands on a membrane, which in turn is made possible by the visualization of protein bands either by chemical staining or by reaction with an antibody of a conjugated ligand. Chemical methods employ staining with organic dyes, metal chelates, autoradiography, fluorescent dyes, complexing with silver, or prelabeling with fluorophores. All of these methods have differing sensitivities and quantitative determinations vary significantly. This review will describe the various protein staining methods applied to membranes after western blotting. "Detection" precedes and is a prerequisite to obtaining qualitative and quantitative data on the proteins in a sample, as much as to comparing the protein composition of different samples. "Detection" is often synonymous to staining, i.e., the reversible or irreversible binding by the proteins of a colored organic or inorganic chemical.

  20. Protein stains to detect antigen on membranes.

    PubMed

    D'souza, Anil; Scofield, R Hal

    2009-01-01

    Western blotting (protein blotting/electroblotting) is the gold standard in the analysis of complex protein mixtures. Electroblotting drives protein molecules from a polyacrylamide (or less commonly, of an agarose) gel to the surface of a binding membrane, thereby facilitating an increased availability of the sites with affinity for both general and specific protein reagents. The analysis of these complex protein mixtures is achieved by the detection of specific protein bands on a membrane, which in turn is made possible by the visualization of protein bands either by chemical staining or by reaction with an antibody of a conjugated ligand. Chemical methods employ staining with organic dyes, metal chelates, autoradiography, fluorescent dyes, complexing with silver, or prelabeling with fluorophores. All of these methods have differing sensitivities and quantitative determinations vary significantly. This review will describe the various protein staining methods applied to membranes after electrophoresis. "Detection" precedes and is a prerequisite to obtaining qualitative and quantitative data on the proteins in a sample, as much as to comparing the protein composition of different samples. Detection is often synonymous to staining, i.e., the reversible or irreversible binding by the proteins of a colored organic or inorganic chemical. PMID:19378080

  1. Selective repression of light harvesting complex 2 formation in Rhodobacter azotoformans by light under semiaerobic conditions.

    PubMed

    Yue, Huiying; Zhao, Chungui; Li, Kai; Yang, Suping

    2015-11-01

    Photosystem formation in anaerobic anoxygenic phototrophic bacteria (APB) is repressed by oxygen but is de-repressed when oxygen tension decreases. Under semiaerobic conditions, the synthesis of photopigments and pigment protein complexes in Rhodobacter (Rba.) sphaeroides are repressed by light. AppA, a blue-light receptor, mediates this regulation. In the present study, it was showed that the synthesis of bacteriochlorophyll, carotenoid, and pigment protein complexes in Rba. azotoformans 134K20 was significantly repressed by oxygen. Oxygen exposure also led to a conversion of spheroidene to spheroidenone. In semiaerobically growing cells, light irradiation resulted in a decrease in the formation of photosystem, and blue light was found to be the most effective light source. Blue light reduced the contents of bacteriochlorophyll and carotenoid slightly, but had negligible effects on light harvesting complex (LH) 1 content, whereas the content of LH2 was significantly decreased indicating that blue light selectively repressed the synthesis of LH2 in semiaerobically growing 134K20. It was concluded that, similar to Rba. sphaeroides, a blue light receptor presented in strain 134K20 played important roles in its light-dependent repression. A possible mechanism involved in controlling the differential inhibitory of blue light on the synthesis of photosystem was discussed. PMID:26193456

  2. Spectroscopic elucidation of uncoupled transition energies in the major photosynthetic light-harvesting complex, LHCII

    PubMed Central

    Schlau-Cohen, Gabriela S.; Calhoun, Tessa R.; Ginsberg, Naomi S.; Ballottari, Matteo; Bassi, Roberto; Fleming, Graham R.

    2010-01-01

    Electrostatic couplings between chromophores in photosynthetic pigment–protein complexes, and interactions of pigments with the surrounding protein environment, produce a complicated energy landscape of delocalized excited states. The resultant electronic structure absorbs light and gives rise to energy transfer steps that direct the excitation toward a site of charge separation with near unity quantum efficiency. Knowledge of the transition energies of the uncoupled chromophores is required to describe how the wave functions of the individual pigments combine to form this manifold of delocalized excited states that effectively harvests light energy. In an investigation of the major light-harvesting complex of photosystem II (LHCII), we develop a method based on polarized 2D electronic spectroscopy to experimentally access the energies of the S0–S1 transitions in the chromophore site basis. Rotating the linear polarization of the incident laser pulses reveals previously hidden off-diagonal features. We exploit the polarization dependence of energy transfer peaks to find the angles between the excited state transition dipole moments. We show that these angles provide a spectroscopic method to directly inform on the relationship between the delocalized excitons and the individual chlorophylls through the site energies of the uncoupled chromophores. PMID:20622154

  3. Biophysical Characterization of Membrane Proteins in Nanodiscs

    PubMed Central

    Inagaki, Sayaka; Ghirlando, Rodolfo; Grisshammer, Reinhard

    2012-01-01

    Nanodiscs are self-assembled discoidal phospholipid bilayers surrounded and stabilized by membrane scaffold proteins (MSP), that have become a powerful and promising tool for the study of membrane proteins. Even though their reconstitution is highly regulated by the type of MSP and phospholipid input, a biophysical characterization leading to the determination of the stoichiometry of MSP, lipid and membrane protein is essential. This is important for biological studies, as the oligomeric state of membrane proteins often correlates with their functional activity. Typically combinations of several methods are applied using, for example, modified samples that incorporate fluorescent labels, along with procedures that result in nanodisc disassembly and lipid dissolution. To obtain a comprehensive understanding of the native properties of nanodiscs, modification-free analysis methods are required. In this work we provide a strategy, using a combination of dynamic light scattering and analytical ultracentrifugation, for the biophysical characterization of unmodified nanodiscs. In this manner we characterize the nanodisc preparation in terms of its overall polydispersity and characterize the hydrodynamically resolved nanodisc of interest in terms of its sedimentation coefficient, Stokes’ radius and overall protein and lipid stoichiometry. Functional and biological applications are also discussed for the study of the membrane protein embedded in nanodiscs under defined experimental conditions. PMID:23219517

  4. Intrinsically disordered proteins drive membrane curvature

    NASA Astrophysics Data System (ADS)

    Busch, David J.; Houser, Justin R.; Hayden, Carl C.; Sherman, Michael B.; Lafer, Eileen M.; Stachowiak, Jeanne C.

    2015-07-01

    Assembly of highly curved membrane structures is essential to cellular physiology. The prevailing view has been that proteins with curvature-promoting structural motifs, such as wedge-like amphipathic helices and crescent-shaped BAR domains, are required for bending membranes. Here we report that intrinsically disordered domains of the endocytic adaptor proteins, Epsin1 and AP180 are highly potent drivers of membrane curvature. This result is unexpected since intrinsically disordered domains lack a well-defined three-dimensional structure. However, in vitro measurements of membrane curvature and protein diffusivity demonstrate that the large hydrodynamic radii of these domains generate steric pressure that drives membrane bending. When disordered adaptor domains are expressed as transmembrane cargo in mammalian cells, they are excluded from clathrin-coated pits. We propose that a balance of steric pressure on the two surfaces of the membrane drives this exclusion. These results provide quantitative evidence for the influence of steric pressure on the content and assembly of curved cellular membrane structures.

  5. Visualizing structural dynamics of thylakoid membranes

    PubMed Central

    Iwai, Masakazu; Yokono, Makio; Nakano, Akihiko

    2014-01-01

    To optimize photosynthesis, light-harvesting antenna proteins regulate light energy dissipation and redistribution in chloroplast thylakoid membranes, which involve dynamic protein reorganization of photosystems I and II. However, direct evidence for such protein reorganization has not been visualized in live cells. Here we demonstrate structural dynamics of thylakoid membranes by live cell imaging in combination with deconvolution. We observed chlorophyll fluorescence in the antibiotics-induced macrochloroplast in the moss Physcomitrella patens. The three-dimensional reconstruction uncovered the fine thylakoid membrane structure in live cells. The time-lapse imaging shows that the entire thylakoid membrane network is structurally stable, but the individual thylakoid membrane structure is flexible in vivo. Our observation indicates that grana serve as a framework to maintain structural integrity of the entire thylakoid membrane network. Both the structural stability and flexibility of thylakoid membranes would be essential for dynamic protein reorganization under fluctuating light environments. PMID:24442007

  6. Visualizing structural dynamics of thylakoid membranes.

    PubMed

    Iwai, Masakazu; Yokono, Makio; Nakano, Akihiko

    2014-01-20

    To optimize photosynthesis, light-harvesting antenna proteins regulate light energy dissipation and redistribution in chloroplast thylakoid membranes, which involve dynamic protein reorganization of photosystems I and II. However, direct evidence for such protein reorganization has not been visualized in live cells. Here we demonstrate structural dynamics of thylakoid membranes by live cell imaging in combination with deconvolution. We observed chlorophyll fluorescence in the antibiotics-induced macrochloroplast in the moss Physcomitrella patens. The three-dimensional reconstruction uncovered the fine thylakoid membrane structure in live cells. The time-lapse imaging shows that the entire thylakoid membrane network is structurally stable, but the individual thylakoid membrane structure is flexible in vivo. Our observation indicates that grana serve as a framework to maintain structural integrity of the entire thylakoid membrane network. Both the structural stability and flexibility of thylakoid membranes would be essential for dynamic protein reorganization under fluctuating light environments.

  7. Crystallization of Membrane Proteins by Vapor Diffusion

    PubMed Central

    Delmar, Jared A.; Bolla, Jani Reddy; Su, Chih-Chia; Yu, Edward W.

    2016-01-01

    X-ray crystallography remains the most robust method to determine protein structure at the atomic level. However, the bottlenecks of protein expression and purification often discourage further study. In this chapter, we address the most common problems encountered at these stages. Based on our experiences in expressing and purifying antimicrobial efflux proteins, we explain how a pure and homogenous protein sample can be successfully crystallized by the vapor diffusion method. We present our current protocols and methodologies for this technique. Case studies show step-by-step how we have overcome problems related to expression and diffraction, eventually producing high quality membrane protein crystals for structural determinations. It is our hope that a rational approach can be made of the often anecdotal process of membrane protein crystallization. PMID:25950974

  8. Protein separation using an electrically tunable membrane

    NASA Astrophysics Data System (ADS)

    Jou, Ining; Melnikov, Dmitriy; Gracheva, Maria

    Separation of small proteins by charge with a solid-state porous membrane requires control over the protein's movement. Semiconductor membrane has this ability due to the electrically tunable electric potential profile inside the nanopore. In this work we investigate the possibility to separate the solution of two similar sized proteins by charge. As an example, we consider two small globular proteins abundant in humans: insulin (negatively charged) and ubiquitin (neutral). We find that the localized electric field inside the pore either attracts or repels the charged protein to or from the pore wall which affects the delay time before a successful translocation of the protein through the nanopore. However, the motion of the uncharged ubiquitin is unaffected. The difference in the delay time (and hence the separation) can be further increased by the application of the electrolyte bias which induces an electroosmotic flow in the pore. NSF DMR and CBET Grant No. 1352218.

  9. Weakly Stable Regions and Protein-Protein Interactions in Beta-Barrel Membrane Proteins

    PubMed Central

    Naveed, Hammad; Liang, Jie

    2014-01-01

    We briefly discuss recent progress in computational characterization of the sequence and structural properties of β-barrel membrane properties. We discuss the emerging concept of weakly stable regions in β-barrel membrane proteins, computational methods to identify these regions and mechanisms adopted by β-barrel membrane proteins in nature to stabilize them. We further discuss computational methods to identify protein-protein interactions in β-barrel membrane proteins and recent experimental studies that aim at altering the biophysical properties including oligomerization state and stability of β-barrel membrane proteins based on the emerging organization principles of these proteins from recent computational studies. PMID:23713778

  10. Curvature-mediated interactions between membrane proteins.

    PubMed Central

    Kim, K S; Neu, J; Oster, G

    1998-01-01

    Membrane proteins can deform the lipid bilayer in which they are embedded. If the bilayer is treated as an elastic medium, then these deformations will generate elastic interactions between the proteins. The interaction between a single pair is repulsive. However, for three or more proteins, we show that there are nonpairwise forces whose magnitude is similar to the pairwise forces. When there are five or more proteins, we show that the nonpairwise forces permit the existence of stable protein aggregates, despite their pairwise repulsions. PMID:9788923

  11. Tracing the evolution of the light-harvesting antennae in chlorophyll a/b-containing organisms.

    PubMed

    Koziol, Adam G; Borza, Tudor; Ishida, Ken-Ichiro; Keeling, Patrick; Lee, Robert W; Durnford, Dion G

    2007-04-01

    The light-harvesting complexes (LHCs) of land plants and green algae have essential roles in light capture and photoprotection. Though the functional diversity of the individual LHC proteins are well described in many land plants, the extent of this family in the majority of green algal groups is unknown. To examine the evolution of the chlorophyll a/b antennae system and to infer its ancestral state, we initiated several expressed sequence tag projects from a taxonomically broad range of chlorophyll a/b-containing protists. This included representatives from the Ulvophyceae (Acetabularia acetabulum), the Mesostigmatophyceae (Mesostigma viride), and the Prasinophyceae (Micromonas sp.), as well as one representative from each of the Euglenozoa (Euglena gracilis) and Chlorarachniophyta (Bigelowiella natans), whose plastids evolved secondarily from a green alga. It is clear that the core antenna system was well developed prior to green algal diversification and likely consisted of the CP29 (Lhcb4) and CP26 (Lhcb5) proteins associated with photosystem II plus a photosystem I antenna composed of proteins encoded by at least Lhca3 and two green algal-specific proteins encoded by the Lhca2 and 9 genes. In organisms containing secondary plastids, we found no evidence for orthologs to the plant/algal antennae with the exception of CP29. We also identified PsbS homologs in the Ulvophyceae and the Prasinophyceae, indicating that this distinctive protein appeared prior to green algal diversification. This analysis provides a snapshot of the antenna systems in diverse green algae, and allows us to infer the changing complexity of the antenna system during green algal evolution.

  12. Identifying the hub proteins from complicated membrane protein network systems.

    PubMed

    Shen, Yi-Zhen; Ding, Yong-Sheng; Gu, Quan; Chou, Kuo-Chen

    2010-05-01

    The so-called "hub proteins" are those proteins in a protein-protein interaction network system that have remarkably higher interaction relations (or degrees) than the others. Therefore, the information of hub proteins can provide very useful insights for selecting or prioritizing targets during drug development. In this paper, by combining the multi-agent-based method with the graphical spectrum analysis and immune-genetic algorithm, a novel simulator for identifying the hub proteins from membrane protein interaction networks is proposed. As a demonstration of using the simulator, two hub membrane proteins, YPL227C and YIL147C, were identified from a complicated network system consisting of 1500 membrane proteins. Meanwhile, along with the two identified hub proteins, their molecular functions, biological processes, and cellular components were also revealed. It is anticipated that the hub-protein-simulator may become a very useful tool for system biology and drug development, particularly in deciphering unknown protein functions, determining protein complexes, and in identifying the key targets from a complicated disease system. PMID:20507268

  13. Rotavirus Capsid Protein VP5* Permeabilizes Membranes

    PubMed Central

    Denisova, Evgeniya; Dowling, William; LaMonica, Rachel; Shaw, Robert; Scarlata, Suzanne; Ruggeri, Franco; Mackow, Erich R.

    1999-01-01

    Proteolytic cleavage of the VP4 outer capsid spike protein into VP8* and VP5* proteins is required for rotavirus infectivity and for rotavirus-induced membrane permeability. In this study we addressed the function of the VP5* cleavage fragment in permeabilizing membranes. Expressed VP5* and truncated VP5* proteins were purified by nickel affinity chromatography and assayed for their ability to permeabilize large unilamellar vesicles (LUVs) preloaded with carboxyfluorescein (CF). VP5* and VP5* truncations, but not VP4 or VP8*, permeabilized LUVs as measured by fluorescence dequenching of released CF. Similar to virus-induced CF release, VP5*-induced CF release was concentration and temperature dependent, with a pH optimum of 7.35 at 37°C, but independent of the presence of divalent cations or cholesterol. VP5*-induced permeability was completely inhibited by VP5*-specific neutralizing monoclonal antibodies (2G4, M2, or M7) which recognize conformational epitopes on VP5* but was not inhibited by VP8*-specific neutralizing antibodies. In addition, N-terminal and C-terminal VP5* truncations including residues 265 to 474 are capable of permeabilizing LUVs. These findings demonstrate that VP5* permeabilizes membranes in the absence of other rotavirus proteins and that membrane-permeabilizing VP5* truncations contain the putative fusion region within predicted virion surface domains. The ability of recombinant expressed VP5* to permeabilize membranes should permit us to functionally define requirements for VP5*-membrane interactions. These findings indicate that VP5* is a specific membrane-permeabilizing capsid protein which is likely to play a role in the cellular entry of rotaviruses. PMID:10074166

  14. Ultrafast spectroscopy of trimeric light-harvesting complex II from higher plants

    SciTech Connect

    Connelly, J.P.; Mueller, G.M.; Hucke, M.; Gatzen, G.; Holzwarth, A.R.; Mullineaux, C.W.; Ruban, A.V.; Horton, P.

    1997-03-06

    Time-resolved femtosecond transient absorption measurements have been carried out at room temperature on light-harvesting chlorophyll a/b protein complex of photosystem II (LHC II) trimers prepared from spinach. Exciting in the chlorophyll (Chl) b region at 650 nm with very low intensity, virtually annihilation-free two-color transient absorption measurement of the kinetics over 100 ps, between 645 and 690 nm, yield global lifetimes of 175 fs, 625 fs, and 5 ps and a long component (>=790 ps) where the three fastest lifetimes reflect Chl b to Chl a energy transfer. On the basis of these results and recent electron diffraction structural data, a preliminary three-pool Ch a, three-pool Chl b kinetic model is proposed. The possible influence of variable xanthophyll composition on quenching in LHC II preparations isolated from light- and dark-adapted leaves has been investigated using time-resolved picosecond fluorescence at room temperature. Global lifetimes of 5 ps and 3.6 ns, the lifetimes of the terminal LHC II excited state, were obtained. No discernable quenching effect due to the presence of zeaxanthin was observed. 38 refs., 6 figs., 3 tabs.

  15. Efficient estimation of energy transfer efficiency in light-harvesting complexes.

    PubMed

    Shabani, A; Mohseni, M; Rabitz, H; Lloyd, S

    2012-07-01

    The fundamental physical mechanisms of energy transfer in photosynthetic complexes is not yet fully understood. In particular, the degree of efficiency or sensitivity of these systems for energy transfer is not known given their realistic with surrounding photonic and phononic environments. One major problem in studying light-harvesting complexes has been the lack of an efficient method for simulation of their dynamics in biological environments. To this end, here we revisit the second order time-convolution (TC2) master equation and examine its reliability beyond extreme Markovian and perturbative limits. In particular, we present a derivation of TC2 without making the usual weak system-bath coupling assumption. Using this equation, we explore the long-time behavior of exciton dynamics of Fenna-Matthews-Olson (FMO) portein complex. Moreover, we introduce a constructive error analysis to estimate the accuracy of TC2 equation in calculating energy transfer efficiency, exhibiting reliable performance for system-bath interactions with weak and intermediate memory and strength. Furthermore, we numerically show that energy transfer efficiency is optimal and robust for the FMO protein complex of green sulfur bacteria with respect to variations in reorganization energy and bath correlation time scales.

  16. Model-building codes for membrane proteins.

    SciTech Connect

    Shirley, David Noyes; Hunt, Thomas W.; Brown, W. Michael; Schoeniger, Joseph S.; Slepoy, Alexander; Sale, Kenneth L.; Young, Malin M.; Faulon, Jean-Loup Michel; Gray, Genetha Anne

    2005-01-01

    We have developed a novel approach to modeling the transmembrane spanning helical bundles of integral membrane proteins using only a sparse set of distance constraints, such as those derived from MS3-D, dipolar-EPR and FRET experiments. Algorithms have been written for searching the conformational space of membrane protein folds matching the set of distance constraints, which provides initial structures for local conformational searches. Local conformation search is achieved by optimizing these candidates against a custom penalty function that incorporates both measures derived from statistical analysis of solved membrane protein structures and distance constraints obtained from experiments. This results in refined helical bundles to which the interhelical loops and amino acid side-chains are added. Using a set of only 27 distance constraints extracted from the literature, our methods successfully recover the structure of dark-adapted rhodopsin to within 3.2 {angstrom} of the crystal structure.

  17. Transmembrane protein sorting driven by membrane curvature

    PubMed Central

    Strahl, H.; Ronneau, S.; González, B. Solana; Klutsch, D.; Schaffner-Barbero, C.; Hamoen, L. W.

    2015-01-01

    The intricate structure of prokaryotic and eukaryotic cells depends on the ability to target proteins to specific cellular locations. In most cases, we have a poor understanding of the underlying mechanisms. A typical example is the assembly of bacterial chemoreceptors at cell poles. Here we show that the classical chemoreceptor TlpA of Bacillus subtilis does not localize according to the consensus stochastic nucleation mechanism but accumulates at strongly curved membrane areas generated during cell division. This preference was confirmed by accumulation at non-septal curved membranes. Localization appears to be an intrinsic property of the protein complex and does not rely on chemoreceptor clustering, as was previously shown for Escherichia coli. By constructing specific amino-acid substitutions, we demonstrate that the preference for strongly curved membranes arises from the curved shape of chemoreceptor trimer of dimers. These findings demonstrate that the intrinsic shape of transmembrane proteins can determine their cellular localization. PMID:26522943

  18. Transmembrane protein sorting driven by membrane curvature

    NASA Astrophysics Data System (ADS)

    Strahl, H.; Ronneau, S.; González, B. Solana; Klutsch, D.; Schaffner-Barbero, C.; Hamoen, L. W.

    2015-11-01

    The intricate structure of prokaryotic and eukaryotic cells depends on the ability to target proteins to specific cellular locations. In most cases, we have a poor understanding of the underlying mechanisms. A typical example is the assembly of bacterial chemoreceptors at cell poles. Here we show that the classical chemoreceptor TlpA of Bacillus subtilis does not localize according to the consensus stochastic nucleation mechanism but accumulates at strongly curved membrane areas generated during cell division. This preference was confirmed by accumulation at non-septal curved membranes. Localization appears to be an intrinsic property of the protein complex and does not rely on chemoreceptor clustering, as was previously shown for Escherichia coli. By constructing specific amino-acid substitutions, we demonstrate that the preference for strongly curved membranes arises from the curved shape of chemoreceptor trimer of dimers. These findings demonstrate that the intrinsic shape of transmembrane proteins can determine their cellular localization.

  19. Atomic-level analysis of membrane-protein structure.

    PubMed

    Hendrickson, Wayne A

    2016-06-01

    Membrane proteins are substantially more challenging than natively soluble proteins as subjects for structural analysis. Thus, membrane proteins are greatly underrepresented in structural databases. Recently, focused consortium efforts and advances in methodology for protein production, crystallographic analysis and cryo-EM analysis have accelerated the pace of atomic-level structure determination of membrane proteins.

  20. Construction and structural analysis of tethered lipid bilayer containing photosynthetic antenna proteins for functional analysis.

    PubMed

    Sumino, Ayumi; Dewa, Takehisa; Takeuchi, Toshikazu; Sugiura, Ryuta; Sasaki, Nobuaki; Misawa, Nobuo; Tero, Ryugo; Urisu, Tsuneo; Gardiner, Alastair T; Cogdell, Richard J; Hashimoto, Hideki; Nango, Mamoru

    2011-07-11

    The construction and structural analysis of a tethered planar lipid bilayer containing bacterial photosynthetic membrane proteins, light-harvesting complex 2 (LH2), and light-harvesting core complex (LH1-RC) is described and establishes this system as an experimental platform for their functional analysis. The planar lipid bilayer containing LH2 and/or LH1-RC complexes was successfully formed on an avidin-immobilized coverglass via an avidin-biotin linkage. Atomic force microscopy (AFM) showed that a smooth continuous membrane was formed there. Lateral diffusion of these membrane proteins, observed by a fluorescence recovery after photobleaching (FRAP), is discussed in terms of the membrane architecture. Energy transfer from LH2 to LH1-RC within the tethered membrane was observed by steady-state fluorescence spectroscopy, indicating that the tethered membrane can mimic the natural situation.

  1. Major intrinsic proteins in biomimetic membranes.

    PubMed

    Nielsen, Claus Hélix

    2010-01-01

    Biological membranes define the structural and functional boundaries in living cells and their organelles. The integrity of the cell depends on its ability to separate inside from outside and yet at the same time allow massive transport of matter in and out the cell. Nature has elegantly met this challenge by developing membranes in the form of lipid bilayers in which specialized transport proteins are incorporated. This raises the question: is it possible to mimic biological membranes and create a membrane based sensor and/or separation device? In the development of a biomimetic sensor/separation technology, a unique class of membrane transport proteins is especially interesting-the major intrinsic proteins (MIPs). Generally, MIPs conduct water molecules and selected solutes in and out of the cell while preventing the passage of other solutes, a property critical for the conservation of the cells internal pH and salt concentration. Also known as water channels or aquaporins they are highly efficient membrane pore proteins some of which are capable of transporting water at very high rates up to 10(9) molecules per second. Some MIPs transport other small, uncharged solutes, such as glycerol and other permeants such as carbon dioxide, nitric oxide, ammonia, hydrogen peroxide and the metalloids antimonite, arsenite, silicic and boric acid depending on the effective restriction mechanism of the protein. The flux properties of MIPs thus lead to the question ifMIPs can be used in separation devices or as sensor devices based on, e.g., the selective permeation of metalloids. In principle a MIP based membrane sensor/separation device requires the supporting biomimetic matrix to be virtually impermeable to anything but water or the solute in question. In practice, however, a biomimetic support matrix will generally have finite permeabilities to both electrolytes and non-electrolytes. The feasibility of a biomimetic MIP device thus depends on the relative transport

  2. Protein permeation through an electrically tunable membrane

    NASA Astrophysics Data System (ADS)

    Jou, Ining A.; Melnikov, Dmitriy V.; Gracheva, Maria E.

    2016-05-01

    Protein filtration is important in many fields of science and technology such as medicine, biology, chemistry, and engineering. Recently, protein separation and filtering with nanoporous membranes has attracted interest due to the possibility of fast separation and high throughput volume. This, however, requires understanding of the protein’s dynamics inside and in the vicinity of the nanopore. In this work, we utilize a Brownian dynamics approach to study the motion of the model protein insulin in the membrane-electrolyte electrostatic potential. We compare the results of the atomic model of the protein with the results of a coarse-grained and a single-bead model, and find that the coarse-grained representation of protein strikes the best balance between the accuracy of the results and the computational effort required. Contrary to common belief, we find that to adequately describe the protein, a single-bead model cannot be utilized without a significant effort to tabulate the simulation parameters. Similar to results for nanoparticle dynamics, our findings also indicate that the electric field and the electro-osmotic flow due to the applied membrane and electrolyte biases affect the capture and translocation of the biomolecule by either attracting or repelling it to or from the nanopore. Our computational model can also be applied to other types of proteins and separation conditions.

  3. A nanoscale bio-inspired light-harvesting system developed from self-assembled alkyl-functionalized metallochlorin nano-aggregates

    NASA Astrophysics Data System (ADS)

    Ocakoglu, Kasim; Joya, Khurram S.; Harputlu, Ersan; Tarnowska, Anna; Gryko, Daniel T.

    2014-07-01

    Self-assembled supramolecular organization of nano-structured biomimetic light-harvesting modules inside solid-state nano-templates can be exploited to develop excellent light-harvesting materials for artificial photosynthetic devices. We present here a hybrid light-harvesting system mimicking the chlorosomal structures of the natural photosynthetic system using synthetic zinc chlorin units (ZnChl-C6, ZnChl-C12 and ZnChl-C18) that are self-aggregated inside the anodic aluminum oxide (AAO) nano-channel membranes. AAO nano-templates were modified with a TiO2 matrix and functionalized with long hydrophobic chains to facilitate the formation of supramolecular Zn-chlorin aggregates. The transparent Zn-chlorin nano-aggregates inside the alkyl-TiO2 modified AAO nano-channels have a diameter of ~120 nm in a 60 μm length channel. UV-Vis studies and fluorescence emission spectra further confirm the formation of the supramolecular ZnChl aggregates from monomer molecules inside the alkyl-functionalized nano-channels. Our results prove that the novel and unique method can be used to produce efficient and stable light-harvesting assemblies for effective solar energy capture through transparent and stable nano-channel ceramic materials modified with bio-mimetic molecular self-assembled nano-aggregates.Self-assembled supramolecular organization of nano-structured biomimetic light-harvesting modules inside solid-state nano-templates can be exploited to develop excellent light-harvesting materials for artificial photosynthetic devices. We present here a hybrid light-harvesting system mimicking the chlorosomal structures of the natural photosynthetic system using synthetic zinc chlorin units (ZnChl-C6, ZnChl-C12 and ZnChl-C18) that are self-aggregated inside the anodic aluminum oxide (AAO) nano-channel membranes. AAO nano-templates were modified with a TiO2 matrix and functionalized with long hydrophobic chains to facilitate the formation of supramolecular Zn-chlorin aggregates. The

  4. Cellular Mechanisms of Membrane Protein Folding

    PubMed Central

    Skach, William R.

    2010-01-01

    The membrane protein folding problem can be articulated by two central questions. How is protein topology established by selective peptide transport to opposite sides of the cellular membrane? And how are transmembrane segments inserted, integrated and folded within the lipid bilayer? In eukaryotes, this process usually takes place in the endoplasmic reticulum (ER) coincident with protein synthesis, and is facilitated by the translating Ribosome and the Sec61 Translocon Complex (RTC). At its core, the RTC forms a dynamic pathway through which the elongating nascent polypeptide moves as it is delivered into cytosolic, lumenal and lipid compartments. This perspective will focus on emerging evidence that the RTC functions as a protein folding machine that restricts conformational space by establishing transmembrane topology and yet provides a permissive environment that enables nascent transmembrane domains to efficiently progress down their folding energy landscape. PMID:19491932

  5. Converting a marginally hydrophobic soluble protein into a membrane protein.

    PubMed

    Nørholm, Morten H H; Cunningham, Fiona; Deber, Charles M; von Heijne, Gunnar

    2011-03-18

    δ-Helices are marginally hydrophobic α-helical segments in soluble proteins that exhibit certain sequence characteristics of transmembrane (TM) helices [Cunningham, F., Rath, A., Johnson, R. M. & Deber, C. M. (2009). Distinctions between hydrophobic helices in globular proteins and TM segments as factors in protein sorting. J. Biol. Chem., 284, 5395-402]. In order to better understand the difference between δ-helices and TM helices, we have studied the insertion of five TM-like δ-helices into dog pancreas microsomal membranes. Using model constructs in which an isolated δ-helix is engineered into a bona fide membrane protein, we find that, for two δ-helices originating from secreted proteins, at least three single-nucleotide mutations are necessary to obtain efficient membrane insertion, whereas one mutation is sufficient in a δ-helix from the cytosolic protein P450BM-3. We further find that only when the entire upstream region of the mutated δ-helix in the intact cytochrome P450BM-3 is deleted does a small fraction of the truncated protein insert into microsomes. Our results suggest that upstream portions of the polypeptide, as well as embedded charged residues, protect δ-helices in globular proteins from being recognized by the signal recognition particle-Sec61 endoplasmic-reticulum-targeting machinery and that δ-helices in secreted proteins are mutationally more distant from TM helices than δ-helices in cytosolic proteins.

  6. Extension of Light-Harvesting Ability of Photosynthetic Light-Harvesting Complex 2 (LH2) through Ultrafast Energy Transfer from Covalently Attached Artificial Chromophores.

    PubMed

    Yoneda, Yusuke; Noji, Tomoyasu; Katayama, Tetsuro; Mizutani, Naoto; Komori, Daisuke; Nango, Mamoru; Miyasaka, Hiroshi; Itoh, Shigeru; Nagasawa, Yutaka; Dewa, Takehisa

    2015-10-14

    Introducing appropriate artificial components into natural biological systems could enrich the original functionality. To expand the available wavelength range of photosynthetic bacterial light-harvesting complex 2 (LH2 from Rhodopseudomonas acidophila 10050), artificial fluorescent dye (Alexa Fluor 647: A647) was covalently attached to N- and C-terminal Lys residues in LH2 α-polypeptides with a molar ratio of A647/LH2 ≃ 9/1. Fluorescence and transient absorption spectroscopies revealed that intracomplex energy transfer from A647 to intrinsic chromophores of LH2 (B850) occurs in a multiexponential manner, with time constants varying from 440 fs to 23 ps through direct and B800-mediated indirect pathways. Kinetic analyses suggested that B800 chromophores mediate faster energy transfer, and the mechanism was interpretable in terms of Förster theory. This study demonstrates that a simple attachment of external chromophores with a flexible linkage can enhance the light harvesting activity of LH2 without affecting inherent functions of energy transfer, and can achieve energy transfer in the subpicosecond range. Addition of external chromophores, thus, represents a useful methodology for construction of advanced hybrid light-harvesting systems that afford solar energy in the broad spectrum.

  7. Post-expression strategies for structural investigations of membrane proteins.

    PubMed

    Columbus, Linda

    2015-06-01

    Currently, membrane proteins only comprise 1.5% of the protein data bank and, thus, still remain a challenge for structural biologists. Expression, stabilization in membrane mimics (e.g. detergent), heterogeneity (conformational and chemical), and crystallization in the presence of a membrane mimic are four major bottlenecks encountered. In response, several post-expression protein modifications have been utilized to facilitate structure determination of membrane proteins. This review highlights four approaches: limited proteolysis, deglycosylation, cysteine alkylation, and lysine methylation. Combined these approaches have facilitated the structure determination of more than 40 membrane proteins and, therefore, are a useful addition to the membrane protein structural biologist's toolkit.

  8. Towards an ab initio description of the optical spectra of light-harvesting antennae: application to the CP29 complex of photosystem II.

    PubMed

    Jurinovich, Sandro; Viani, Lucas; Prandi, Ingrid G; Renger, Thomas; Mennucci, Benedetta

    2015-06-14

    Light-harvesting pigment-protein complexes (PPC) represent the fundamental units through which the photosynthetic organisms absorb sunlight and funnel the energy to the reaction centre for carrying out the primary energy conversion reactions of photosynthesis. Here we apply a multiscale computational strategy to a specific PPC present in the photosystem II of plants and algae (CP29) to investigate in what detail should the environment effects due to protein and membrane/solvent be included for an accurate description of optical spectra. We find that a refinement of the crystal structure is needed before any meaningful quantum chemical calculations of pigment transition energies can be performed. For this purpose we apply classical molecular dynamics simulations of the PPC within its natural environment and we perform ab initio computations of the exciton Hamiltonian of the complex, including the environment either implicitly by the polarizable continuum model (PCM) or explicitly using the polarizable QM/MM methodology (MMPol). However, PCM essentially leads to an unspecific redshift of all transition energies, and MMPol is able to reveal site-specific changes in the optical properties of the pigments. Based on the latter and the excitonic couplings obtained within a polarizable QM/MM methodology, optical spectra are calculated, which are in good qualitative agreement with experimental data. A weakness of the approach is however found in the overestimation of the fluctuations of the excitonic parameters of the pigments along the MD trajectory. An explanation for such a finding in terms of the limits of the force fields commonly used for protein cofactors is presented and discussed.

  9. Towards an ab initio description of the optical spectra of light-harvesting antennae: application to the CP29 complex of photosystem II.

    PubMed

    Jurinovich, Sandro; Viani, Lucas; Prandi, Ingrid G; Renger, Thomas; Mennucci, Benedetta

    2015-06-14

    Light-harvesting pigment-protein complexes (PPC) represent the fundamental units through which the photosynthetic organisms absorb sunlight and funnel the energy to the reaction centre for carrying out the primary energy conversion reactions of photosynthesis. Here we apply a multiscale computational strategy to a specific PPC present in the photosystem II of plants and algae (CP29) to investigate in what detail should the environment effects due to protein and membrane/solvent be included for an accurate description of optical spectra. We find that a refinement of the crystal structure is needed before any meaningful quantum chemical calculations of pigment transition energies can be performed. For this purpose we apply classical molecular dynamics simulations of the PPC within its natural environment and we perform ab initio computations of the exciton Hamiltonian of the complex, including the environment either implicitly by the polarizable continuum model (PCM) or explicitly using the polarizable QM/MM methodology (MMPol). However, PCM essentially leads to an unspecific redshift of all transition energies, and MMPol is able to reveal site-specific changes in the optical properties of the pigments. Based on the latter and the excitonic couplings obtained within a polarizable QM/MM methodology, optical spectra are calculated, which are in good qualitative agreement with experimental data. A weakness of the approach is however found in the overestimation of the fluctuations of the excitonic parameters of the pigments along the MD trajectory. An explanation for such a finding in terms of the limits of the force fields commonly used for protein cofactors is presented and discussed. PMID:25872495

  10. Elucidation of structure-function relationships in plant major light-harvesting complex (LHC II) by nonlinear spectroscopy.

    PubMed

    Lokstein, Heiko; Betke, Alexander; Krikunova, Maria; Teuchner, Klaus; Voigt, Bernd

    2012-03-01

    Conventional linear and time-resolved spectroscopic techniques are often not appropriate to elucidate specific pigment-pigment interactions in light-harvesting pigment-protein complexes (LHCs). Nonlinear (laser-) spectroscopic techniques, including nonlinear polarization spectroscopy in the frequency domain (NLPF) as well as step-wise (resonant) and simultaneous (non-resonant) two-photon excitation spectroscopies may be advantageous in this regard. Nonlinear spectroscopies have been used to elucidate substructure(s) of very complex spectra, including analyses of strong excitonic couplings between chlorophylls and of interactions between (bacterio)chlorophylls and "optically dark" states of carotenoids in LHCs, including the major antenna complex of higher plants, LHC II. This article shortly reviews our previous study and outlines perspectives regarding the application of selected nonlinear laser-spectroscopic techniques to disentangle structure-function relationships in LHCs and other pigment-protein complexes.

  11. Self-assembled photosynthesis-inspired light harvesting material and solar cells containing the same

    DOEpatents

    Lindsey, Jonathan S.; Chinnasamy, Muthiah; Fan, Dazhong

    2009-12-15

    A solar cell is described that comprises: (a) a semiconductor charge separation material; (b) at least one electrode connected to the charge separation material; and (c) a light-harvesting film on the charge separation material, the light-harvesting film comprising non-covalently coupled, self-assembled units of porphyrinic macrocycles. The porphyrinic macrocycles preferably comprise: (i) an intramolecularly coordinated metal; (ii) a first coordinating substituent; and (iii) a second coordinating substituent opposite the first coordinating substituent. The porphyrinic macrocycles can be assembled by repeating intermolecular coordination complexes of the metal, the first coordinating substituent and the second coordinating substituent.

  12. Membrane Protein Solubilization and Composition of Protein Detergent Complexes.

    PubMed

    Duquesne, Katia; Prima, Valérie; Sturgis, James N

    2016-01-01

    Membrane proteins are typically expressed in heterologous systems with a view to in vitro characterization. A critical step in the preparation of membrane proteins after expression in any system is the solubilization of the protein in aqueous solution, typically using detergents and lipids, to obtain the protein in a form suitable for purification, structural or functional analysis. This process is particularly difficult as the objective is to prepare the protein in an unnatural environment, a protein detergent complex, separating it from its natural lipid partners while causing the minimum destabilization or modification of the structure. Although the process is difficult, and relatively hard to master, an increasing number of membrane proteins have been successfully isolated after expression in a wide variety of systems. In this chapter we give a general protocol for preparing protein detergent complexes that is aimed at guiding the reader through the different critical steps. In the second part of the chapter we illustrate how to analyze the composition of protein detergent complexes; this analysis is important as it has been found that compositional variation often causes irreproducible results. PMID:27485340

  13. Unified analysis of ensemble and single-complex optical spectral data from light-harvesting complex-2 chromoproteins for gaining deeper insight into bacterial photosynthesis

    NASA Astrophysics Data System (ADS)

    Pajusalu, Mihkel; Kunz, Ralf; Rätsep, Margus; Timpmann, Kõu; Köhler, Jürgen; Freiberg, Arvi

    2015-11-01

    Bacterial light-harvesting pigment-protein complexes are very efficient at converting photons into excitons and transferring them to reaction centers, where the energy is stored in a chemical form. Optical properties of the complexes are known to change significantly in time and also vary from one complex to another; therefore, a detailed understanding of the variations on the level of single complexes and how they accumulate into effects that can be seen on the macroscopic scale is required. While experimental and theoretical methods exist to study the spectral properties of light-harvesting complexes on both individual complex and bulk ensemble levels, they have been developed largely independently of each other. To fill this gap, we simultaneously analyze experimental low-temperature single-complex and bulk ensemble optical spectra of the light-harvesting complex-2 (LH2) chromoproteins from the photosynthetic bacterium Rhodopseudomonas acidophila in order to find a unique theoretical model consistent with both experimental situations. The model, which satisfies most of the observations, combines strong exciton-phonon coupling with significant disorder, characteristic of the proteins. We establish a detailed disorder model that, in addition to containing a C2-symmetrical modulation of the site energies, distinguishes between static intercomplex and slow conformational intracomplex disorders. The model evaluations also verify that, despite best efforts, the single-LH2-complex measurements performed so far may be biased toward complexes with higher Huang-Rhys factors.

  14. Engineering Lipid Bilayer Membranes for Protein Studies

    PubMed Central

    Khan, Muhammad Shuja; Dosoky, Noura Sayed; Williams, John Dalton

    2013-01-01

    Lipid membranes regulate the flow of nutrients and communication signaling between cells and protect the sub-cellular structures. Recent attempts to fabricate artificial systems using nanostructures that mimic the physiological properties of natural lipid bilayer membranes (LBM) fused with transmembrane proteins have helped demonstrate the importance of temperature, pH, ionic strength, adsorption behavior, conformational reorientation and surface density in cellular membranes which all affect the incorporation of proteins on solid surfaces. Much of this work is performed on artificial templates made of polymer sponges or porous materials based on alumina, mica, and porous silicon (PSi) surfaces. For example, porous silicon materials have high biocompatibility, biodegradability, and photoluminescence, which allow them to be used both as a support structure for lipid bilayers or a template to measure the electrochemical functionality of living cells grown over the surface as in vivo. The variety of these media, coupled with the complex physiological conditions present in living systems, warrant a summary and prospectus detailing which artificial systems provide the most promise for different biological conditions. This study summarizes the use of electrochemical impedance spectroscopy (EIS) data on artificial biological membranes that are closely matched with previously published biological systems using both black lipid membrane and patch clamp techniques. PMID:24185908

  15. MemProtMD: Automated Insertion of Membrane Protein Structures into Explicit Lipid Membranes

    PubMed Central

    Stansfeld, Phillip J.; Goose, Joseph E.; Caffrey, Martin; Carpenter, Elisabeth P.; Parker, Joanne L.; Newstead, Simon; Sansom, Mark S.P.

    2015-01-01

    Summary There has been exponential growth in the number of membrane protein structures determined. Nevertheless, these structures are usually resolved in the absence of their lipid environment. Coarse-grained molecular dynamics (CGMD) simulations enable insertion of membrane proteins into explicit models of lipid bilayers. We have automated the CGMD methodology, enabling membrane protein structures to be identified upon their release into the PDB and embedded into a membrane. The simulations are analyzed for protein-lipid interactions, identifying lipid binding sites, and revealing local bilayer deformations plus molecular access pathways within the membrane. The coarse-grained models of membrane protein/bilayer complexes are transformed to atomistic resolution for further analysis and simulation. Using this automated simulation pipeline, we have analyzed a number of recently determined membrane protein structures to predict their locations within a membrane, their lipid/protein interactions, and the functional implications of an enhanced understanding of the local membrane environment of each protein. PMID:26073602

  16. Subdiffusion of proteins and oligomers on membranes

    NASA Astrophysics Data System (ADS)

    Lepzelter, David; Zaman, Muhammad

    2012-11-01

    Diffusion of proteins on lipid membranes plays a central role in cell signaling processes. From a mathematical perspective, most membrane diffusion processes are explained by the Saffman-Delbrück theory. However, recent studies have suggested a major limitation in the theoretical framework, the lack of complexity in the modeled lipid membrane. Lipid domains (sometimes termed membrane rafts) are known to slow protein diffusion, but there have been no quantitative theoretical examinations of how much diffusion is slowed in a general case. We provide an overall theoretical framework for confined-domain ("corralled") diffusion. Further, there have been multiple apparent contradictions of the basic conclusions of Saffman and Delbrück, each involving cases in which a single protein or an oligomer has multiple transmembrane regions passing through a lipid phase barrier. We present a set of corrections to the Saffman-Delbrück theory to account for these experimental observations. Our corrections are able to provide a quantitative explanation of numerous cellular signaling processes that have been considered beyond the scope of the Saffman-Delbrück theory, and may be extendable to other forms of subdiffusion.

  17. Helix-packing motifs in membrane proteins.

    PubMed

    Walters, R F S; DeGrado, W F

    2006-09-12

    The fold of a helical membrane protein is largely determined by interactions between membrane-imbedded helices. To elucidate recurring helix-helix interaction motifs, we dissected the crystallographic structures of membrane proteins into a library of interacting helical pairs. The pairs were clustered according to their three-dimensional similarity (rmsd membrane proteins.

  18. When physics takes over: BAR proteins and membrane curvature

    PubMed Central

    Simunovic, Mijo; Voth, Gregory A.; Callan-Jones, Andrew; Bassereau, Patricia

    2016-01-01

    Cell membranes become highly curved during membrane trafficking, cytokinesis, infection, immune response or cell motion. Bin/amphiphysin/Rvs (BAR) domain proteins with their intrinsically curved and anisotropic shape are involved in many of these processes, but with a large spectrum of modes of action. In vitro experiments and multiscale computer simulations have contributed in identifying a minimal set of physical parameters, namely protein density on the membrane, membrane tension, and membrane shape, that control how bound BAR domain proteins behave on the membrane. In this review, we summarize the multifaceted coupling of BAR proteins to membrane mechanics and propose a simple phase diagram that recapitulates the effects of these parameters. PMID:26519988

  19. Identification of extracellularly phosphorylated membrane proteins.

    PubMed

    Burghoff, Sandra; Willberg, Wibke; Schrader, Jürgen

    2015-10-01

    Ecto-protein kinases phosphorylate extracellular membrane proteins and exhibit similarities to casein kinases and protein kinases A and C. However, the identification of their protein substrates still remains a challenge because a clear separation from intracellular phosphoproteins is difficult. Here, we describe a straightforward method for the identification of extracellularly phosphorylated membrane proteins in human umbilical vein endothelial cells (HUVECs) and K562 cells which used the protease bromelain to selectively remove ectoproteins from intact cells and combined this with the subsequent analysis using IMAC and LC-MS/MS. A "false-positive" strategy in which cells without protease treatment served as controls was applied. Using this approach we identified novel phosphorylation sites on five ectophosphoproteins (NOTCH1, otopetrin 1, regulator of G-protein signalling 13 (RGS13), protein tyrosine phosphatase receptor type D isoform 3 (PTPRD), usherin isoform B (USH2A)). Use of bromelain appears to be a reliable technique for the further identification of phosphorylated surface-exposed peptides when extracellular adenosine-5'-triphosphate is elevated during purinergic signalling.

  20. Protein permeation through an electrically tunable membrane

    NASA Astrophysics Data System (ADS)

    Jou, Ining A.; Melnikov, Dmitriy V.; Gracheva, Maria E.

    2016-05-01

    Protein filtration is important in many fields of science and technology such as medicine, biology, chemistry, and engineering. Recently, protein separation and filtering with nanoporous membranes has attracted interest due to the possibility of fast separation and high throughput volume. This, however, requires understanding of the protein’s dynamics inside and in the vicinity of the nanopore. In this work, we utilize a Brownian dynamics approach to study the motion of the model protein insulin in the membrane–electrolyte electrostatic potential. We compare the results of the atomic model of the protein with the results of a coarse-grained and a single-bead model, and find that the coarse-grained representation of protein strikes the best balance between the accuracy of the results and the computational effort required. Contrary to common belief, we find that to adequately describe the protein, a single-bead model cannot be utilized without a significant effort to tabulate the simulation parameters. Similar to results for nanoparticle dynamics, our findings also indicate that the electric field and the electro-osmotic flow due to the applied membrane and electrolyte biases affect the capture and translocation of the biomolecule by either attracting or repelling it to or from the nanopore. Our computational model can also be applied to other types of proteins and separation conditions.

  1. Exploiting Microbeams for Membrane Protein Structure Determination.

    PubMed

    Warren, Anna J; Axford, Danny; Paterson, Neil G; Owen, Robin L

    2016-01-01

    A reproducible, and sample independent means of predictably obtaining large, well-ordered crystals has proven elusive in macromolecular crystallography. In the structure determination pipeline, crystallisation often proves to be a rate-limiting step, and the process of obtaining even small or badly ordered crystals can prove time-consuming and laborious. This is particularly true in the field of membrane protein crystallography and this is reflected in the limited number of unique membrane protein structures deposited in the protein data bank (less than 650 by June 2016 - http://blanco.biomol.uci.edu/mpstruc ). Over recent years the requirement for, and time and cost associated with obtaining, large crystals has been partially alleviated through the development of beamline instrumentation allowing data collection, and structure solution, from ever-smaller crystals. Advances in several areas have led to a step change in what might be considered achievable during a synchrotron trip over the last decade. This chapter will briefly review the current status of the field, the tools available to ease data collection and processing, and give some examples of exploitation of these for membrane protein microfocus macromolecular crystallography. PMID:27553238

  2. Efficient light-harvesting using non-carbonyl carotenoids: Energy transfer dynamics in the VCP complex from Nannochloropsis oceanica.

    PubMed

    Keşan, Gürkan; Litvín, Radek; Bína, David; Durchan, Milan; Šlouf, Václav; Polívka, Tomáš

    2016-04-01

    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%. PMID:26744091

  3. Efficient light-harvesting using non-carbonyl carotenoids: Energy transfer dynamics in the VCP complex from Nannochloropsis oceanica.

    PubMed

    Keşan, Gürkan; Litvín, Radek; Bína, David; Durchan, Milan; Šlouf, Václav; Polívka, Tomáš

    2016-04-01

    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%.

  4. Membrane association of presynaptic cytomatrix protein bassoon.

    PubMed

    Sanmartí-Vila, L; tom Dieck, S; Richter, K; Altrock, W; Zhang, L; Volknandt, W; Zimmermann, H; Garner, C C; Gundelfinger, E D; Dresbach, T

    2000-08-18

    Components of the specialized cytomatrix at active zones of presynaptic nerve terminals are thought to be involved in organizing synaptic events such as immobilisation or translocation of synaptic vesicles and assemblingactive zone components. The 420-kDa non-transmembraneprotein Bassoon is a specific componentof the presynaptic cytomatrix that shares features with both cytoskeleton-associated and peripheral-membrane proteins. Using immunogold electron microscopy we show here that synapse associated Bassoon is distributed in a subregion of active zones. Using a biochemical assay we show that a fraction of Bassoon is membrane associated. Electron microscopy performed on the same biochemical fraction further revealed that Bassoon is associated with vesicular structures. Together these data suggest that at least a fraction of Bassoon is associated with a membraneous compartment in neurons.

  5. Self-assembly strategies for integrating light harvesting and charge separation in artificial photosynthetic systems.

    PubMed

    Wasielewski, Michael R

    2009-12-21

    In natural photosynthesis, organisms optimize solar energy conversion through organized assemblies of photofunctional chromophores and catalysts within proteins that provide specifically tailored environments for chemical reactions. As with their natural counterparts, artificial photosynthetic systems for practical solar fuels production must collect light energy, separate charge, and transport charge to catalytic sites where multielectron redox processes will occur. While encouraging progress has been made on each aspect of this complex problem, researchers have not yet developed self-ordering and self-assembling components and the tailored environments necessary to realize a fully-functional artificial system. Previously researchers have used complex, covalent molecular systems comprised of chromophores, electron donors, and electron acceptors to mimic both the light-harvesting and the charge separation functions of photosynthetic proteins. These systems allow for study of the dependencies of electron transfer rate constants on donor-acceptor distance and orientation, electronic interaction, and the free energy of the reaction. The most useful and informative systems are those in which structural constraints control both the distance and the orientation between the electron donors and acceptors. Self-assembly provides a facile means for organizing large numbers of molecules into supramolecular structures that can bridge length scales from nanometers to macroscopic dimensions. The resulting structures must provide pathways for migration of light excitation energy among antenna chromophores, and from antennas to reaction centers. They also must incorporate charge conduits, that is, molecular "wires" that can efficiently move electrons and holes between reaction centers and catalytic sites. The central scientific challenge is to develop small, functional building blocks with a minimum number of covalent linkages, which also have the appropriate molecular recognition

  6. Strong antenna-enhanced fluorescence of a single light-harvesting complex shows photon antibunching.

    PubMed

    Wientjes, Emilie; Renger, Jan; Curto, Alberto G; Cogdell, Richard; van Hulst, Niek F

    2014-06-23

    The nature of the highly efficient energy transfer in photosynthetic light-harvesting complexes is a subject of intense research. Unfortunately, the low fluorescence efficiency and limited photostability hampers the study of individual light-harvesting complexes at ambient conditions. Here we demonstrate an over 500-fold fluorescence enhancement of light-harvesting complex 2 (LH2) at the single-molecule level by coupling to a gold nanoantenna. The resonant antenna produces an excitation enhancement of circa 100 times and a fluorescence lifetime shortening to ~20 ps. The radiative rate enhancement results in a 5.5-fold-improved fluorescence quantum efficiency. Exploiting the unique brightness, we have recorded the first photon antibunching of a single light-harvesting complex under ambient conditions, showing that the 27 bacteriochlorophylls coordinated by LH2 act as a non-classical single-photon emitter. The presented bright antenna-enhanced LH2 emission is a highly promising system to study energy transfer and the role of quantum coherence at the level of single complexes.

  7. Atomistic study of energy funneling in the light-harvesting complex of green sulfur bacteria.

    PubMed

    Huh, Joonsuk; Saikin, Semion K; Brookes, Jennifer C; Valleau, Stéphanie; Fujita, Takatoshi; Aspuru-Guzik, Alán

    2014-02-01

    Phototrophic organisms such as plants, photosynthetic bacteria, and algae use microscopic complexes of pigment molecules to absorb sunlight. Within the light-harvesting complexes, which frequently have several functional and structural subunits, the energy is transferred in the form of molecular excitations with very high efficiency. Green sulfur bacteria are considered to be among the most efficient light-harvesting organisms. Despite multiple experimental and theoretical studies of these bacteria, the physical origin of the efficient and robust energy transfer in their light-harvesting complexes is not well understood. To study excitation dynamics at the systems level, we introduce an atomistic model that mimics a complete light-harvesting apparatus of green sulfur bacteria. The model contains approximately 4000 pigment molecules and comprises a double wall roll for the chlorosome, a baseplate, and six Fenna-Matthews-Olson trimer complexes. We show that the fast relaxation within functional subunits combined with the transfer between collective excited states of pigments can result in robust energy funneling to the initial excitation conditions and temperature changes. Moreover, the same mechanism describes the coexistence of multiple time scales of excitation dynamics frequently observed in ultrafast optical experiments. While our findings support the hypothesis of supertransfer, the model reveals energy transport through multiple channels on different length scales.

  8. Bio serves nano: biological light-harvesting complex as energy donor for semiconductor quantum dots.

    PubMed

    Werwie, Mara; Xu, Xiangxing; Haase, Mathias; Basché, Thomas; Paulsen, Harald

    2012-04-01

    Light-harvesting complex (LHCII) of the photosynthetic apparatus in plants is attached to type-II core-shell CdTe/CdSe/ZnS nanocrystals (quantum dots, QD) exhibiting an absorption band at 710 nm and carrying a dihydrolipoic acid coating for water solubility. LHCII stays functional upon binding to the QD surface and enhances the light utilization of the QDs significantly, similar to its light-harvesting function in photosynthesis. Electronic excitation energy transfer of about 50% efficiency is shown by donor (LHCII) fluorescence quenching as well as sensitized acceptor (QD) emission and corroborated by time-resolved fluorescence measurements. The energy transfer efficiency is commensurable with the expected efficiency calculated according to Förster theory on the basis of the estimated donor-acceptor separation. Light harvesting is particularly efficient in the red spectral domain where QD absorption is relatively low. Excitation over the entire visible spectrum is further improved by complementing the biological pigments in LHCII with a dye attached to the apoprotein; the dye has been chosen to absorb in the "green gap" of the LHCII absorption spectrum and transfers its excitation energy ultimately to QD. This is the first report of a biological light-harvesting complex serving an inorganic semiconductor nanocrystal. Due to the charge separation between the core and the shell in type-II QDs the presented LHCII-QD hybrid complexes are potentially interesting for sensitized charge-transfer and photovoltaic applications.

  9. Studying the Effect of Light Quality on the Size of the Photosystem II Light Harvesting Complex

    ERIC Educational Resources Information Center

    Muhoz, Romualdo; Quiles, Maria J.

    2003-01-01

    In this article the effect of light quality on the size of the photosystem II (PSII) light harvesting complex (LHCII) is studied by measuring the chlorophyll fluorescence emitted by leaf sections of oat ("Avena sativa," var. Prevision) plants previously treated with either white light or with light filtered through blue, green, red or farred…

  10. Membrane Compartmentalization Reducing the Mobility of Lipids and Proteins within a Model Plasma Membrane.

    PubMed

    Koldsø, Heidi; Reddy, Tyler; Fowler, Philip W; Duncan, Anna L; Sansom, Mark S P

    2016-09-01

    The cytoskeleton underlying cell membranes may influence the dynamic organization of proteins and lipids within the bilayer by immobilizing certain transmembrane (TM) proteins and forming corrals within the membrane. Here, we present coarse-grained resolution simulations of a biologically realistic membrane model of asymmetrically organized lipids and TM proteins. We determine the effects of a model of cytoskeletal immobilization of selected membrane proteins using long time scale coarse-grained molecular dynamics simulations. By introducing compartments with varying degrees of restraints within the membrane models, we are able to reveal how compartmentalization caused by cytoskeletal immobilization leads to reduced and anomalous diffusional mobility of both proteins and lipids. This in turn results in a reduced rate of protein dimerization within the membrane and of hopping of membrane proteins between compartments. These simulations provide a molecular realization of hierarchical models often invoked to explain single-molecule imaging studies of membrane proteins.

  11. Outer membrane proteins of pathogenic spirochetes

    PubMed Central

    Cullen, Paul A.; Haake, David A.; Adler, Ben

    2009-01-01

    Pathogenic spirochetes are the causative agents of several important diseases including syphilis, Lyme disease, leptospirosis, swine dysentery, periodontal disease and some forms of relapsing fever. Spirochetal bacteria possess two membranes and the proteins present in the outer membrane are at the site of interaction with host tissue and the immune system. This review describes the current knowledge in the field of spirochetal outer membrane protein (OMP) biology. What is known concerning biogenesis and structure of OMPs, with particular regard to the atypical signal peptide cleavage sites observed amongst the spirochetes, is discussed. We examine the functions that have been determined for several spirochetal OMPs including those that have been demonstrated to function as adhesins, porins or to have roles in complement resistance. A detailed description of the role of spirochetal OMPs in immunity, including those that stimulate protective immunity or that are involved in antigenic variation, is given. A final section is included which covers experimental considerations in spirochetal outer membrane biology. This section covers contentious issues concerning cellular localization of putative OMPs, including determination of surface exposure. A more detailed knowledge of spirochetal OMP biology will hopefully lead to the design of new vaccines and a better understanding of spirochetal pathogenesis. PMID:15449605

  12. Hybrid artificial photosynthetic systems comprising semiconductors as light harvesters and biomimetic complexes as molecular cocatalysts.

    PubMed

    Wen, Fuyu; Li, Can

    2013-11-19

    Solar fuel production through artificial photosynthesis may be a key to generating abundant and clean energy, thus addressing the high energy needs of the world's expanding population. As the crucial components of photosynthesis, the artificial photosynthetic system should be composed of a light harvester (e.g., semiconductor or molecular dye), a reduction cocatalyst (e.g., hydrogenase mimic, noble metal), and an oxidation cocatalyst (e.g., photosystem II mimic for oxygen evolution from water oxidation). Solar fuel production catalyzed by an artificial photosynthetic system starts from the absorption of sunlight by the light harvester, where charge separation takes place, followed by a charge transfer to the reduction and oxidation cocatalysts, where redox reaction processes occur. One of the most challenging problems is to develop an artificial photosynthetic solar fuel production system that is both highly efficient and stable. The assembly of cocatalysts on the semiconductor (light harvester) not only can facilitate the charge separation, but also can lower the activation energy or overpotential for the reactions. An efficient light harvester loaded with suitable reduction and oxidation cocatalysts is the key for high efficiency of artificial photosynthetic systems. In this Account, we describe our strategy of hybrid photocatalysts using semiconductors as light harvesters with biomimetic complexes as molecular cocatalysts to construct efficient and stable artificial photosynthetic systems. We chose semiconductor nanoparticles as light harvesters because of their broad spectral absorption and relatively robust properties compared with a natural photosynthesis system. Using biomimetic complexes as cocatalysts can significantly facilitate charge separation via fast charge transfer from the semiconductor to the molecular cocatalysts and also catalyze the chemical reactions of solar fuel production. The hybrid photocatalysts supply us with a platform to study the

  13. Serial Millisecond Crystallography of Membrane Proteins.

    PubMed

    Jaeger, Kathrin; Dworkowski, Florian; Nogly, Przemyslaw; Milne, Christopher; Wang, Meitian; Standfuss, Joerg

    2016-01-01

    Serial femtosecond crystallography (SFX) at X-ray free-electron lasers (XFELs) is a powerful method to determine high-resolution structures of pharmaceutically relevant membrane proteins. Recently, the technology has been adapted to carry out serial millisecond crystallography (SMX) at synchrotron sources, where beamtime is more abundant. In an injector-based approach, crystals grown in lipidic cubic phase (LCP) or embedded in viscous medium are delivered directly into the unattenuated beam of a microfocus beamline. Pilot experiments show the application of microjet-based SMX for solving the structure of a membrane protein and compatibility of the method with de novo phasing. Planned synchrotron upgrades, faster detectors and software developments will go hand-in-hand with developments at free-electron lasers to provide a powerful methodology for solving structures from microcrystals at room temperature, ligand screening or crystal optimization for time-resolved studies with minimal or no radiation damage. PMID:27553240

  14. Vibrational dynamics of plant light-harvesting complex LHC II investigated by quasi- and inelastic neutron scattering

    NASA Astrophysics Data System (ADS)

    Golub, Maksym; Irrgang, Klaus-Dieter; Rusevich, Leonid; Pieper, Jörg

    2015-01-01

    Vibrational dynamics of the light-harvesting complex II (LHC II) from spinach was investigated by quasi- and inelastic neutron scattering (QENS and INS) at three different temperatures of 80, 160, and 285 K. QENS/INS spectra of solubilised LHC II and of the corresponding buffer solution were obtained separately and exhibit characteristic inelastic features. After subtraction of the buffer contribution, the INS spectrum of LHC II reveals a distinct Boson peak at ˜ 2.5 meV at 80 K that shifts towards lower energies if the temperature is increased to 285 K. This effect is interpreted in terms of a "softening" of the protein matrix along with the dynamical transition at ˜ 240 K. Our findings indicate that INS is a valuable method to obtain the density of vibrational states not only at cryogenic, but also at physiological temperatures.

  15. Lipidic phase membrane protein serial femtosecond crystallography

    PubMed Central

    Johansson, Linda C; Arnlund, David; White, Thomas A; Katona, Gergely; DePonte, Daniel P; Weierstall, Uwe; Doak, R Bruce; Shoeman, Robert L; Lomb, Lukas; Malmerberg, Erik; Davidsson, Jan; Nass, Karol; Liang, Mengning; Andreasson, Jakob; Aquila, Andrew; Bajt, Sasa; Barthelmess, Miriam; Barty, Anton; Bogan, Michael J; Bostedt, Christoph; Bozek, John D; Caleman, Carl; Coffee, Ryan; Coppola, Nicola; Ekeberg, Tomas; Epp, Sascha W; Erk, Benjamin; Fleckenstein, Holger; Foucar, Lutz; Graafsma, Heinz; Gumprecht, Lars; Hajdu, Janos; Hampton, Christina Y; Hartmann, Robert; Hartmann, Andreas; Hauser, Günter; Hirsemann, Helmut; Holl, Peter; Hunter, Mark S; Kassemeyer, Stephan; Kimmel, Nils; Kirian, Richard A; Maia, Filipe R N C; Marchesini, Stefano; Martin, Andrew V; Reich, Christian; Rolles, Daniel; Rudek, Benedikt; Rudenko, Artem; Schlichting, Ilme; Schulz, Joachim; Seibert, M Marvin; Sierra, Raymond G; Soltau, Heike; Starodub, Dmitri; Stellato, Francesco; Stern, Stephan; Strüder, Lothar; Timneanu, Nicusor; Ullrich, Joachim; Wahlgren, Weixiao Y; Wang, Xiaoyu; Weidenspointner, Georg; Wunderer, Cornelia; Fromme, Petra; Chapman, Henry N; Spence, John C H; Neutze, Richard

    2012-01-01

    X-ray free electron laser (X-feL)-based serial femtosecond crystallography is an emerging method with potential to rapidly advance the challenging field of membrane protein structural biology. here we recorded interpretable diffraction data from micrometer-sized lipidic sponge phase crystals of the Blastochloris viridis photosynthetic reaction center delivered into an X-feL beam using a sponge phase micro-jet. PMID:22286383

  16. Lipidic phase membrane protein serial femtosecond crystallography.

    PubMed

    Johansson, Linda C; Arnlund, David; White, Thomas A; Katona, Gergely; Deponte, Daniel P; Weierstall, Uwe; Doak, R Bruce; Shoeman, Robert L; Lomb, Lukas; Malmerberg, Erik; Davidsson, Jan; Nass, Karol; Liang, Mengning; Andreasson, Jakob; Aquila, Andrew; Bajt, Saša; Barthelmess, Miriam; Barty, Anton; Bogan, Michael J; Bostedt, Christoph; Bozek, John D; Caleman, Carl; Coffee, Ryan; Coppola, Nicola; Ekeberg, Tomas; Epp, Sascha W; Erk, Benjamin; Fleckenstein, Holger; Foucar, Lutz; Graafsma, Heinz; Gumprecht, Lars; Hajdu, Janos; Hampton, Christina Y; Hartmann, Robert; Hartmann, Andreas; Hauser, Günter; Hirsemann, Helmut; Holl, Peter; Hunter, Mark S; Kassemeyer, Stephan; Kimmel, Nils; Kirian, Richard A; Maia, Filipe R N C; Marchesini, Stefano; Martin, Andrew V; Reich, Christian; Rolles, Daniel; Rudek, Benedikt; Rudenko, Artem; Schlichting, Ilme; Schulz, Joachim; Seibert, M Marvin; Sierra, Raymond G; Soltau, Heike; Starodub, Dmitri; Stellato, Francesco; Stern, Stephan; Strüder, Lothar; Timneanu, Nicusor; Ullrich, Joachim; Wahlgren, Weixiao Y; Wang, Xiaoyu; Weidenspointner, Georg; Wunderer, Cornelia; Fromme, Petra; Chapman, Henry N; Spence, John C H; Neutze, Richard

    2012-03-01

    X-ray free electron laser (X-FEL)-based serial femtosecond crystallography is an emerging method with potential to rapidly advance the challenging field of membrane protein structural biology. Here we recorded interpretable diffraction data from micrometer-sized lipidic sponge phase crystals of the Blastochloris viridis photosynthetic reaction center delivered into an X-FEL beam using a sponge phase micro-jet.

  17. Lipidic phase membrane protein serial femtosecond crystallography.

    PubMed

    Johansson, Linda C; Arnlund, David; White, Thomas A; Katona, Gergely; Deponte, Daniel P; Weierstall, Uwe; Doak, R Bruce; Shoeman, Robert L; Lomb, Lukas; Malmerberg, Erik; Davidsson, Jan; Nass, Karol; Liang, Mengning; Andreasson, Jakob; Aquila, Andrew; Bajt, Saša; Barthelmess, Miriam; Barty, Anton; Bogan, Michael J; Bostedt, Christoph; Bozek, John D; Caleman, Carl; Coffee, Ryan; Coppola, Nicola; Ekeberg, Tomas; Epp, Sascha W; Erk, Benjamin; Fleckenstein, Holger; Foucar, Lutz; Graafsma, Heinz; Gumprecht, Lars; Hajdu, Janos; Hampton, Christina Y; Hartmann, Robert; Hartmann, Andreas; Hauser, Günter; Hirsemann, Helmut; Holl, Peter; Hunter, Mark S; Kassemeyer, Stephan; Kimmel, Nils; Kirian, Richard A; Maia, Filipe R N C; Marchesini, Stefano; Martin, Andrew V; Reich, Christian; Rolles, Daniel; Rudek, Benedikt; Rudenko, Artem; Schlichting, Ilme; Schulz, Joachim; Seibert, M Marvin; Sierra, Raymond G; Soltau, Heike; Starodub, Dmitri; Stellato, Francesco; Stern, Stephan; Strüder, Lothar; Timneanu, Nicusor; Ullrich, Joachim; Wahlgren, Weixiao Y; Wang, Xiaoyu; Weidenspointner, Georg; Wunderer, Cornelia; Fromme, Petra; Chapman, Henry N; Spence, John C H; Neutze, Richard

    2012-03-01

    X-ray free electron laser (X-FEL)-based serial femtosecond crystallography is an emerging method with potential to rapidly advance the challenging field of membrane protein structural biology. Here we recorded interpretable diffraction data from micrometer-sized lipidic sponge phase crystals of the Blastochloris viridis photosynthetic reaction center delivered into an X-FEL beam using a sponge phase micro-jet. PMID:22286383

  18. Mammalian plasma membrane proteins as potential biomarkers and drug targets.

    PubMed

    Rucevic, Marijana; Hixson, Douglas; Josic, Djuro

    2011-06-01

    Defining the plasma membrane proteome is crucial to understand the role of plasma membrane in fundamental biological processes. Change in membrane proteins is one of the first events that take place under pathological conditions, making plasma membrane proteins a likely source of potential disease biomarkers with prognostic or diagnostic potential. Membrane proteins are also potential targets for monoclonal antibodies and other drugs that block receptors or inhibit enzymes essential to the disease progress. Despite several advanced methods recently developed for the analysis of hydrophobic proteins and proteins with posttranslational modifications, integral membrane proteins are still under-represented in plasma membrane proteome. Recent advances in proteomic investigation of plasma membrane proteins, defining their roles as diagnostic and prognostic disease biomarkers and as target molecules in disease treatment, are presented.

  19. Membrane-associated proteins and peptides.

    PubMed

    Lensink, Marc F

    2015-01-01

    This chapter discusses the practical aspects of setting up molecular dynamics simulations of membrane-associated proteins and peptides, and the analysis thereof. Topology files for selected lipids are provided and selected analysis tools presented. These include tools for the creation of lipid bilayers of mixed lipid content (DOPE) and easy extraction of lipid coordinates (g_zcoor, g_xycoor), the calculation of helical axes (g_helixaxis) and aromatic order parameters (g_arom), the determination of peptide- or protein-interacting lipids (g_under), and the investigation of lipid-specific interactions through the calculation of lipid-bridged residue-residue contacts (g_prolip). PMID:25330961

  20. Thermal Quantum Correlations in Photosynthetic Light-Harvesting Complexes

    NASA Astrophysics Data System (ADS)

    Mahdian, M.; Kouhestani, H.

    2015-08-01

    Photosynthesis is one of the ancient biological processes, playing crucial role converting solar energy to cellular usable currency. Environmental factors and external perturbations has forced nature to choose systems with the highest efficiency and performance. Recent theoretical and experimental studies have proved the presence of quantum properties in biological systems. Energy transfer systems like Fenna-Matthews-Olson (FMO) complex shows quantum entanglement between sites of Bacteriophylla molecules in protein environment and presence of decoherence. Complex biological systems implement more truthful mechanisms beside chemical-quantum correlations to assure system's efficiency. In this study we investigate thermal quantum correlations in FMO protein of the photosynthetic apparatus of green sulfur bacteria by quantum discord measure. The results confirmed existence of remarkable quantum correlations of of BChla pigments in room temperature. This results approve involvement of quantum correlation mechanisms for information storage and retention in living organisms that could be useful for further evolutionary studies. Inspired idea of this study is potentially interesting to practice by the same procedure in genetic data transfer mechanisms.

  1. Disentangling the low-energy states of the major light-harvesting complex of plants and their role in photoprotection.

    PubMed

    Krüger, Tjaart P J; Ilioaia, Cristian; Johnson, Matthew P; Ruban, Alexander V; van Grondelle, Rienk

    2014-07-01

    The ability to dissipate large fractions of their absorbed light energy as heat is a vital photoprotective function of the peripheral light-harvesting pigment-protein complexes in photosystem II of plants. The major component of this process, known as qE, is characterised by the appearance of low-energy (red-shifted) absorption and fluorescence bands. Although the appearance of these red states has been established, the molecular mechanism, their site and particularly their involvement in qE are strongly debated. Here, room-temperature single-molecule fluorescence spectroscopy was used to study the red emission states of the major plant light-harvesting complex (LHCII) in different environments, in particular conditions mimicking qE. It was found that most states correspond to peak emission at around 700nm and are unrelated to energy dissipative states, though their frequency of occurrence increased under conditions that mimicked qE. Longer-wavelength emission appeared to be directly related to energy dissipative states, in particular emission beyond 770nm. The ensemble average of the red emission bands shares many properties with those obtained from previous bulk in vitro and in vivo studies. We propose the existence of at least three excitation energy dissipating mechanisms in LHCII, each of which is associated with a different spectral signature and whose contribution to qE is determined by environmental control of protein conformational disorder. Emission at 700nm is attributed to a conformational change in the Lut 2 domain, which is facilitated by the conformational change associated with the primary quenching mechanism involving Lut 1.

  2. Electrophoretic transfer of proteins across polyacrylamide membranes.

    PubMed

    Rylatt, D B; Napoli, M; Ogle, D; Gilbert, A; Lim, S; Nair, C H

    1999-12-31

    The electrophoretic transfer of purified proteins has been examined in a Gradiflow "Babyflow BF100" unit. A number of factors affect protein separation within this preparative electrophoresis system. We established that the rate of protein transfer was proportional to the applied voltage. The transfer is slowest at the isoelectric point (pI) and increased the further away the pH was from the pI of the protein. Protein transfer was found to be independent of the ionic strength of the buffer, for buffers that excluded the addition of strong acids or strong bases or sodium chloride. Transfer decreased as the pore size of the membrane decreased. Finally, transfer was inhibited at high salt concentrations in the protein solution, but remained unaffected when urea and non-ionic detergents were added to the solution. To increase the speed of protein separations, buffers with low conductivity should be used. A pH for the optimal separation should be selected on the basis of the relative pI and size of the target proteins and that of the major contaminants. PMID:10674937

  3. Crystallizing Membrane Proteins Using Lipidic Mesophases

    PubMed Central

    Caffrey, Martin; Cherezov, Vadim

    2009-01-01

    A detailed protocol for crystallizing membrane proteins that makes use of lipidic mesophases is described. This has variously been referred to as the lipid cubic phase or in meso method. The method has been shown to be quite general in that it has been used to solve X-ray crystallographic structures of prokaryotic and eukaryotic proteins, proteins that are monomeric, homo- and hetero-multimeric, chromophore-containing and chromophore-free, and α-helical and β-barrel proteins. Its most recent successes are the human engineered β2-adrenergic and adenosine A2A G protein-coupled receptors. Protocols are provided for preparing and characterizing the lipidic mesophase, for reconstituting the protein into the monoolein-based mesophase, for functional assay of the protein in the mesophase, and for setting up crystallizations in manual mode. Methods for harvesting micro-crystals are also described. The time required to prepare the protein-loaded mesophase and to set up a crystallization plate manually is about one hour. PMID:19390528

  4. Quantification of detergent using colorimetric methods in membrane protein crystallography.

    PubMed

    Prince, Chelsy; Jia, Zongchao

    2015-01-01

    Membrane protein crystallography has the potential to greatly aid our understanding of membrane protein biology. Yet, membrane protein crystals remain challenging to produce. Although robust methods for the expression and purification of membrane proteins continue to be developed, the detergent component of membrane protein samples is equally important to crystallization efforts. This chapter describes the development of three colorimetric assays for the quantitation of detergent in membrane protein samples and provides detailed protocols. All of these techniques use small sample volumes and have potential applications in crystallography. The application of these techniques in crystallization prescreening, detergent concentration modification, and detergent exchange experiments is demonstrated. It has been observed that the concentration of detergent in a membrane protein sample can be just as important as the protein concentration when attempting to reproduce crystallization lead conditions.

  5. Identifying the quantum correlations in light-harvesting complexes

    SciTech Connect

    Bradler, Kamil; Wilde, Mark M.; Vinjanampathy, Sai; Uskov, Dmitry B.

    2010-12-15

    One of the major efforts in the quantum biological program is to subject biological systems to standard tests or measures of quantumness. These tests and measures should elucidate whether nontrivial quantum effects may be present in biological systems. Two such measures of quantum correlations are the quantum discord and the relative entropy of entanglement. Here, we show that the relative entropy of entanglement admits a simple analytic form when dynamics and accessible degrees of freedom are restricted to a zero- and single-excitation subspace. We also simulate and calculate the amount of quantum discord that is present in the Fenna-Matthews-Olson protein complex during the transfer of an excitation from a chlorosome antenna to a reaction center. We find that the single-excitation quantum discord and single-excitation relative entropy of entanglement are equal for all of our numerical simulations, but a proof of their general equality for this setting evades us for now. Also, some of our simulations demonstrate that the relative entropy of entanglement without the single-excitation restriction is much lower than the quantum discord. The first picosecond of dynamics is the relevant time scale for the transfer of the excitation, according to some sources in the literature. Our simulation results indicate that quantum correlations contribute a significant fraction of the total correlation during this first picosecond in many cases, at both cryogenic and physiological temperatures.

  6. Practical considerations of membrane protein instability during purification and crystallisation.

    PubMed

    Tate, Christopher G

    2010-01-01

    Crystallisation of integral membranes requires milligrams of purified protein in a homogeneous, monodisperse state, and crucially, the membrane protein must also be fully functional and stable. The stability of membrane proteins in solution is dependent on the type of detergents used, but unfortunately the use of the most stabilising detergent can often decrease the probability of obtaining crystals that diffract to high resolution, especially of small membrane proteins. A number of strategies have been developed to facilitate the purification of membrane proteins in a functional form, which have led to new possibilities for structure determination.

  7. Evolution under the sun: optimizing light harvesting in photosynthesis.

    PubMed

    Ruban, Alexander V

    2015-01-01

    The emergence and evolution of life on our planet was possible because the sun provides energy to our biosphere. Indeed, all life forms need energy for existence and proliferation in space and time. Light-energy conversion takes place in photosynthetic organisms that evolve in various environments featuring an impressive range of light intensities that span several orders of magnitude. This property is achieved by the evolution of mechanisms of efficient energy capture that involved development of antenna pigments and pigment-protein complexes as well as the emergence of various strategies on the organismal, cellular, and molecular levels to counteract the detrimental effects of high light intensity on the delicate photosynthetic apparatus. Darwin was one of the first to describe the behaviour of plants towards light. He noticed that some plants try to avoid full daylight and called this reaction paraheliotropism. However, it was only in the second half of the 20th century, when scientists began to discover the structure and molecular mechanisms of the photosynthetic machinery, that the reasons for paraheliotropisms became clear. This review explains the need for the evolution of light adaptations using the example of higher plants. The review focuses on short-term adaptation mechanisms that occur on the minute scale, showing that these processes are fast enough to track rapid fluctuations in light intensity and that they evolved to be effective, allowing for the expansion of plant habitats and promoting diversification and survival. Also introduced are the most recent developments in methods that enable quantification of the light intensities that can be tolerated by plants.

  8. Reconstitution of the membrane protein OmpF into biomimetic block copolymer–phospholipid hybrid membranes

    PubMed Central

    Bieligmeyer, Matthias; Artukovic, Franjo; Hirth, Thomas; Schiestel, Thomas

    2016-01-01

    Summary Structure and function of many transmembrane proteins are affected by their environment. In this respect, reconstitution of a membrane protein into a biomimetic polymer membrane can alter its function. To overcome this problem we used membranes formed by poly(1,4-isoprene-block-ethylene oxide) block copolymers blended with 1,2-diphytanoyl-sn-glycero-3-phosphocholine. By reconstituting the outer membrane protein OmpF from Escherichia coli into these membranes, we demonstrate functionality of this protein in biomimetic lipopolymer membranes, independent of the molecular weight of the block copolymers. At low voltages, the channel conductance of OmpF in 1 M KCl was around 2.3 nS. In line with these experiments, integration of OmpF was also revealed by impedance spectroscopy. Our results indicate that blending synthetic polymer membranes with phospholipids allows for the reconstitution of transmembrane proteins under preservation of protein function, independent of the membrane thickness. PMID:27547605

  9. Reconstitution of the membrane protein OmpF into biomimetic block copolymer-phospholipid hybrid membranes.

    PubMed

    Bieligmeyer, Matthias; Artukovic, Franjo; Nussberger, Stephan; Hirth, Thomas; Schiestel, Thomas; Müller, Michaela

    2016-01-01

    Structure and function of many transmembrane proteins are affected by their environment. In this respect, reconstitution of a membrane protein into a biomimetic polymer membrane can alter its function. To overcome this problem we used membranes formed by poly(1,4-isoprene-block-ethylene oxide) block copolymers blended with 1,2-diphytanoyl-sn-glycero-3-phosphocholine. By reconstituting the outer membrane protein OmpF from Escherichia coli into these membranes, we demonstrate functionality of this protein in biomimetic lipopolymer membranes, independent of the molecular weight of the block copolymers. At low voltages, the channel conductance of OmpF in 1 M KCl was around 2.3 nS. In line with these experiments, integration of OmpF was also revealed by impedance spectroscopy. Our results indicate that blending synthetic polymer membranes with phospholipids allows for the reconstitution of transmembrane proteins under preservation of protein function, independent of the membrane thickness. PMID:27547605

  10. Calcium ions are involved in the unusual red shift of the light-harvesting 1 Qy transition of the core complex in thermophilic purple sulfur bacterium Thermochromatium tepidum.

    PubMed

    Kimura, Yukihiro; Hirano, Yu; Yu, Long-Jiang; Suzuki, Hiroaki; Kobayashi, Masayuki; Wang, Zheng-Yu

    2008-05-16

    Thermophilic purple sulfur bacterium, Thermochromatium tepidum, can grow at temperatures up to 58 degrees C and exhibits an unusual Qy absorption at 915 nm for the core light-harvesting complex (LH1), an approximately 35-nm red shift from those of its mesophilic counterparts. We demonstrate in this study, using a highly purified LH1-reaction center complex, that the LH1 Qy transition is strongly dependent on metal cations and Ca2+ is involved in the unusual red shift. Removal of the Ca2+ resulted in formation of a species with the LH1 Qy absorption at 880 nm, and addition of the Ca2+ to the 880-nm species recovered the native 915-nm form. Interchange between the two forms is fully reversible. Based on spectroscopic and isothermal titration calorimetry analyses, the Ca2+ binding to the LH1 complex was estimated to occur in a stoichiometric ratio of Ca2+/alphabeta-subunit = 1:1 and the binding constant was in 10(5) m(-1) order of magnitude, which is comparable with those for EF-hand Ca2+-binding proteins. Despite the high affinity, conformational changes in the LH1 complex upon Ca2+ binding were small and occurred slowly, with a typical time constant of approximately 6 min. Replacement of the Ca2+ with other metal cations caused blue shifts of the Qy bands depending on the property of the cations, indicating that the binding site is highly selective. Based on the amino acid sequences of the LH1 complex, possible Ca2+-binding sites are proposed that consist of several acidic amino acid residues near the membrane interfaces of the C-terminal region of the alpha-polypeptide and the N-terminal region of the beta-polypeptide.

  11. Hydrophobic mismatch sorts SNARE proteins into distinct membrane domains

    NASA Astrophysics Data System (ADS)

    Milovanovic, Dragomir; Honigmann, Alf; Koike, Seiichi; Göttfert, Fabian; Pähler, Gesa; Junius, Meike; Müllar, Stefan; Diederichsen, Ulf; Janshoff, Andreas; Grubmüller, Helmut; Risselada, Herre J.; Eggeling, Christian; Hell, Stefan W.; van den Bogaart, Geert; Jahn, Reinhard

    2015-01-01

    The clustering of proteins and lipids in distinct microdomains is emerging as an important principle for the spatial patterning of biological membranes. Such domain formation can be the result of hydrophobic and ionic interactions with membrane lipids as well as of specific protein-protein interactions. Here using plasma membrane-resident SNARE proteins as model, we show that hydrophobic mismatch between the length of transmembrane domains (TMDs) and the thickness of the lipid membrane suffices to induce clustering of proteins. Even when the TMDs differ in length by only a single residue, hydrophobic mismatch can segregate structurally closely homologous membrane proteins in distinct membrane domains. Domain formation is further fine-tuned by interactions with polyanionic phosphoinositides and homo and heterotypic protein interactions. Our findings demonstrate that hydrophobic mismatch contributes to the structural organization of membranes.

  12. Chiroptical nature of two-exciton states of light-harvesting complex: Doubly resonant three-wave-mixing spectroscopy

    NASA Astrophysics Data System (ADS)

    Lee, Hochan; Cheon, Sangheon; Cho, Minhaeng

    2010-06-01

    Photosynthetic light-harvesting complex is a coupled multichromophore system. Due to electronic couplings between neighboring chlorophylls in the complex, the one- and two-exciton states are delocalized and they can be written as linear combinations of singly and doubly excited configurations, respectively. Despite that the chiroptical properties of one-exciton states in such a multichromophore system have been investigated by using linear optical activity measurement techniques; those of two-exciton states have not been studied before due to a lack of appropriate measurement methods. Here, we present a theoretical description on chiroptical χ(2) spectroscopy and show that it can be used to investigate such properties of a photosynthetic light-harvesting system, which is the Fenna-Matthews-Olson complex, consisting of seven bacteriochlorophylls in its protein subunit. To simulate the doubly resonant sum- and difference-frequency-generation spectra of the complex, one- and two-exciton transition dipoles were calculated. Carrying out quantum chemistry calculations of electronically excited states of a model bacteriochlorophyll system and taking into account the dipole-induced dipole electronic transition processes between the ground state and two-exciton states, we could calculate the two-dimensional sum-frequency-generation spectra revealing dominant second-order chiroptical transition pathways and involved one- and two-exciton states. It is believed that the present computational scheme and the theoretically proposed doubly resonant two-dimensional three-wave-mixing spectroscopy would be of use to shed light on the chiroptical natures of two-exciton states of arbitrary coupled multichromophore systems.

  13. Maximizing photosynthetic efficiency and culture productivity in cyanobacteria upon minimizing the phycobilisome light-harvesting antenna size.

    PubMed

    Kirst, Henning; Formighieri, Cinzia; Melis, Anastasios

    2014-10-01

    A phycocyanin-deletion mutant of Synechocystis (cyanobacteria) was generated upon replacement of the CPC-operon with a kanamycin resistance cassette. The Δcpc transformant strains (Δcpc) exhibited a green phenotype, compared to the blue-green of the wild type (WT), lacked the distinct phycocyanin absorbance at 625nm, and had a lower Chl per cell content and a lower PSI/PSII reaction center ratio compared to the WT. Molecular and genetic analyses showed replacement of all WT copies of the Synechocystis DNA with the transgenic version, thereby achieving genomic DNA homoplasmy. Biochemical analyses showed the absence of the phycocyanin α- and β-subunits, and the overexpression of the kanamycin resistance NPTI protein in the Δcpc. Physiological analyses revealed a higher, by a factor of about 2, intensity for the saturation of photosynthesis in the Δcpc compared to the WT. Under limiting intensities of illumination, growth of the Δcpc was slower than that of the WT. This difference in the rate of cell duplication diminished gradually as growth irradiance increased. Identical rates of cell duplication of about 13h for both WT and Δcpc were observed at about 800μmolphotonsm(-2)s(-1) or greater. Culture productivity analyses under simulated bright sunlight and high cell-density conditions showed that biomass accumulation by the Δcpc was 1.57-times greater than that achieved by the WT. Thus, the work provides first-time direct evidence of the applicability of the Truncated Light-harvesting Antenna (TLA)-concept in cyanobacteria, entailing substantial improvements in the photosynthetic efficiency and productivity of mass cultures upon minimizing the phycobilisome light-harvesting antenna size.

  14. Anisotropic organization and microscopic manipulation of self-assembling synthetic porphyrin microrods that mimic chlorosomes: bacterial light-harvesting systems.

    PubMed

    Chappaz-Gillot, Cyril; Marek, Peter L; Blaive, Bruno J; Canard, Gabriel; Bürck, Jochen; Garab, Gyozo; Hahn, Horst; Jávorfi, Tamás; Kelemen, Loránd; Krupke, Ralph; Mössinger, Dennis; Ormos, Pál; Reddy, Chilla Malla; Roussel, Christian; Steinbach, Gábor; Szabó, Milán; Ulrich, Anne S; Vanthuyne, Nicolas; Vijayaraghavan, Aravind; Zupcanova, Anita; Balaban, Teodor Silviu

    2012-01-18

    Being able to control in time and space the positioning, orientation, movement, and sense of rotation of nano- to microscale objects is currently an active research area in nanoscience, having diverse nanotechnological applications. In this paper, we demonstrate unprecedented control and maneuvering of rod-shaped or tubular nanostructures with high aspect ratios which are formed by self-assembling synthetic porphyrins. The self-assembly algorithm, encoded by appended chemical-recognition groups on the periphery of these porphyrins, is the same as the one operating for chlorosomal bacteriochlorophylls (BChl's). Chlorosomes, rod-shaped organelles with relatively long-range molecular order, are the most efficient naturally occurring light-harvesting systems. They are used by green photosynthetic bacteria to trap visible and infrared light of minute intensities even at great depths, e.g., 100 m below water surface or in volcanic vents in the absence of solar radiation. In contrast to most other natural light-harvesting systems, the chlorosomal antennae are devoid of a protein scaffold to orient the BChl's; thus, they are an attractive goal for mimicry by synthetic chemists, who are able to engineer more robust chromophores to self-assemble. Functional devices with environmentally friendly chromophores-which should be able to act as photosensitizers within hybrid solar cells, leading to high photon-to-current conversion efficiencies even under low illumination conditions-have yet to be fabricated. The orderly manner in which the BChl's and their synthetic counterparts self-assemble imparts strong diamagnetic and optical anisotropies and flow/shear characteristics to their nanostructured assemblies, allowing them to be manipulated by electrical, magnetic, or tribomechanical forces. PMID:22148684

  15. A saposin-lipoprotein nanoparticle system for membrane proteins.

    PubMed

    Frauenfeld, Jens; Löving, Robin; Armache, Jean-Paul; Sonnen, Andreas F-P; Guettou, Fatma; Moberg, Per; Zhu, Lin; Jegerschöld, Caroline; Flayhan, Ali; Briggs, John A G; Garoff, Henrik; Löw, Christian; Cheng, Yifan; Nordlund, Pär

    2016-04-01

    A limiting factor in membrane protein research is the ability to solubilize and stabilize such proteins. Detergents are used most often for solubilizing membrane proteins, but they are associated with protein instability and poor compatibility with structural and biophysical studies. Here we present a saposin-lipoprotein nanoparticle system, Salipro, which allows for the reconstitution of membrane proteins in a lipid environment that is stabilized by a scaffold of saposin proteins. We demonstrate the applicability of the method on two purified membrane protein complexes as well as by the direct solubilization and nanoparticle incorporation of a viral membrane protein complex from the virus membrane. Our approach facilitated high-resolution structural studies of the bacterial peptide transporter PeptTSo2 by single-particle cryo-electron microscopy (cryo-EM) and allowed us to stabilize the HIV envelope glycoprotein in a functional state.

  16. Large-scale proteomic analysis of membrane proteins.

    PubMed

    Ahram, Mamoun; Springer, David L

    2004-10-01

    Proteomic analysis of membrane proteins is a promising approach for the identification of novel drug targets and/or disease biomarkers. Despite notable technological developments, obstacles related to extraction and solublization of membrane proteins are encountered. A critical discussion of the different preparative methods of membrane proteins is offered in relation to downstream proteomic applications, mainly gel-based analyses and mass spectrometry. Frequently, unknown proteins are identified by high-throughput profiling of membrane proteins. In search for novel membrane proteins, analysis of protein sequences using computational tools is performed to predict the presence of transmembrane domains. This review also presents these bioinformatic tools with the human proteome as a case study. Along with technological innovations, advancements in the areas of sample preparation and computational prediction of membrane proteins will lead to exciting discoveries.

  17. Dynamic nuclear polarization methods in solids and solutions to explore membrane proteins and membrane systems.

    PubMed

    Cheng, Chi-Yuan; Han, Songi

    2013-01-01

    Membrane proteins regulate vital cellular processes, including signaling, ion transport, and vesicular trafficking. Obtaining experimental access to their structures, conformational fluctuations, orientations, locations, and hydration in membrane environments, as well as the lipid membrane properties, is critical to understanding their functions. Dynamic nuclear polarization (DNP) of frozen solids can dramatically boost the sensitivity of current solid-state nuclear magnetic resonance tools to enhance access to membrane protein structures in native membrane environments. Overhauser DNP in the solution state can map out the local and site-specific hydration dynamics landscape of membrane proteins and lipid membranes, critically complementing the structural and dynamics information obtained by electron paramagnetic resonance spectroscopy. Here, we provide an overview of how DNP methods in solids and solutions can significantly increase our understanding of membrane protein structures, dynamics, functions, and hydration in complex biological membrane environments. PMID:23331309

  18. Dynamic Nuclear Polarization Methods in Solids and Solutions to Explore Membrane Proteins and Membrane Systems

    NASA Astrophysics Data System (ADS)

    Cheng, Chi-Yuan; Han, Songi

    2013-04-01

    Membrane proteins regulate vital cellular processes, including signaling, ion transport, and vesicular trafficking. Obtaining experimental access to their structures, conformational fluctuations, orientations, locations, and hydration in membrane environments, as well as the lipid membrane properties, is critical to understanding their functions. Dynamic nuclear polarization (DNP) of frozen solids can dramatically boost the sensitivity of current solid-state nuclear magnetic resonance tools to enhance access to membrane protein structures in native membrane environments. Overhauser DNP in the solution state can map out the local and site-specific hydration dynamics landscape of membrane proteins and lipid membranes, critically complementing the structural and dynamics information obtained by electron paramagnetic resonance spectroscopy. Here, we provide an overview of how DNP methods in solids and solutions can significantly increase our understanding of membrane protein structures, dynamics, functions, and hydration in complex biological membrane environments.

  19. Mass Spectrometry of Membrane Proteins: A Focus on Aquaporins

    PubMed Central

    Schey, Kevin L.; Grey, Angus C.; Nicklay, Joshua J.

    2015-01-01

    Membrane proteins are abundant, critically important biomolecules that conduct essential functions in all cells and are the targets of a significant number of therapeutic drugs. However, the analysis of their expression, modification, protein–protein interactions, and structure by mass spectrometry has lagged behind similar studies of soluble proteins. Here we review the limitations to analysis of integral membrane and membrane-associated proteins and highlight advances in sample preparation and mass spectrometry methods that have led to the successful analysis of this protein class. Advances in the analysis of membrane protein posttranslational modification, protein–protein interaction, protein structure, and tissue distributions by imaging mass spectrometry are discussed. Furthermore, we focus our discussion on the application of mass spectrometry for the analysis of aquaporins as a prototypical integral membrane protein and how advances in analytical methods have revealed new biological insights into the structure and function of this family of proteins. PMID:23394619

  20. Proteopolymersomes: in vitro production of a membrane protein in polymersome membranes.

    PubMed

    Nallani, Madhavan; Andreasson-Ochsner, Mirjam; Tan, Cherng-Wen Darren; Sinner, Eva-Kathrin; Wisantoso, Yudi; Geifman-Shochat, Susana; Hunziker, Walter

    2011-12-01

    Polymersomes are stable self-assembled architectures which mimic cell membranes. For characterization, membrane proteins can be incorporated into such bio-mimetic membranes by reconstitution methods, leading to so-called proteopolymersomes. In this work, we demonstrate the direct incorporation of a membrane protein into polymersome membranes by a cell-free expression system. Firstly, we demonstrate pore formation in the preformed polymersome membrane using α-hemolysin. Secondly, we use claudin-2, a protein involved in cell-cell interactions, to demonstrate the in vitro expression of a membrane protein into these polymersomes. Surface plasmon resonance (Biacore) binding studies with the claudin-2 proteopolymersomes and claudin-2 specific antibodies are performed to show the presence of the in vitro expressed protein in polymersome membranes.

  1. A Prediction Model for Membrane Proteins Using Moments Based Features.

    PubMed

    Butt, Ahmad Hassan; Khan, Sher Afzal; Jamil, Hamza; Rasool, Nouman; Khan, Yaser Daanial

    2016-01-01

    The most expedient unit of the human body is its cell. Encapsulated within the cell are many infinitesimal entities and molecules which are protected by a cell membrane. The proteins that are associated with this lipid based bilayer cell membrane are known as membrane proteins and are considered to play a significant role. These membrane proteins exhibit their effect in cellular activities inside and outside of the cell. According to the scientists in pharmaceutical organizations, these membrane proteins perform key task in drug interactions. In this study, a technique is presented that is based on various computationally intelligent methods used for the prediction of membrane protein without the experimental use of mass spectrometry. Statistical moments were used to extract features and furthermore a Multilayer Neural Network was trained using backpropagation for the prediction of membrane proteins. Results show that the proposed technique performs better than existing methodologies.

  2. Biogenesis of inner membrane proteins in Escherichia coli.

    PubMed

    Luirink, Joen; Yu, Zhong; Wagner, Samuel; de Gier, Jan-Willem

    2012-06-01

    The inner membrane proteome of the model organism Escherichia coli is composed of inner membrane proteins, lipoproteins and peripherally attached soluble proteins. Our knowledge of the biogenesis of inner membrane proteins is rapidly increasing. This is in particular true for the early steps of biogenesis - protein targeting to and insertion into the membrane. However, our knowledge of inner membrane protein folding and quality control is still fragmentary. Furthering our knowledge in these areas will bring us closer to understand the biogenesis of individual inner membrane proteins in the context of the biogenesis of the inner membrane proteome of Escherichia coli as a whole. This article is part of a Special Issue entitled: Biogenesis/Assembly of Respiratory Enzyme Complexes.

  3. A Prediction Model for Membrane Proteins Using Moments Based Features

    PubMed Central

    Butt, Ahmad Hassan; Khan, Sher Afzal; Jamil, Hamza; Rasool, Nouman; Khan, Yaser Daanial

    2016-01-01

    The most expedient unit of the human body is its cell. Encapsulated within the cell are many infinitesimal entities and molecules which are protected by a cell membrane. The proteins that are associated with this lipid based bilayer cell membrane are known as membrane proteins and are considered to play a significant role. These membrane proteins exhibit their effect in cellular activities inside and outside of the cell. According to the scientists in pharmaceutical organizations, these membrane proteins perform key task in drug interactions. In this study, a technique is presented that is based on various computationally intelligent methods used for the prediction of membrane protein without the experimental use of mass spectrometry. Statistical moments were used to extract features and furthermore a Multilayer Neural Network was trained using backpropagation for the prediction of membrane proteins. Results show that the proposed technique performs better than existing methodologies. PMID:26966690

  4. Hydrophobic mismatch sorts SNARE proteins into distinct membrane domains

    PubMed Central

    Milovanovic, Dragomir; Honigmann, Alf; Koike, Seiichi; Göttfert, Fabian; Pähler, Gesa; Junius, Meike; Müllar, Stefan; Diederichsen, Ulf; Janshoff, Andreas; Grubmüller, Helmut; Risselada, Herre J.; Eggeling, Christian; Hell, Stefan W.; van den Bogaart, Geert; Jahn, Reinhard

    2015-01-01

    The clustering of proteins and lipids in distinct microdomains is emerging as an important principle for the spatial patterning of biological membranes. Such domain formation can be the result of hydrophobic and ionic interactions with membrane lipids as well as of specific protein–protein interactions. Here using plasma membrane-resident SNARE proteins as model, we show that hydrophobic mismatch between the length of transmembrane domains (TMDs) and the thickness of the lipid membrane suffices to induce clustering of proteins. Even when the TMDs differ in length by only a single residue, hydrophobic mismatch can segregate structurally closely homologous membrane proteins in distinct membrane domains. Domain formation is further fine-tuned by interactions with polyanionic phosphoinositides and homo and heterotypic protein interactions. Our findings demonstrate that hydrophobic mismatch contributes to the structural organization of membranes. PMID:25635869

  5. Fingerprinting the macro-organisation of pigment-protein complexes in plant thylakoid membranes in vivo by circular-dichroism spectroscopy.

    PubMed

    Tóth, Tünde N; Rai, Neha; Solymosi, Katalin; Zsiros, Ottó; Schröder, Wolfgang P; Garab, Győző; van Amerongen, Herbert; Horton, Peter; Kovács, László

    2016-09-01

    Macro-organisation of the protein complexes in plant thylakoid membranes plays important roles in the regulation and fine-tuning of photosynthetic activity. These delicate structures might, however, undergo substantial changes during isolating the thylakoid membranes or during sample preparations, e.g., for electron microscopy. Circular-dichroism (CD) spectroscopy is a non-invasive technique which can thus be used on intact samples. Via excitonic and psi-type CD bands, respectively, it carries information on short-range excitonic pigment-pigment interactions and the macro-organisation (chiral macrodomains) of pigment-protein complexes (psi, polymer or salt-induced). In order to obtain more specific information on the origin of the major psi-type CD bands, at around (+)506, (-)674 and (+)690nm, we fingerprinted detached leaves and isolated thylakoid membranes of wild-type and mutant plants and also tested the effects of different environmental conditions in vivo. We show that (i) the chiral macrodomains disassemble upon mild detergent treatments, but not after crosslinking the protein complexes; (ii) in different wild-type leaves of dicotyledonous and monocotyledonous angiosperms the CD features are quite robust, displaying very similar excitonic and psi-type bands, suggesting similar protein composition and (macro-) organisation of photosystem II (PSII) supercomplexes in the grana; (iii) the main positive psi-type bands depend on light-harvesting protein II contents of the membranes; (iv) the (+)506nm band appears only in the presence of PSII-LHCII supercomplexes and does not depend on the xanthophyll composition of the membranes. Hence, CD spectroscopy can be used to detect different macro-domains in the thylakoid membranes with different outer antenna compositions in vivo. PMID:27154055

  6. Fingerprinting the macro-organisation of pigment-protein complexes in plant thylakoid membranes in vivo by circular-dichroism spectroscopy.

    PubMed

    Tóth, Tünde N; Rai, Neha; Solymosi, Katalin; Zsiros, Ottó; Schröder, Wolfgang P; Garab, Győző; van Amerongen, Herbert; Horton, Peter; Kovács, László

    2016-09-01

    Macro-organisation of the protein complexes in plant thylakoid membranes plays important roles in the regulation and fine-tuning of photosynthetic activity. These delicate structures might, however, undergo substantial changes during isolating the thylakoid membranes or during sample preparations, e.g., for electron microscopy. Circular-dichroism (CD) spectroscopy is a non-invasive technique which can thus be used on intact samples. Via excitonic and psi-type CD bands, respectively, it carries information on short-range excitonic pigment-pigment interactions and the macro-organisation (chiral macrodomains) of pigment-protein complexes (psi, polymer or salt-induced). In order to obtain more specific information on the origin of the major psi-type CD bands, at around (+)506, (-)674 and (+)690nm, we fingerprinted detached leaves and isolated thylakoid membranes of wild-type and mutant plants and also tested the effects of different environmental conditions in vivo. We show that (i) the chiral macrodomains disassemble upon mild detergent treatments, but not after crosslinking the protein complexes; (ii) in different wild-type leaves of dicotyledonous and monocotyledonous angiosperms the CD features are quite robust, displaying very similar excitonic and psi-type bands, suggesting similar protein composition and (macro-) organisation of photosystem II (PSII) supercomplexes in the grana; (iii) the main positive psi-type bands depend on light-harvesting protein II contents of the membranes; (iv) the (+)506nm band appears only in the presence of PSII-LHCII supercomplexes and does not depend on the xanthophyll composition of the membranes. Hence, CD spectroscopy can be used to detect different macro-domains in the thylakoid membranes with different outer antenna compositions in vivo.

  7. Light Harvesting as a Simple and Versatile Way to Enhance Brightness of Luminescent Sensors

    PubMed Central

    2009-01-01

    The emissive output of indicator dyes in luminescent sensors can be amplified by the addition of antenna dyes with a higher brightness. The highly concentrated antenna dye molecules absorb the excitation light and transfer the energy to an indicator dye. This harvesting of light makes thin sensor layers (thickness <500 nm) and nanometer sized sensor particles with exceptionally high brightness and compatible with the most powerful LEDs available. The performance of sensor layers of ∼250 nm thickness employing light harvesting was investigated and compared with established sensors. The principle is demonstrated for oxygen and ammonia sensors. An overview of possible application of light harvesting to various reagent mediated optical sensing schemes is given.

  8. Vibronic origin of long-lived coherence in an artificial molecular light harvester

    NASA Astrophysics Data System (ADS)

    Lim, James; Paleček, David; Caycedo-Soler, Felipe; Lincoln, Craig N.; Prior, Javier; von Berlepsch, Hans; Huelga, Susana F.; Plenio, Martin B.; Zigmantas, Donatas; Hauer, Jürgen

    2015-07-01

    Natural and artificial light-harvesting processes have recently gained new interest. Signatures of long-lasting coherence in spectroscopic signals of biological systems have been repeatedly observed, albeit their origin is a matter of ongoing debate, as it is unclear how the loss of coherence due to interaction with the noisy environments in such systems is averted. Here we report experimental and theoretical verification of coherent exciton-vibrational (vibronic) coupling as the origin of long-lasting coherence in an artificial light harvester, a molecular J-aggregate. In this macroscopically aligned tubular system, polarization-controlled 2D spectroscopy delivers an uncongested and specific optical response as an ideal foundation for an in-depth theoretical description. We derive analytical expressions that show under which general conditions vibronic coupling leads to prolonged excited-state coherence.

  9. Nature-inspired light-harvesting liquid crystalline porphyrins for organic photovoltaics

    SciTech Connect

    Li, Lanfang; Kang, Shin-Woong; Harden, John; Sun, Qingjiang; Zhou, Xiaoli; Dai, Liming; Jakli, Antal; Kumar, Satyendra; Li, Quan

    2008-12-22

    A new class of nanoscale light-harvesting discotic liquid crystalline porphyrins, with the same basic structure of the best photoreceptor in nature (chlorophyll), was synthesized. These materials can be exceptionally aligned into a highly ordered architecture in which the columns formed by intermolecular {pi}-{pi} stacking are spontaneously perpendicular to the substrate. The homeotropic alignment, well confirmed by synchrotron X-ray diffraction, could not only provide the most efficient pathway for hole conduction along the columnar axis crossing the device thickness, but also offer the largest area to the incident light for optimized light harvesting. Their preliminary photocurrent generation and photovoltaic performances were also demonstrated. The results provide new and efficient pathways to the development of organic photovoltaics by using homeotropically aligned liquid crystal thin films.

  10. Vibronic origin of long-lived coherence in an artificial molecular light harvester

    PubMed Central

    Lim, James; Paleček, David; Caycedo-Soler, Felipe; Lincoln, Craig N.; Prior, Javier; von Berlepsch, Hans; Huelga, Susana F.; Plenio, Martin B.; Zigmantas, Donatas; Hauer, Jürgen

    2015-01-01

    Natural and artificial light-harvesting processes have recently gained new interest. Signatures of long-lasting coherence in spectroscopic signals of biological systems have been repeatedly observed, albeit their origin is a matter of ongoing debate, as it is unclear how the loss of coherence due to interaction with the noisy environments in such systems is averted. Here we report experimental and theoretical verification of coherent exciton–vibrational (vibronic) coupling as the origin of long-lasting coherence in an artificial light harvester, a molecular J-aggregate. In this macroscopically aligned tubular system, polarization-controlled 2D spectroscopy delivers an uncongested and specific optical response as an ideal foundation for an in-depth theoretical description. We derive analytical expressions that show under which general conditions vibronic coupling leads to prolonged excited-state coherence. PMID:26158602

  11. Vibronic origin of long-lived coherence in an artificial molecular light harvester.

    PubMed

    Lim, James; Paleček, David; Caycedo-Soler, Felipe; Lincoln, Craig N; Prior, Javier; von Berlepsch, Hans; Huelga, Susana F; Plenio, Martin B; Zigmantas, Donatas; Hauer, Jürgen

    2015-07-09

    Natural and artificial light-harvesting processes have recently gained new interest. Signatures of long-lasting coherence in spectroscopic signals of biological systems have been repeatedly observed, albeit their origin is a matter of ongoing debate, as it is unclear how the loss of coherence due to interaction with the noisy environments in such systems is averted. Here we report experimental and theoretical verification of coherent exciton-vibrational (vibronic) coupling as the origin of long-lasting coherence in an artificial light harvester, a molecular J-aggregate. In this macroscopically aligned tubular system, polarization-controlled 2D spectroscopy delivers an uncongested and specific optical response as an ideal foundation for an in-depth theoretical description. We derive analytical expressions that show under which general conditions vibronic coupling leads to prolonged excited-state coherence.

  12. Molecular cloning and characterization of the light-harvesting chlorophyll a/b gene from the pigeon pea (Cajanus cajan).

    PubMed

    Qiao, Guang; Wen, Xiao-Peng; Zhang, Ting

    2015-12-01

    Light-harvesting chlorophyll a/b-binding proteins (LHCB) have been implicated in the stress response. In this study, a gene encoding LHCB in the pigeon pea was cloned and characterized. Based on the sequence of a previously obtained 327 bp Est, a full-length 793 bp cDNA was cloned using the rapid amplification of cDNA ends (RACE) method. It was designated CcLHCB1 and encoded a 262 amino acid protein. The calculated molecular weight of the CcLHCB1 protein was 27.89 kDa, and the theoretical isoelectric point was 5.29. Homology search and sequence multi-alignment demonstrated that the CcLHCB1 protein sequence shared a high identity with LHCB from other plants. Bioinformatics analysis revealed that CcLHCB1 was a hydrophobic protein with three transmembrane domains. By fluorescent quantitative real-time polymerase chain reaction (PCR), CcLHCB1 mRNA transcripts were detectable in different tissues (leaf, stem, and root), with the highest level found in the leaf. The expression of CcLHCB1 mRNA in the leaves was up-regulated by drought stimulation and AM inoculation. Our results provide the basis for a better understanding of the molecular organization of LCHB and might be useful for understanding the interaction between plants and microbes in the future. PMID:26329890

  13. Molecular cloning and characterization of the light-harvesting chlorophyll a/b gene from the pigeon pea (Cajanus cajan).

    PubMed

    Qiao, Guang; Wen, Xiao-Peng; Zhang, Ting

    2015-12-01

    Light-harvesting chlorophyll a/b-binding proteins (LHCB) have been implicated in the stress response. In this study, a gene encoding LHCB in the pigeon pea was cloned and characterized. Based on the sequence of a previously obtained 327 bp Est, a full-length 793 bp cDNA was cloned using the rapid amplification of cDNA ends (RACE) method. It was designated CcLHCB1 and encoded a 262 amino acid protein. The calculated molecular weight of the CcLHCB1 protein was 27.89 kDa, and the theoretical isoelectric point was 5.29. Homology search and sequence multi-alignment demonstrated that the CcLHCB1 protein sequence shared a high identity with LHCB from other plants. Bioinformatics analysis revealed that CcLHCB1 was a hydrophobic protein with three transmembrane domains. By fluorescent quantitative real-time polymerase chain reaction (PCR), CcLHCB1 mRNA transcripts were detectable in different tissues (leaf, stem, and root), with the highest level found in the leaf. The expression of CcLHCB1 mRNA in the leaves was up-regulated by drought stimulation and AM inoculation. Our results provide the basis for a better understanding of the molecular organization of LCHB and might be useful for understanding the interaction between plants and microbes in the future.

  14. Dynamic membrane protein topological switching upon changes in phospholipid environment

    PubMed Central

    Vitrac, Heidi; MacLean, David M.; Jayaraman, Vasanthi; Bogdanov, Mikhail; Dowhan, William

    2015-01-01

    A fundamental objective in membrane biology is to understand and predict how a protein sequence folds and orients in a lipid bilayer. Establishing the principles governing membrane protein folding is central to understanding the molecular basis for membrane proteins that display multiple topologies, the intrinsic dynamic organization of membrane proteins, and membrane protein conformational disorders resulting in disease. We previously established that lactose permease of Escherichia coli displays a mixture of topological conformations and undergoes postassembly bidirectional changes in orientation within the lipid bilayer triggered by a change in membrane phosphatidylethanolamine content, both in vivo and in vitro. However, the physiological implications and mechanism of dynamic structural reorganization of membrane proteins due to changes in lipid environment are limited by the lack of approaches addressing the kinetic parameters of transmembrane protein flipping. In this study, real-time fluorescence spectroscopy was used to determine the rates of protein flipping in the lipid bilayer in both directions and transbilayer flipping of lipids triggered by a change in proteoliposome lipid composition. Our results provide, for the first time to our knowledge, a dynamic picture of these events and demonstrate that membrane protein topological rearrangements in response to lipid modulations occur rapidly following a threshold change in proteoliposome lipid composition. Protein flipping was not accompanied by extensive lipid-dependent unfolding of transmembrane domains. Establishment of lipid bilayer asymmetry was not required but may accelerate the rate of protein flipping. Membrane protein flipping was found to accelerate the rate of transbilayer flipping of lipids. PMID:26512118

  15. Light-Harvesting Nanotubes Formed by Supramolecular Assembly of Aromatic Oligophosphates.

    PubMed

    Bösch, Caroline D; Langenegger, Simon M; Häner, Robert

    2016-08-16

    A 2,7-disubstituted phosphodiester-linked phenanthrene trimer forms tubular structures in aqueous media. Chromophores are arranged in H-aggregates. Incorporation of small quantities of pyrene results in the development of light-harvesting nanotubes in which phenanthrenes act as antenna chromophores and pyrenes as energy acceptors. Energy collection is most efficient after excitation at the phenanthrene H-band. Fluorescence quantum yields up to 23 % are reached in pyrene doped, supramolecular nanotubes. PMID:27375116

  16. Atomistic mechanisms of rapid energy transport in light-harvesting molecules

    NASA Astrophysics Data System (ADS)

    Ohmura, Satoshi; Koga, Shiro; Akai, Ichiro; Shimojo, Fuyuki; Kalia, Rajiv K.; Nakano, Aiichiro; Vashishta, Priya

    2011-03-01

    Synthetic supermolecules such as π-conjugated light-harvesting dendrimers efficiently harvest energy from sunlight, which is of significant importance for the global energy problem. Key to their success is rapid transport of electronic excitation energy from peripheral antennas to photochemical reaction cores, the atomistic mechanisms of which remains elusive. Here, quantum-mechanical molecular dynamics simulation incorporating nonadiabatic electronic transitions reveals the key molecular motion that significantly accelerates the energy transport based on the Dexter mechanism.

  17. Marginally hydrophobic transmembrane α-helices shaping membrane protein folding

    PubMed Central

    De Marothy, Minttu T; Elofsson, Arne

    2015-01-01

    Cells have developed an incredible machinery to facilitate the insertion of membrane proteins into the membrane. While we have a fairly good understanding of the mechanism and determinants of membrane integration, more data is needed to understand the insertion of membrane proteins with more complex insertion and folding pathways. This review will focus on marginally hydrophobic transmembrane helices and their influence on membrane protein folding. These weakly hydrophobic transmembrane segments are by themselves not recognized by the translocon and therefore rely on local sequence context for membrane integration. How can such segments reside within the membrane? We will discuss this in the light of features found in the protein itself as well as the environment it resides in. Several characteristics in proteins have been described to influence the insertion of marginally hydrophobic helices. Additionally, the influence of biological membranes is significant. To begin with, the actual cost for having polar groups within the membrane may not be as high as expected; the presence of proteins in the membrane as well as characteristics of some amino acids may enable a transmembrane helix to harbor a charged residue. The lipid environment has also been shown to directly influence the topology as well as membrane boundaries of transmembrane helices—implying a dynamic relationship between membrane proteins and their environment. PMID:25970811

  18. Dark States in the Light-Harvesting complex 2 Revealed by Two-dimensional Electronic Spectroscopy

    NASA Astrophysics Data System (ADS)

    Ferretti, Marco; Hendrikx, Ruud; Romero, Elisabet; Southall, June; Cogdell, Richard J.; Novoderezhkin, Vladimir I.; Scholes, Gregory D.; van Grondelle, Rienk

    2016-02-01

    Energy transfer and trapping in the light harvesting antennae of purple photosynthetic bacteria is an ultrafast process, which occurs with a quantum efficiency close to unity. However the mechanisms behind this process have not yet been fully understood. Recently it was proposed that low-lying energy dark states, such as charge transfer states and polaron pairs, play an important role in the dynamics and directionality of energy transfer. However, it is difficult to directly detect those states because of their small transition dipole moment and overlap with the B850/B870 exciton bands. Here we present a new experimental approach, which combines the selectivity of two-dimensional electronic spectroscopy with the availability of genetically modified light harvesting complexes, to reveal the presence of those dark states in both the genetically modified and the wild-type light harvesting 2 complexes of Rhodopseudomonas palustris. We suggest that Nature has used the unavoidable charge transfer processes that occur when LH pigments are concentrated to enhance and direct the flow of energy.

  19. Light harvesting in photonic crystals revisited: why do slow photons at the blue edge enhance absorption?

    PubMed

    Deparis, O; Mouchet, S R; Su, B-L

    2015-11-11

    Light harvesting enhancement by slow photons in photonic crystal catalysts or dye-sensitized solar cells is a promising approach for increasing the efficiency of photoreactions. This structural effect is exploited in inverse opal TiO2 photocatalysts by tuning the red edge of the photonic band gap to the TiO2 electronic excitation band edge. In spite of many experimental demonstrations, the slow photon effect is not fully understood yet. In particular, observed enhancement by tuning the blue edge has remained unexplained. Based on rigorous couple wave analysis simulations, we quantify light harvesting enhancement in terms of absorption increase at a specific wavelength (monochromatic UV illumination) or photocurrent increase (solar light illumination), with respect to homogeneous flat slab of equivalent material thickness. We show that the commonly accepted explanation relying on light intensity confinement in high (low) dielectric constant regions at the red (blue) edge is challenged in the case of TiO2 inverse opals because of the sub-wavelength size of the material skeleton. The reason why slow photons at the blue edge are also able to enhance light harvesting is the loose confinement of the field, which leads to significant resonantly enhanced field intensity overlap with the skeleton in both red and blue edge tuning cases, yet with different intensity patterns. PMID:26517229

  20. Dark States in the Light-Harvesting complex 2 Revealed by Two-dimensional Electronic Spectroscopy.

    PubMed

    Ferretti, Marco; Hendrikx, Ruud; Romero, Elisabet; Southall, June; Cogdell, Richard J; Novoderezhkin, Vladimir I; Scholes, Gregory D; van Grondelle, Rienk

    2016-02-09

    Energy transfer and trapping in the light harvesting antennae of purple photosynthetic bacteria is an ultrafast process, which occurs with a quantum efficiency close to unity. However the mechanisms behind this process have not yet been fully understood. Recently it was proposed that low-lying energy dark states, such as charge transfer states and polaron pairs, play an important role in the dynamics and directionality of energy transfer. However, it is difficult to directly detect those states because of their small transition dipole moment and overlap with the B850/B870 exciton bands. Here we present a new experimental approach, which combines the selectivity of two-dimensional electronic spectroscopy with the availability of genetically modified light harvesting complexes, to reveal the presence of those dark states in both the genetically modified and the wild-type light harvesting 2 complexes of Rhodopseudomonas palustris. We suggest that Nature has used the unavoidable charge transfer processes that occur when LH pigments are concentrated to enhance and direct the flow of energy.

  1. Dark States in the Light-Harvesting complex 2 Revealed by Two-dimensional Electronic Spectroscopy

    PubMed Central

    Ferretti, Marco; Hendrikx, Ruud; Romero, Elisabet; Southall, June; Cogdell, Richard J.; Novoderezhkin, Vladimir I.; Scholes, Gregory D.; van Grondelle, Rienk

    2016-01-01

    Energy transfer and trapping in the light harvesting antennae of purple photosynthetic bacteria is an ultrafast process, which occurs with a quantum efficiency close to unity. However the mechanisms behind this process have not yet been fully understood. Recently it was proposed that low-lying energy dark states, such as charge transfer states and polaron pairs, play an important role in the dynamics and directionality of energy transfer. However, it is difficult to directly detect those states because of their small transition dipole moment and overlap with the B850/B870 exciton bands. Here we present a new experimental approach, which combines the selectivity of two-dimensional electronic spectroscopy with the availability of genetically modified light harvesting complexes, to reveal the presence of those dark states in both the genetically modified and the wild-type light harvesting 2 complexes of Rhodopseudomonas palustris. We suggest that Nature has used the unavoidable charge transfer processes that occur when LH pigments are concentrated to enhance and direct the flow of energy. PMID:26857477

  2. Dark States in the Light-Harvesting complex 2 Revealed by Two-dimensional Electronic Spectroscopy

    DOE PAGES

    Ferretti, Marco; Hendrikx, Ruud; Romero, Elisabet; Southall, June; Cogdell, Richard J.; Novoderezhkin, Vladimir I.; Scholes, Gregory D.; van Grondelle, Rienk

    2016-02-09

    Energy transfer and trapping in the light harvesting antennae of purple photosynthetic bacteria is an ultrafast process, which occurs with a quantum efficiency close to unity. However the mechanisms behind this process have not yet been fully understood. Recently it was proposed that low-lying energy dark states, such as charge transfer states and polaron pairs, play an important role in the dynamics and directionality of energy transfer. However, it is difficult to directly detect those states because of their small transition dipole moment and overlap with the B850/B870 exciton bands. Here we present a new experimental approach, which combines themore » selectivity of two-dimensional electronic spectroscopy with the availability of genetically modified light harvesting complexes, to reveal the presence of those dark states in both the genetically modified and the wild-type light harvesting 2 complexes of Rhodopseudomonas palustris. In conclusion, we suggest that Nature has used the unavoidable charge transfer processes that occur when LH pigments are concentrated to enhance and direct the flow of energy.« less

  3. Directed assembly of hierarchical light-harvesting complexes using virus capsid scaffolds and DNA origami tiles

    NASA Astrophysics Data System (ADS)

    Wang, Debin; Capehart, Stacy; Pal, Suchetan; Liu, Minghui; Lau, Jolene; Yan, Hao; Francis, Matthew; Deyoreo, Jim; LBNL Team; UCB Team; ASU Team

    2013-03-01

    Directed assembly of nanostructures with molecular precision is of great importance to develop an insightful understanding of assembly pathways and dynamics as well as to derive new functionalities. In this work, we explore the use of virus capsids and DNA origami tiles as 3D scaffolds and 2D templates for directed assembly of light-harvesting molecules and plasmonic gold nanoparticles to achieve tunable photoemission. Bacteriophage MS2 virus capsids with well-defined spherical macromolecular structures are genetically modified to provide predictable steric arrangements of light-harvesting molecules. DNA origami tiles act as programmable planar templates to provide higher-order organization of oligonucleotide-functionalized light-harvesting capsids and plasmonic gold nanoparticles. The direct observation of distance dependent photoluminescence emission is carried out by our correlative approach combining atomic force microscopy and confocal fluorescence microscopy, which is in good agreement with our numerical simulation and theoretical calculation. This work will facilitate the construction of multicomponent biological-metal hybrid plasmonic nanostructures for nanophotonics and biosensing applications.

  4. Light harvesting in photonic crystals revisited: why do slow photons at the blue edge enhance absorption?

    PubMed

    Deparis, O; Mouchet, S R; Su, B-L

    2015-11-11

    Light harvesting enhancement by slow photons in photonic crystal catalysts or dye-sensitized solar cells is a promising approach for increasing the efficiency of photoreactions. This structural effect is exploited in inverse opal TiO2 photocatalysts by tuning the red edge of the photonic band gap to the TiO2 electronic excitation band edge. In spite of many experimental demonstrations, the slow photon effect is not fully understood yet. In particular, observed enhancement by tuning the blue edge has remained unexplained. Based on rigorous couple wave analysis simulations, we quantify light harvesting enhancement in terms of absorption increase at a specific wavelength (monochromatic UV illumination) or photocurrent increase (solar light illumination), with respect to homogeneous flat slab of equivalent material thickness. We show that the commonly accepted explanation relying on light intensity confinement in high (low) dielectric constant regions at the red (blue) edge is challenged in the case of TiO2 inverse opals because of the sub-wavelength size of the material skeleton. The reason why slow photons at the blue edge are also able to enhance light harvesting is the loose confinement of the field, which leads to significant resonantly enhanced field intensity overlap with the skeleton in both red and blue edge tuning cases, yet with different intensity patterns.

  5. Adaptable Lipid Matrix Promotes Protein-Protein Association in Membranes.

    PubMed

    Kuznetsov, Andrey S; Polyansky, Anton A; Fleck, Markus; Volynsky, Pavel E; Efremov, Roman G

    2015-09-01

    The cell membrane is "stuffed" with proteins, whose transmembrane (TM) helical domains spontaneously associate to form functionally active complexes. For a number of membrane receptors, a modulation of TM domains' oligomerization has been shown to contribute to the development of severe pathological states, thus calling for detailed studies of the atomistic aspects of the process. Despite considerable progress achieved so far, several crucial questions still remain: How do the helices recognize each other in the membrane? What is the driving force of their association? Here, we assess the dimerization free energy of TM helices along with a careful consideration of the interplay between the structure and dynamics of protein and lipids using atomistic molecular dynamics simulations in the hydrated lipid bilayer for three different model systems - TM fragments of glycophorin A, polyalanine and polyleucine peptides. We observe that the membrane driven association of TM helices exhibits a prominent entropic character, which depends on the peptide sequence. Thus, a single TM peptide of a given composition induces strong and characteristic perturbations in the hydrophobic core of the bilayer, which may facilitate the initial "communication" between TM helices even at the distances of 20-30 Å. Upon tight helix-helix association, the immobilized lipids accommodate near the peripheral surfaces of the dimer, thus disturbing the packing of the surrounding. The dimerization free energy of the modeled peptides corresponds to the strength of their interactions with lipids inside the membrane being the lowest for glycophorin A and similarly higher for both homopolymers. We propose that the ability to accommodate lipid tails determines the dimerization strength of TM peptides and that the lipid matrix directly governs their association. PMID:26575933

  6. Vertebrate Membrane Proteins: Structure, Function, and Insights from Biophysical Approaches

    PubMed Central

    MÜLLER, DANIEL J.; WU, NAN; PALCZEWSKI, KRZYSZTOF

    2008-01-01

    Membrane proteins are key targets for pharmacological intervention because they are vital for cellular function. Here, we analyze recent progress made in the understanding of the structure and function of membrane proteins with a focus on rhodopsin and development of atomic force microscopy techniques to study biological membranes. Membrane proteins are compartmentalized to carry out extra- and intracellular processes. Biological membranes are densely populated with membrane proteins that occupy approximately 50% of their volume. In most cases membranes contain lipid rafts, protein patches, or paracrystalline formations that lack the higher-order symmetry that would allow them to be characterized by diffraction methods. Despite many technical difficulties, several crystal structures of membrane proteins that illustrate their internal structural organization have been determined. Moreover, high-resolution atomic force microscopy, near-field scanning optical microscopy, and other lower resolution techniques have been used to investigate these structures. Single-molecule force spectroscopy tracks interactions that stabilize membrane proteins and those that switch their functional state; this spectroscopy can be applied to locate a ligand-binding site. Recent development of this technique also reveals the energy landscape of a membrane protein, defining its folding, reaction pathways, and kinetics. Future development and application of novel approaches during the coming years should provide even greater insights to the understanding of biological membrane organization and function. PMID:18321962

  7. Systematically Ranking the Tightness of Membrane Association for Peripheral Membrane Proteins (PMPs)*

    PubMed Central

    Gao, Liyan; Ge, Haitao; Huang, Xiahe; Liu, Kehui; Zhang, Yuanya; Xu, Wu; Wang, Yingchun

    2015-01-01

    Large-scale quantitative evaluation of the tightness of membrane association for nontransmembrane proteins is important for identifying true peripheral membrane proteins with functional significance. Herein, we simultaneously ranked more than 1000 proteins of the photosynthetic model organism Synechocystis sp. PCC 6803 for their relative tightness of membrane association using a proteomic approach. Using multiple precisely ranked and experimentally verified peripheral subunits of photosynthetic protein complexes as the landmarks, we found that proteins involved in two-component signal transduction systems and transporters are overall tightly associated with the membranes, whereas the associations of ribosomal proteins are much weaker. Moreover, we found that hypothetical proteins containing the same domains generally have similar tightness. This work provided a global view of the structural organization of the membrane proteome with respect to divergent functions, and built the foundation for future investigation of the dynamic membrane proteome reorganization in response to different environmental or internal stimuli. PMID:25505158

  8. Bio-Photoelectrochemical Solar Cells Incorporating Reaction Center and Reaction Center Plus Light Harvesting Complexes

    NASA Astrophysics Data System (ADS)

    Yaghoubi, Houman

    onto Au electrodes via surface exposed cysteine residues. This resulted in photocurrent densities as large as ~600 nA cm-2 while still the incident photon to generated electron quantum efficiency was as low as %3 x 10-4. 2- The second approach is to immobilize wild type RCs of Rhodobacter sphaeroides on the surface of a Au underlying electrode using self-assembled monolayers of carboxylic acid terminated oligomers and cytochrome c charge mediating layers, with a preferential orientation from the primary electron donor site. This approach resulted in EQE of up to 0.06%, which showed 200 times efficiency improvement comparing to the first approach. In the third approach, instead of isolated protein complexes, RCs plus light harvesting (LH) complexes were employed for a better photon absorption. Direct attachment of RC-LH1 complexes on Au working electrodes, resulted in 0.21% EQE which showed 3.5 times efficiency improvement over the second approach (700 times higher than the first approach). The main impact of this work is the harnessing of biological RCs for efficient energy harvesting in man-made structures. Specifically, the results in this work will advance the application of RCs in devices for energy harvesting and will enable a better understanding of bio and nanomaterial interfaces, thereby advancing the application of biological materials in electronic devices. At the end, this work offers general guidelines that can serve to improve the performance of bio-hybrid solar cells.

  9. A novel lipoprotein nanoparticle system for membrane proteins

    PubMed Central

    Frauenfeld, Jens; Löving, Robin; Armache, Jean-Paul; Sonnen, Andreas; Guettou, Fatma; Moberg, Per; Zhu, Lin; Jegerschöld, Caroline; Flayhan, Ali; Briggs, John A.G.; Garoff, Henrik; Löw, Christian; Cheng, Yifan; Nordlund, Pär

    2016-01-01

    Membrane proteins are of outstanding importance in biology, drug discovery and vaccination. A common limiting factor in research and applications involving membrane proteins is the ability to solubilize and stabilize membrane proteins. Although detergents represent the major means for solubilizing membrane proteins, they are often associated with protein instability and poor applicability in structural and biophysical studies. Here, we present a novel lipoprotein nanoparticle system that allows for the reconstitution of membrane proteins into a lipid environment that is stabilized by a scaffold of Saposin proteins. We showcase the applicability of the method on two purified membrane protein complexes as well as the direct solubilization and nanoparticle-incorporation of a viral membrane protein complex from the virus membrane. We also demonstrate that this lipid nanoparticle methodology facilitates high-resolution structural studies of membrane proteins in a lipid environment by single-particle electron cryo-microscopy (cryo-EM) and allows for the stabilization of the HIV-envelope glycoprotein in a functional state. PMID:26950744

  10. Water-dispersible nanospheres of hydrogen-bonded supramolecular polymers and their application for mimicking light-harvesting systems.

    PubMed

    Peng, Hui-Qing; Xu, Jiang-Fei; Chen, Yu-Zhe; Wu, Li-Zhu; Tung, Chen-Ho; Yang, Qing-Zheng

    2014-02-01

    Water-dispersible nanospheres of hydrogen-bonded supramolecular polymers have been prepared for the first time by using the miniemulsion method. Nanospheres containing chromophores with high fluorescence quantum yields were fabricated to mimic the natural light-harvesting system.

  11. Hierarchical self-assembly of a biomimetic light-harvesting antenna based on DNA G-quadruplexes.

    PubMed

    Sancho Oltra, Núria; Browne, Wesley R; Roelfes, Gerard

    2013-02-11

    A new modular approach to an artificial light-harvesting antenna system is presented. The approach involves the hierarchical self-assembly of porphyrin acceptor molecules to G-quadruplexes tethered to coumarin donor moieties.

  12. Chitosan-based membrane chromatography for protein adsorption and separation.

    PubMed

    Liu, Yezhuo; Feng, Zhicheng; Shao, Zhengzhong; Chen, Xin

    2012-08-01

    A chitosan-based membrane chromatography was set up by using natural chitosan/carboxymethylchitosan (CS/CMCS) blend membrane as the matrix. The dynamic adsorption property for protein (lysozyme as model protein) was detailed discussed with the change in pore size of the membrane, the flow rate and the initial concentration of the feed solution, and the layer of membrane in membrane stack. The best dynamic adsorption capacity of lysozyme on the CS/CMCS membrane chromatography was found to be 15.3mg/mL under the optimal flow conditions. Moreover, the CS/CMCS membrane chromatography exhibited good repeatability and reusability with the desorption efficiency of ~90%. As an application, lysozyme and ovalbumin were successfully separated from their binary mixture through the CS/CMCS membrane chromatography. This implies that such a natural chitosan-based membrane chromatography may have great potential on the bioseparation field in the future.

  13. An Integrated Framework Advancing Membrane Protein Modeling and Design

    PubMed Central

    Weitzner, Brian D.; Duran, Amanda M.; Tilley, Drew C.; Elazar, Assaf; Gray, Jeffrey J.

    2015-01-01

    Membrane proteins are critical functional molecules in the human body, constituting more than 30% of open reading frames in the human genome. Unfortunately, a myriad of difficulties in overexpression and reconstitution into membrane mimetics severely limit our ability to determine their structures. Computational tools are therefore instrumental to membrane protein structure prediction, consequently increasing our understanding of membrane protein function and their role in disease. Here, we describe a general framework facilitating membrane protein modeling and design that combines the scientific principles for membrane protein modeling with the flexible software architecture of Rosetta3. This new framework, called RosettaMP, provides a general membrane representation that interfaces with scoring, conformational sampling, and mutation routines that can be easily combined to create new protocols. To demonstrate the capabilities of this implementation, we developed four proof-of-concept applications for (1) prediction of free energy changes upon mutation; (2) high-resolution structural refinement; (3) protein-protein docking; and (4) assembly of symmetric protein complexes, all in the membrane environment. Preliminary data show that these algorithms can produce meaningful scores and structures. The data also suggest needed improvements to both sampling routines and score functions. Importantly, the applications collectively demonstrate the potential of combining the flexible nature of RosettaMP with the power of Rosetta algorithms to facilitate membrane protein modeling and design. PMID:26325167

  14. An Integrated Framework Advancing Membrane Protein Modeling and Design.

    PubMed

    Alford, Rebecca F; Koehler Leman, Julia; Weitzner, Brian D; Duran, Amanda M; Tilley, Drew C; Elazar, Assaf; Gray, Jeffrey J

    2015-09-01

    Membrane proteins are critical functional molecules in the human body, constituting more than 30% of open reading frames in the human genome. Unfortunately, a myriad of difficulties in overexpression and reconstitution into membrane mimetics severely limit our ability to determine their structures. Computational tools are therefore instrumental to membrane protein structure prediction, consequently increasing our understanding of membrane protein function and their role in disease. Here, we describe a general framework facilitating membrane protein modeling and design that combines the scientific principles for membrane protein modeling with the flexible software architecture of Rosetta3. This new framework, called RosettaMP, provides a general membrane representation that interfaces with scoring, conformational sampling, and mutation routines that can be easily combined to create new protocols. To demonstrate the capabilities of this implementation, we developed four proof-of-concept applications for (1) prediction of free energy changes upon mutation; (2) high-resolution structural refinement; (3) protein-protein docking; and (4) assembly of symmetric protein complexes, all in the membrane environment. Preliminary data show that these algorithms can produce meaningful scores and structures. The data also suggest needed improvements to both sampling routines and score functions. Importantly, the applications collectively demonstrate the potential of combining the flexible nature of RosettaMP with the power of Rosetta algorithms to facilitate membrane protein modeling and design. PMID:26325167

  15. Membrane interacting regions of Dengue virus NS2A protein.

    PubMed

    Nemésio, Henrique; Villalaín, José

    2014-08-28

    The Dengue virus (DENV) NS2A protein, essential for viral replication, is a poorly characterized membrane protein. NS2A displays both protein/protein and membrane/protein interactions, yet neither its functions in the viral cycle nor its active regions are known with certainty. To highlight the different membrane-active regions of NS2A, we characterized the effects of peptides derived from a peptide library encompassing this protein's full length on different membranes by measuring their membrane leakage induction and modulation of lipid phase behavior. Following this initial screening, one region, peptide dens25, had interesting effects on membranes; therefore, we sought to thoroughly characterize this region's interaction with membranes. This peptide presents an interfacial/hydrophobic pattern characteristic of a membrane-proximal segment. We show that dens25 strongly interacts with membranes that contain a large proportion of lipid molecules with a formal negative charge, and that this effect has a major electrostatic contribution. Considering its membrane modulating capabilities, this region might be involved in membrane rearrangements and thus be important for the viral cycle.

  16. A topological and conformational stability alphabet for multipass membrane proteins.

    PubMed

    Feng, Xiang; Barth, Patrick

    2016-03-01

    Multipass membrane proteins perform critical signal transduction and transport across membranes. How transmembrane helix (TMH) sequences encode the topology and conformational flexibility regulating these functions remains poorly understood. Here we describe a comprehensive analysis of the sequence-structure relationships at multiple interacting TMHs from all membrane proteins with structures in the Protein Data Bank (PDB). We found that membrane proteins can be deconstructed in interacting TMH trimer units, which mostly fold into six distinct structural classes of topologies and conformations. Each class is enriched in recurrent sequence motifs from functionally unrelated proteins, revealing unforeseen consensus and evolutionary conserved networks of stabilizing interhelical contacts. Interacting TMHs' topology and local protein conformational flexibility were remarkably well predicted in a blinded fashion from the identified binding-hotspot motifs. Our results reveal universal sequence-structure principles governing the complex anatomy and plasticity of multipass membrane proteins that may guide de novo structure prediction, design, and studies of folding and dynamics. PMID:26780406

  17. Reversible Switching between Nonquenched and Quenched States in Nanoscale Linear Arrays of Plant Light-Harvesting Antenna Complexes

    PubMed Central

    2014-01-01

    A simple and robust nanolithographic method that allows sub-100 nm chemical patterning on a range of oxide surfaces was developed in order to fabricate nanoarrays of plant light-harvesting LHCII complexes. The site-specific immobilization and the preserved functionality of the LHCII complexes were confirmed by fluorescence emission spectroscopy. Nanopatterned LHCII trimers could be reversibly switched between fluorescent and quenched states by controlling the detergent concentration in the imaging buffer. A 3-fold quenching of the average fluorescence intensity was accompanied by a decrease in the average (amplitude-weighted) fluorescence lifetime from approximately 2.24 ns to approximately 0.4 ns, attributed to the intrinsic ability of LHCII to switch between fluorescent and quenched states upon changes in its conformational state. The nanopatterning methodology was extended by immobilizing a second protein, the enhanced green fluorescent protein (EGFP), onto LHCII-free areas of the chemically patterned surfaces. This very simple surface chemistry, which allows simultaneous selective immobilization and therefore sorting of the two types of protein molecules on the surface, is a key underpinning step toward the integration of LHCII into switchable biohybrid antenna constructs. PMID:24988144

  18. Derivation of coarse-grained simulation models of chlorophyll molecules in lipid bilayers for applications in light harvesting systems.

    PubMed

    Debnath, Ananya; Wiegand, Sabine; Paulsen, Harald; Kremer, Kurt; Peter, Christine

    2015-09-14

    The correct interplay of interactions between protein, pigment and lipid molecules is highly relevant for our understanding of the association behavior of the light harvesting complex (LHCII) of green plants. To cover the relevant time and length scales in this multicomponent system, a multi-scale simulation ansatz is employed that subsequently uses a classical all atomistic (AA) model to derive a suitable coarse grained (CG) model which can be backmapped into the AA resolution, aiming for a seamless conversion between two scales. Such an approach requires a faithful description of not only the protein and lipid components, but also the interaction functions for the indispensable pigment molecules, chlorophyll b and chlorophyll a (referred to as chl b/chl a). In this paper we develop a CG model for chl b and chl a in a dipalmitoylphosphatidyl choline (DPPC) bilayer system. The structural properties and the distribution behavior of chl within the lipid bilayer in the CG simulations are consistent with those of AA reference simulations. The non-bonded potentials are parameterized such that they fit to the thermodynamics based MARTINI force-field for the lipid bilayer and the protein. The CG simulation shows chl aggregation in the lipid bilayer which is supported by fluorescence quenching experiments. It is shown that the derived chl model is well suited for CG simulations of stable, structurally consistent, trimeric LHCII and can in the future be used to study their large scale aggregation behavior.

  19. Reshaping biological membranes in endocytosis: crossing the configurational space of membrane-protein interactions.

    PubMed

    Simunovic, Mijo; Bassereau, Patricia

    2014-03-01

    Lipid membranes are highly dynamic. Over several decades, physicists and biologists have uncovered a number of ways they can change the shape of membranes or alter their phase behavior. In cells, the intricate action of membrane proteins drives these processes. Considering the highly complex ways proteins interact with biological membranes, molecular mechanisms of membrane remodeling still remain unclear. When studying membrane remodeling phenomena, researchers often observe different results, leading them to disparate conclusions on the physiological course of such processes. Here we discuss how combining research methodologies and various experimental conditions contributes to the understanding of the entire phase space of membrane-protein interactions. Using the example of clathrin-mediated endocytosis we try to distinguish the question 'how can proteins remodel the membrane?' from 'how do proteins remodel the membrane in the cell?' In particular, we consider how altering physical parameters may affect the way membrane is remodeled. Uncovering the full range of physical conditions under which membrane phenomena take place is key in understanding the way cells take advantage of membrane properties in carrying out their vital tasks.

  20. The Use of Detergents to Purify Membrane Proteins.

    PubMed

    Orwick-Rydmark, Marcella; Arnold, Thomas; Linke, Dirk

    2016-04-01

    Extraction of membrane proteins from biological membranes is usually accomplished with the help of detergents. This unit describes the use of detergents to solubilize and purify membrane proteins. The chemical and physical properties of the different classes of detergents typically used with biological samples are discussed. A separate section addresses the compatibility of detergents with applications downstream of the membrane protein purification process, such as optical spectroscopy, mass spectrometry, protein crystallography, biomolecular NMR, or electron microscopy. A brief summary of alternative membrane protein solubilizing and stabilizing systems is also included. Protocols in this unit include the isolation and solubilization of biological membranes and phase separation; support protocols for detergent removal, detergent exchange, and the determination of critical micelle concentration using different methods are also included.

  1. Detergent-Specific Membrane Protein Crystallization Screens

    NASA Technical Reports Server (NTRS)

    Wiener, Michael

    2007-01-01

    A suite of reagents has been developed for three-dimensional crystallization of integral membranes present in solution as protein-detergent complexes (PDCs). The compositions of these reagents have been determined in part by proximity to the phase boundaries (lower consolute boundaries) of the detergents present in the PDCs. The acquisition of some of the requisite phase-boundary data and the preliminary design of several of the detergent- specific screens was supported by a NASA contract. At the time of expiration of the contract, a partial set of preliminary screens had been developed. This work has since been extended under non-NASA sponsorship, leading to near completion of a set of 20 to 30 different and unique detergent- specific 96-condition screens.

  2. Direct interaction of the major light-harvesting complex II and PsbS in nonphotochemical quenching.

    PubMed

    Wilk, Laura; Grunwald, Matthias; Liao, Pen-Nan; Walla, Peter Jomo; Kühlbrandt, Werner

    2013-04-01

    The photosystem II (PSII) subunit S (PsbS) plays a key role in nonphotochemical quenching, a photoprotective mechanism for dissipation of excess excitation energy in plants. The precise function of PsbS in nonphotochemical quenching is unknown. By reconstituting PsbS together with the major light-harvesting complex of PSII (LHC-II) and the xanthophyll zeaxanthin (Zea) into proteoliposomes, we have tested the individual contributions of PSII complexes and Zea to chlorophyll (Chl) fluorescence quenching in a membrane environment. We demonstrate that PsbS is stable in the absence of pigments in vitro. Significant Chl fluorescence quenching of reconstituted LHC-II was observed in the presence of PsbS and Zea, although neither Zea nor PsbS alone was sufficient to induce the same quenching. Coreconstitution with PsbS resulted in the formation of LHC-II/PsbS heterodimers, indicating their direct interaction in the lipid bilayer. Two-photon excitation measurements on liposomes containing LHC-II, PsbS, and Zea showed an increase of electronic interactions between carotenoid S1 and Chl states, Φ(Coupling)(CarS1-Chl), that correlated directly with Chl fluorescence quenching. These findings are in agreement with a carotenoid-dependent Chl fluorescence quenching by direct interactions of LHCs of PSII with PsbS monomers.

  3. Bilayer-thickness-mediated interactions between integral membrane proteins.

    PubMed

    Kahraman, Osman; Koch, Peter D; Klug, William S; Haselwandter, Christoph A

    2016-04-01

    Hydrophobic thickness mismatch between integral membrane proteins and the surrounding lipid bilayer can produce lipid bilayer thickness deformations. Experiment and theory have shown that protein-induced lipid bilayer thickness deformations can yield energetically favorable bilayer-mediated interactions between integral membrane proteins, and large-scale organization of integral membrane proteins into protein clusters in cell membranes. Within the continuum elasticity theory of membranes, the energy cost of protein-induced bilayer thickness deformations can be captured by considering compression and expansion of the bilayer hydrophobic core, membrane tension, and bilayer bending, resulting in biharmonic equilibrium equations describing the shape of lipid bilayers for a given set of bilayer-protein boundary conditions. Here we develop a combined analytic and numerical methodology for the solution of the equilibrium elastic equations associated with protein-induced lipid bilayer deformations. Our methodology allows accurate prediction of thickness-mediated protein interactions for arbitrary protein symmetries at arbitrary protein separations and relative orientations. We provide exact analytic solutions for cylindrical integral membrane proteins with constant and varying hydrophobic thickness, and develop perturbative analytic solutions for noncylindrical protein shapes. We complement these analytic solutions, and assess their accuracy, by developing both finite element and finite difference numerical solution schemes. We provide error estimates of our numerical solution schemes and systematically assess their convergence properties. Taken together, the work presented here puts into place an analytic and numerical framework which allows calculation of bilayer-mediated elastic interactions between integral membrane proteins for the complicated protein shapes suggested by structural biology and at the small protein separations most relevant for the crowded membrane

  4. Discriminating lysosomal membrane protein types using dynamic neural network.

    PubMed

    Tripathi, Vijay; Gupta, Dwijendra Kumar

    2014-01-01

    This work presents a dynamic artificial neural network methodology, which classifies the proteins into their classes from their sequences alone: the lysosomal membrane protein classes and the various other membranes protein classes. In this paper, neural networks-based lysosomal-associated membrane protein type prediction system is proposed. Different protein sequence representations are fused to extract the features of a protein sequence, which includes seven feature sets; amino acid (AA) composition, sequence length, hydrophobic group, electronic group, sum of hydrophobicity, R-group, and dipeptide composition. To reduce the dimensionality of the large feature vector, we applied the principal component analysis. The probabilistic neural network, generalized regression neural network, and Elman regression neural network (RNN) are used as classifiers and compared with layer recurrent network (LRN), a dynamic network. The dynamic networks have memory, i.e. its output depends not only on the input but the previous outputs also. Thus, the accuracy of LRN classifier among all other artificial neural networks comes out to be the highest. The overall accuracy of jackknife cross-validation is 93.2% for the data-set. These predicted results suggest that the method can be effectively applied to discriminate lysosomal associated membrane proteins from other membrane proteins (Type-I, Outer membrane proteins, GPI-Anchored) and Globular proteins, and it also indicates that the protein sequence representation can better reflect the core feature of membrane proteins than the classical AA composition.

  5. Bacteriophage membrane protein P9 as a fusion partner for the efficient expression of membrane proteins in Escherichia coli.

    PubMed

    Jung, Yuna; Jung, Hyeim; Lim, Dongbin

    2015-12-01

    Despite their important roles and economic values, studies of membrane proteins have been hampered by the difficulties associated with obtaining sufficient amounts of protein. Here, we report a novel membrane protein expression system that uses the major envelope protein (P9) of phage φ6 as an N-terminal fusion partner. Phage membrane protein P9 facilitated the synthesis of target proteins and their integration into the Escherichia coli cell membrane. This system was used to produce various multi-pass transmembrane proteins, including G-protein-coupled receptors, transporters, and ion channels of human origin. Green fluorescent protein fusion was used to confirm the correct folding of the expressed proteins. Of the 14 membrane proteins tested, eight were highly expressed, three were moderately expressed, and three were barely expressed in E. coli. Seven of the eight highly expressed proteins could be purified after extraction with the mild detergent lauryldimethylamine-oxide. Although a few proteins have previously been developed as fusion partners to augment membrane protein production, we believe that the major envelope protein P9 described here is better suited to the efficient expression of eukaryotic transmembrane proteins in E. coli.

  6. Intermolecular detergent-membrane protein noes for the characterization of the dynamics of membrane protein-detergent complexes.

    PubMed

    Eichmann, Cédric; Orts, Julien; Tzitzilonis, Christos; Vögeli, Beat; Smrt, Sean; Lorieau, Justin; Riek, Roland

    2014-12-11

    The interaction between membrane proteins and lipids or lipid mimetics such as detergents is key for the three-dimensional structure and dynamics of membrane proteins. In NMR-based structural studies of membrane proteins, qualitative analysis of intermolecular nuclear Overhauser enhancements (NOEs) or paramagnetic resonance enhancement are used in general to identify the transmembrane segments of a membrane protein. Here, we employed a quantitative characterization of intermolecular NOEs between (1)H of the detergent and (1)H(N) of (2)H-perdeuterated, (15)N-labeled α-helical membrane protein-detergent complexes following the exact NOE (eNOE) approach. Structural considerations suggest that these intermolecular NOEs should show a helical-wheel-type behavior along a transmembrane helix or a membrane-attached helix within a membrane protein as experimentally demonstrated for the complete influenza hemagglutinin fusion domain HAfp23. The partial absence of such a NOE pattern along the amino acid sequence as shown for a truncated variant of HAfp23 and for the Escherichia coli inner membrane protein YidH indicates the presence of large tertiary structure fluctuations such as an opening between helices or the presence of large rotational dynamics of the helices. Detergent-protein NOEs thus appear to be a straightforward probe for a qualitative characterization of structural and dynamical properties of membrane proteins embedded in detergent micelles.

  7. Membrane Interacting Regions of Dengue Virus NS2A Protein

    PubMed Central

    2015-01-01

    The Dengue virus (DENV) NS2A protein, essential for viral replication, is a poorly characterized membrane protein. NS2A displays both protein/protein and membrane/protein interactions, yet neither its functions in the viral cycle nor its active regions are known with certainty. To highlight the different membrane-active regions of NS2A, we characterized the effects of peptides derived from a peptide library encompassing this protein’s full length on different membranes by measuring their membrane leakage induction and modulation of lipid phase behavior. Following this initial screening, one region, peptide dens25, had interesting effects on membranes; therefore, we sought to thoroughly characterize this region’s interaction with membranes. This peptide presents an interfacial/hydrophobic pattern characteristic of a membrane-proximal segment. We show that dens25 strongly interacts with membranes that contain a large proportion of lipid molecules with a formal negative charge, and that this effect has a major electrostatic contribution. Considering its membrane modulating capabilities, this region might be involved in membrane rearrangements and thus be important for the viral cycle. PMID:25119664

  8. Toward understanding driving forces in membrane protein folding.

    PubMed

    Hong, Heedeok

    2014-12-15

    α-Helical membrane proteins are largely composed of nonpolar residues that are embedded in the lipid bilayer. An enigma in the folding of membrane proteins is how a polypeptide chain can be condensed into the compact folded state in the environment where the hydrophobic effect cannot strongly drive molecular interactions. Probably other forces such as van der Waals packing, hydrogen bonding, and weakly polar interactions, which are regarded less important in the folding of water-soluble proteins, should emerge. However, it is not clearly understood how those individual forces operate and how they are balanced for stabilizing membrane proteins. Studying this problem is not a trivial task mainly because of the methodological challenges in controlling the reversible folding of membrane proteins in the lipid bilayer. Overcoming the hurdles, meaningful progress has been made in the field in the last few decades. This review will focus on recent studies tackling the problem of driving forces in membrane protein folding. PMID:25107533

  9. Visualizing active membrane protein complexes by electron cryotomography

    PubMed Central

    Gold, Vicki A.M.; Ieva, Raffaele; Walter, Andreas; Pfanner, Nikolaus; van der Laan, Martin; Kühlbrandt, Werner

    2014-01-01

    Unravelling the structural organization of membrane protein machines in their active state and native lipid environment is a major challenge in modern cell biology research. Here we develop the STAMP (Specifically TArgeted Membrane nanoParticle) technique as a strategy to localize protein complexes in situ by electron cryotomography (cryo-ET). STAMP selects active membrane protein complexes and marks them with quantum dots. Taking advantage of new electron detector technology that is currently revolutionizing cryotomography in terms of achievable resolution, this approach enables us to visualize the three-dimensional distribution and organization of protein import sites in mitochondria. We show that import sites cluster together in the vicinity of crista membranes, and we reveal unique details of the mitochondrial protein import machinery in action. STAMP can be used as a tool for site-specific labelling of a multitude of membrane proteins by cryo-ET in the future. PMID:24942077

  10. Expression strategies for structural studies of eukaryotic membrane proteins.

    PubMed

    Lyons, Joseph A; Shahsavar, Azadeh; Paulsen, Peter Aasted; Pedersen, Bjørn Panyella; Nissen, Poul

    2016-06-01

    Integral membrane proteins in eukaryotes are central to various cellular processes and key targets in structural biology, biotechnology and drug development. However, the number of available structures for eukaryotic membrane protein belies their physiological importance. Recently, the number of available eukaryotic membrane protein structures has been steadily increasing due to the development of novel strategies in construct design, expression and structure determination. Here, we examine the major expression systems exploited for eukaryotic membrane proteins. Additionally we strive to tabulate and describe the recent expression strategies in eukaryotic membrane protein structural biology. We find that a majority of targets have been expressed in advanced host systems and modified from their wild-type form with distinct focus on conformation and thermostabilisation. However, strategies for native protein purification should also be considered where possible, particularly in light of the recent advances in single particle cryo electron microscopy.

  11. Expression strategies for structural studies of eukaryotic membrane proteins.

    PubMed

    Lyons, Joseph A; Shahsavar, Azadeh; Paulsen, Peter Aasted; Pedersen, Bjørn Panyella; Nissen, Poul

    2016-06-01

    Integral membrane proteins in eukaryotes are central to various cellular processes and key targets in structural biology, biotechnology and drug development. However, the number of available structures for eukaryotic membrane protein belies their physiological importance. Recently, the number of available eukaryotic membrane protein structures has been steadily increasing due to the development of novel strategies in construct design, expression and structure determination. Here, we examine the major expression systems exploited for eukaryotic membrane proteins. Additionally we strive to tabulate and describe the recent expression strategies in eukaryotic membrane protein structural biology. We find that a majority of targets have been expressed in advanced host systems and modified from their wild-type form with distinct focus on conformation and thermostabilisation. However, strategies for native protein purification should also be considered where possible, particularly in light of the recent advances in single particle cryo electron microscopy. PMID:27362979

  12. Organization and Dynamics of Receptor Proteins in a Plasma Membrane.

    PubMed

    Koldsø, Heidi; Sansom, Mark S P

    2015-11-25

    The interactions of membrane proteins are influenced by their lipid environment, with key lipid species able to regulate membrane protein function. Advances in high-resolution microscopy can reveal the organization and dynamics of proteins and lipids within living cells at resolutions <200 nm. Parallel advances in molecular simulations provide near-atomic-resolution models of the dynamics of the organization of membranes of in vivo-like complexity. We explore the dynamics of proteins and lipids in crowded and complex plasma membrane models, thereby closing the gap in length and complexity between computations and experiments. Our simulations provide insights into the mutual interplay between lipids and proteins in determining mesoscale (20-100 nm) fluctuations of the bilayer, and in enabling oligomerization and clustering of membrane proteins.

  13. Size-dependent protein segregation at membrane interfaces

    NASA Astrophysics Data System (ADS)

    Schmid, Eva M.; Bakalar, Matthew H.; Choudhuri, Kaushik; Weichsel, Julian; Ann, Hyoung Sook; Geissler, Phillip L.; Dustin, Michael L.; Fletcher, Daniel A.

    2016-07-01

    Membrane interfaces formed at cell-cell junctions are associated with characteristic patterns of membrane proteins whose organization is critical for intracellular signalling. To isolate the role of membrane protein size in pattern formation, we reconstituted model membrane interfaces in vitro using giant unilamellar vesicles decorated with synthetic binding and non-binding proteins. We show that size differences between membrane proteins can drastically alter their organization at membrane interfaces, with as little as a ~5 nm increase in non-binding protein size driving its exclusion from the interface. Combining in vitro measurements with Monte Carlo simulations, we find that non-binding protein exclusion is also influenced by lateral crowding, binding protein affinity, and thermally driven membrane height fluctuations that transiently limit access to the interface. This sensitive and highly effective means of physically segregating proteins has implications for cell-cell contacts such as T-cell immunological synapses (for example, CD45 exclusion) and epithelial cell junctions (for example, E-cadherin enrichment), as well as for protein sorting at intracellular contact points between membrane-bound organelles.

  14. Membrane protein structures without crystals, by single particle electron cryomicroscopy

    PubMed Central

    Vinothkumar, Kutti R

    2015-01-01

    It is an exciting period in membrane protein structural biology with a number of medically important protein structures determined at a rapid pace. However, two major hurdles still remain in the structural biology of membrane proteins. One is the inability to obtain large amounts of protein for crystallization and the other is the failure to get well-diffracting crystals. With single particle electron cryomicroscopy, both these problems can be overcome and high-resolution structures of membrane proteins and other labile protein complexes can be obtained with very little protein and without the need for crystals. In this review, I highlight recent advances in electron microscopy, detectors and software, which have allowed determination of medium to high-resolution structures of membrane proteins and complexes that have been difficult to study by other structural biological techniques. PMID:26435463

  15. The peripheral myelin protein 22 and epithelial membrane protein family.

    PubMed

    Jetten, A M; Suter, U

    2000-01-01

    The peripheral myelin protein 22 (PMP22) and the epithelial membrane proteins (EMP-1, -2, and -3) comprise a subfamily of small hydrophobic membrane proteins. The putative four-transmembrane domain structure as well as the genomic structure are highly conserved among family members. PMP22 and EMPs are expressed in many tissues, and functions in cell growth, differentiation, and apoptosis have been reported. EMP-1 is highly up-regulated during squamous differentiation and in certain tumors, and a role in tumorigenesis has been proposed. PMP22 is most highly expressed in peripheral nerves, where it is localized in the compact portion of myelin. It plays a crucial role in normal physiological and pathological processes in the peripheral nervous system. Progress in molecular genetics has revealed that genetic alterations in the PMP22 gene, including duplications, deletions, and point mutations, are responsible for several forms of hereditary peripheral neuropathies, including Charcot-Marie-Tooth disease type 1A (CMT1A), Dejerine-Sottas syndrome (DDS), and hereditary neuropathy with liability to pressure palsies (HNPP). The natural mouse mutants Trembler and Trembler-J contain a missense mutation in different hydrophobic domains of PMP22, resulting in demyelination and Schwann cell proliferation. Transgenic mice carrying many copies of the PMP22 gene and PMP22-null mice display a variety of defects in the initial steps of myelination and/or maintenance of myelination, whereas no pathological alterations are detected in other tissues normally expressing PMP22. Further characterization of the interactions of PMP22 and EMPs with other proteins as well as their regulation will provide additional insight into their normal physiological function and their roles in disease and possibly will result in the development of therapeutic tools. PMID:10697408

  16. One-Dimensional Multichromophor Arrays Based on DNA: From Self-Assembly to Light-Harvesting.

    PubMed

    Ensslen, Philipp; Wagenknecht, Hans-Achim

    2015-10-20

    Light-harvesting complexes collect light energy and deliver it by a cascade of energy and electron transfer processes to the reaction center where charge separation leads to storage as chemical energy. The design of artificial light-harvesting assemblies faces enormous challenges because several antenna chromophores need to be kept in close proximity but self-quenching needs to be avoided. Double stranded DNA as a supramolecular scaffold plays a promising role due to its characteristic structural properties. Automated DNA synthesis allows incorporation of artificial chromophore-modified building blocks, and sequence design allows precise control of the distances and orientations between the chromophores. The helical twist between the chromophores, which is induced by the DNA framework, controls energy and electron transfer and thereby reduces the self-quenching that is typically observed in chromophore aggregates. This Account summarizes covalently multichromophore-modified DNA and describes how such multichromophore arrays were achieved by Watson-Crick-specific and DNA-templated self-assembly. The covalent DNA systems were prepared by incorporation of chromophores as DNA base substitutions (either as C-nucleosides or with acyclic linkers as substitutes for the 2'-deoxyribofuranoside) and as DNA base modifications. Studies with DNA base substitutions revealed that distances but more importantly relative orientations of the chromophores govern the energy transfer efficiencies and thereby the light-harvesting properties. With DNA base substitutions, duplex stabilization was faced and could be overcome, for instance, by zipper-like placement of the chromophores in both strands. For both principal structural approaches, DNA-based light-harvesting antenna could be realized. The major disadvantages, however, for covalent multichromophore DNA conjugates are the poor yields of synthesis and the solubility issues for oligonucleotides with more than 5-10 chromophore

  17. Development of photochemical activity in relation to pigment and membrane protein accumulation in chloroplasts of barley and its virescens mutant.

    PubMed

    Kyle, D J; Zalik, S

    1982-06-01

    The development of photochemical activity in relation to pigment and membrane protein accumulation in chloroplasts of greening wild-type barley (Hordeum vulgare L. cv. Gateway) and its virescens mutant were studied. The rate of chlorophyll accumulation per plastid was faster in the wild-type than in the mutant seedlings upon illumination after 6 days of etiolation, but was not different after 8 days. Although the protein content per plastid did not vary during greening, there was a change in the sodium dodecyl sulfate-polyacrylamide gel polypeptide profiles. High molecular weight proteins of 96,000 and 66,000 decreased whereas those at 34,000, 27,000 and 22,000 increased in relative quantity as a function of greening. The fully greened mutant seedlings were not deficient in the light-harvesting chlorophyll protein complex (LHC) or the reaction centers of photosystem I and photosystem II. Photosystem I-associated photochemical activities appeared within the first hour of plastid development and photosystem II associated activities and O(2) evolution within the next 6 hours. In all cases, the developmental rates per unit protein were slower in the mutant following 6 days of etiolation, but no differences between the two genotypes could be seen after 8 days due to a decrease in the developmental rate of the wild-type chloroplasts. An increase in photosynthetic unit size associated with plastid morphogenesis was faster in the wild-type seedlings after 6 days, but again the difference was negligible after 8 days. It was concluded that no single measured photochemical parameter is affected by this mutation, but rather, all aspects of chloroplast development are affected similarly by an overall reduction in the rate of chloroplast morphogenesis. This mutant, therefore, undergoes the normal pattern of proplastid to chloroplast development, but at a markedly reduced rate.

  18. Membrane Protein Mobility and Orientation Preserved in Supported Bilayers Created Directly from Cell Plasma Membrane Blebs.

    PubMed

    Richards, Mark J; Hsia, Chih-Yun; Singh, Rohit R; Haider, Huma; Kumpf, Julia; Kawate, Toshimitsu; Daniel, Susan

    2016-03-29

    Membrane protein interactions with lipids are crucial for their native biological behavior, yet traditional characterization methods are often carried out on purified protein in the absence of lipids. We present a simple method to transfer membrane proteins expressed in mammalian cells to an assay-friendly, cushioned, supported lipid bilayer platform using cell blebs as an intermediate. Cell blebs, expressing either GPI-linked yellow fluorescent proteins or neon-green fused transmembrane P2X2 receptors, were induced to rupture on glass surfaces using PEGylated lipid vesicles, which resulted in planar supported membranes with over 50% mobility for multipass transmembrane proteins and over 90% for GPI-linked proteins. Fluorescent proteins were tracked, and their diffusion in supported bilayers characterized, using single molecule tracking and moment scaling spectrum (MSS) analysis. Diffusion was characterized for individual proteins as either free or confined, revealing details of the local lipid membrane heterogeneity surrounding the protein. A particularly useful result of our bilayer formation process is the protein orientation in the supported planar bilayer. For both the GPI-linked and transmembrane proteins used here, an enzymatic assay revealed that protein orientation in the planar bilayer results in the extracellular domains facing toward the bulk, and that the dominant mode of bleb rupture is via the "parachute" mechanism. Mobility, orientation, and preservation of the native lipid environment of the proteins using cell blebs offers advantages over proteoliposome reconstitution or disrupted cell membrane preparations, which necessarily result in significant scrambling of protein orientation and typically immobilized membrane proteins in SLBs. The bleb-based bilayer platform presented here is an important step toward integrating membrane proteomic studies on chip, especially for future studies aimed at understanding fundamental effects of lipid interactions

  19. Membrane Protein Mobility and Orientation Preserved in Supported Bilayers Created Directly from Cell Plasma Membrane Blebs.

    PubMed

    Richards, Mark J; Hsia, Chih-Yun; Singh, Rohit R; Haider, Huma; Kumpf, Julia; Kawate, Toshimitsu; Daniel, Susan

    2016-03-29

    Membrane protein interactions with lipids are crucial for their native biological behavior, yet traditional characterization methods are often carried out on purified protein in the absence of lipids. We present a simple method to transfer membrane proteins expressed in mammalian cells to an assay-friendly, cushioned, supported lipid bilayer platform using cell blebs as an intermediate. Cell blebs, expressing either GPI-linked yellow fluorescent proteins or neon-green fused transmembrane P2X2 receptors, were induced to rupture on glass surfaces using PEGylated lipid vesicles, which resulted in planar supported membranes with over 50% mobility for multipass transmembrane proteins and over 90% for GPI-linked proteins. Fluorescent proteins were tracked, and their diffusion in supported bilayers characterized, using single molecule tracking and moment scaling spectrum (MSS) analysis. Diffusion was characterized for individual proteins as either free or confined, revealing details of the local lipid membrane heterogeneity surrounding the protein. A particularly useful result of our bilayer formation process is the protein orientation in the supported planar bilayer. For both the GPI-linked and transmembrane proteins used here, an enzymatic assay revealed that protein orientation in the planar bilayer results in the extracellular domains facing toward the bulk, and that the dominant mode of bleb rupture is via the "parachute" mechanism. Mobility, orientation, and preservation of the native lipid environment of the proteins using cell blebs offers advantages over proteoliposome reconstitution or disrupted cell membrane preparations, which necessarily result in significant scrambling of protein orientation and typically immobilized membrane proteins in SLBs. The bleb-based bilayer platform presented here is an important step toward integrating membrane proteomic studies on chip, especially for future studies aimed at understanding fundamental effects of lipid interactions

  20. Iterative linearized density matrix propagation for modeling coherent excitation energy transfer in photosynthetic light harvesting.

    PubMed

    Huo, P; Coker, D F

    2010-11-14

    Rather than incoherent hopping between chromophores, experimental evidence suggests that the excitation energy transfer in some biological light harvesting systems initially occurs coherently, and involves coherent superposition states in which excitation spreads over multiple chromophores separated by several nanometers. Treating such delocalized coherent superposition states in the presence of decoherence and dissipation arising from coupling to an environment is a significant challenge for conventional theoretical tools that either use a perturbative approach or make the Markovian approximation. In this paper, we extend the recently developed iterative linearized density matrix (ILDM) propagation scheme [E. R. Dunkel et al., J. Chem. Phys. 129, 114106 (2008)] to study coherent excitation energy transfer in a model of the Fenna-Matthews-Olsen light harvesting complex from green sulfur bacteria. This approach is nonperturbative and uses a discrete path integral description employing a short time approximation to the density matrix propagator that accounts for interference between forward and backward paths of the quantum excitonic system while linearizing the phase in the difference between the forward and backward paths of the environmental degrees of freedom resulting in a classical-like treatment of these variables. The approach avoids making the Markovian approximation and we demonstrate that it successfully describes the coherent beating of the site populations on different chromophores and gives good agreement with other methods that have been developed recently for going beyond the usual approximations, thus providing a new reliable theoretical tool to study coherent exciton transfer in light harvesting systems. We conclude with a discussion of decoherence in independent bilinearly coupled harmonic chromophore baths. The ILDM propagation approach in principle can be applied to more general descriptions of the environment.

  1. Double-membraned Liposomes Sculpted by Poliovirus 3AB Protein*

    PubMed Central

    Wang, Jing; Ptacek, Jennifer B.; Kirkegaard, Karla; Bullitt, Esther

    2013-01-01

    Infection with many positive-strand RNA viruses dramatically remodels cellular membranes, resulting in the accumulation of double-membraned vesicles that resemble cellular autophagosomes. In this study, a single protein encoded by poliovirus, 3AB, is shown to be sufficient to induce the formation of double-membraned liposomes via the invagination of single-membraned liposomes. Poliovirus 3AB is a 109-amino acid protein with a natively unstructured N-terminal domain. HeLa cells transduced with 3AB protein displayed intracellular membrane disruption; specifically, the formation of cytoplasmic invaginations. The ability of a single viral protein to produce structures of similar topology to cellular autophagosomes should facilitate the understanding of both cellular and viral mechanisms for membrane remodeling. PMID:23908350

  2. Disturbed vesicular trafficking of membrane proteins in prion disease.

    PubMed

    Uchiyama, Keiji; Miyata, Hironori; Sakaguchi, Suehiro

    2013-01-01

    The pathogenic mechanism of prion diseases remains unknown. We recently reported that prion infection disturbs post-Golgi trafficking of certain types of membrane proteins to the cell surface, resulting in reduced surface expression of membrane proteins and abrogating the signal from the proteins. The surface expression of the membrane proteins was reduced in the brains of mice inoculated with prions, well before abnormal symptoms became evident. Prions or pathogenic prion proteins were mainly detected in endosomal compartments, being particularly abundant in recycling endosomes. Some newly synthesized membrane proteins are delivered to the surface from the Golgi apparatus through recycling endosomes, and some endocytosed membrane proteins are delivered back to the surface through recycling endosomes. These results suggest that prions might cause neuronal dysfunctions and cell loss by disturbing post-Golgi trafficking of membrane proteins via accumulation in recycling endosomes. Interestingly, it was recently shown that delivery of a calcium channel protein to the cell surface was impaired and its function was abrogated in a mouse model of hereditary prion disease. Taken together, these results suggest that impaired delivery of membrane proteins to the cell surface is a common pathogenic event in acquired and hereditary prion diseases.

  3. Long-distance electronic energy transfer in light-harvesting supramolecular polymers.

    PubMed

    Winiger, Christian B; Li, Shaoguang; Kumar, Ganesh R; Langenegger, Simon M; Häner, Robert

    2014-12-01

    The efficient collection of solar energy relies on the design and construction of well-organized light-harvesting systems. Herein we report that supramolecular phenanthrene polymers doped with pyrene are effective collectors of light energy. The linear polymers are formed through the assembly of short amphiphilic oligomers in water. Absorption of light by phenanthrene residues is followed by electronic energy transfer along the polymer over long distances (>100 nm) to the accepting pyrene molecules. The high efficiency of the energy transfer, which is documented by large fluorescence quantum yields, suggests a quantum coherent process.

  4. Path induced coherent energy transfer in light-harvesting complexes in purple bacteria.

    PubMed

    Sun, Kewei; Ye, Jun; Zhao, Yang

    2014-09-28

    Features of path dependent energy transfer in a dual-ring light-harvesting (LH2) complexes (B850) system have been examined in detail systematically. The Frenkel-Dirac time dependent variational method with the Davydov D1 Ansatz is employed with detailed evolution of polaron dynamics in real space readily obtained. It is found that the phase of the transmission amplitude through the LH2 complexes plays an important role in constructing the coherent excitonic energy transfer. It is also found that the symmetry breaking caused by the dimerization of bacteriochlorophylls and coherence or correlation between two rings will be conducive in enhancing the exciton transfer efficiency. PMID:25273408

  5. The dynamics of structural deformations of immobilized single light-harvesting complexes

    PubMed Central

    Bopp, Martin A.; Sytnik, Alexander; Howard, Tina D.; Cogdell, Richard J.; Hochstrasser, Robin M.

    1999-01-01

    Single assemblies of the intact light-harvesting complex LH2 from Rhodopseudomonas acidophila were bound to mica surfaces at 300 K and examined by observing their fluorescence after polarized light excitation. The complexes are generally not cylindrically symmetric. They act like elliptic absorbers, indicating that the high symmetry found in crystals of LH2 is not present when the molecules are immobilized on mica. The ellipticity and the principal axes of the ellipses fluctuate on the time scale of seconds, indicating that there is a mobile structural deformation. The B850 ring of cofactors shows significantly less asymmetry than B800. The photobleaching strongly depends on the presence of oxygen. PMID:10500166

  6. Neutron and light scattering studies of light-harvesting photosynthetic antenna complexes

    SciTech Connect

    Tang, Kuo-Hsiang; Blankenship, Robert E.

    2011-06-28

    Small-angle neutron scattering (SANS) and dynamic light scattering (DLS) have been employed in studying the structural information of various biological systems, particularly in systems without high-resolution structural information available. In this report, we briefly present some principles and biological applications of neutron scattering and DLS, compare the differences in information that can be obtained with small-angle X-ray scattering (SAXS), and then report recent studies of SANS and DLS, together with other biophysical approaches, for light-harvesting antenna complexes and reaction centers of purple and green phototrophic bacteria.

  7. Functionalized Nanoparticles and Surfaces for Controlled Chemical Catalysis and Effective Light Harvesting

    SciTech Connect

    Marye Anne Fox, James K. Whitesell

    2012-11-02

    We have prepared a range of such arrays as key components for biotechnology and photonic applications. These involve self-assembled arrays of increasing complexity with three-dimensionally disposed multilayer interactions. These arrays also include dendrimers as the distinguishing structural building blocks. These photoactive integrated systems have a regular, highly-branched, three-dimensional architecture. Structural modifications of these units include variation of the core, bridging layers, and terminal groups. These modifications result in a large array of dendritic molecules with potential applications for light harvesting.

  8. Energy transfer from conjugated polymer to bacterial light-harvesting complex

    NASA Astrophysics Data System (ADS)

    Buczynska, D.; Bujak, Ł.; Loi, M. A.; Brotosudarmo, T. H. P.; Cogdell, R.; Mackowski, S.

    2012-10-01

    Energy transfer from a conjugated polymer blend (poly(9,9-dioctylfluorenyl-2,7-diyl):poly (2-methoxy-5-(2-ethylhexyloxy)-1, 4-phenylenevinylene) to a light-harvesting complex 2 from purple bacteria has been demonstrated using time-resolved fluorescence spectroscopy. For our hybrid nanostructure, we observe a 30% reduction of the fluorescence lifetime of the polymer emission as compared to the pure polymer layer. This result is an important step towards integrating naturally evolved biomolecules with synthetic materials into biohybrid organic electronic systems.

  9. Efficient Light Harvester Layer Prepared by Solid/Mist Interface Reaction for Perovskite Solar Cells.

    PubMed

    Xia, Xiang; Li, Hongcui; Wu, Wenyi; Li, Yanhua; Fei, Dehou; Gao, Chunxiao; Liu, Xizhe

    2015-08-12

    A solid/mist reaction method is developed to produce well-crystallized light harvester layers without pinhole defects for perovskite solar cells. The reaction based on mist precursor can be facilely operated with low process temperature. And it can effectively control the volume of CH3NH3I solution and the reaction temperature, which affect the quality of perovskite harvester layers and the performance of perovskite solar cells remarkably. Under optimized condition, the efficiencies of devices reach 16.2% with the average efficiency of 14.9%. The solid/mist reaction is also used to fabricate planar junction solar cells and a PCE of 14.9% is obtained.

  10. Toward understanding as photosynthetic biosignatures: light harvesting and energy transfer calculation

    NASA Astrophysics Data System (ADS)

    Komatsu, Y.; Umemura, M.; Shoji, M.; Shiraishi, K.; Kayanuma, M.; Yabana, K.

    2014-03-01

    Among several proposed biosignatures, red edge is a direct evidence of photosynthetic life if it is detected (Kiang et al 2007). Red edge is a sharp change in reflectance spectra of vegetation in NIR region (about 700-750 nm). The sign of red edge is observed by Earthshine or remote sensing (Wolstencroft & Raven 2002, Woolf et al 2002). But, why around 700-750 nm? The photosynthetic organisms on Earth have evolved to optimize the sunlight condition. However, if we consider about photosynthetic organism on extrasolar planets, they should have developed to utilize the spectra of its principal star. Thus, it is not strange even if it shows different vegetation spectra. In this study, we focused on the light absorption mechanism of photosynthetic organisms on Earth and investigated the fundamental properties of the light harvesting mechanisms, which is the first stage for the light absorption. Light harvesting complexes contain photosynthetic pigments like chlorophylls. Effective light absorption and the energy transfer are accomplished by the electronic excitations of collective photosynthetic pigments. In order to investigate this mechanism, we constructed an energy transfer model by using a dipole-dipole approximation for the interactions between electronic excitations. Transition moments and transition energies of each pigment are calculated at the time-dependent density functional theory (TDDFT) level (Marques & Gross 2004). Quantum dynamics simulation for the excitation energy transfer was calculated by the Liouvelle's equation. We adopted the model to purple bacteria, which has been studied experimentally and known to absorb lower energy. It is meaningful to focus on the mechanism of this bacteria, since in the future mission, M planets will become a important target. We calculated the oscillator strengths in one light harvesting complex and confirmed the validity by comparing to the experimental data. This complex is made of an inner and an outer ring. The

  11. Fluorescence enhancement of light-harvesting complex 2 from purple bacteria coupled to spherical gold nanoparticles

    SciTech Connect

    Bujak, Ł.; Czechowski, N.; Piatkowski, D.; Litvin, R.; Mackowski, S.; Brotosudarmo, T. H. P.; Pichler, S.; Cogdell, R. J.; Heiss, W.

    2011-10-24

    The influence of plasmon excitations in spherical gold nanoparticles on the optical properties of a light-harvesting complex 2 (LH2) from the purple bacteria Rhodopseudomonas palustris has been studied. Systematic analysis is facilitated by controlling the thickness of a silica layer between Au nanoparticles and LH2 complexes. Fluorescence of LH2 complexes features substantial increase when these complexes are separated by 12 nm from the gold nanoparticles. At shorter distances, non-radiative quenching leads to a decrease of fluorescence emission. The enhancement of fluorescence originates predominantly from an increase of absorption of pigments comprising the LH2 complex.

  12. High-Throughput Baculovirus Expression System for Membrane Protein Production.

    PubMed

    Kalathur, Ravi C; Panganiban, Marinela; Bruni, Renato

    2016-01-01

    The ease of use, robustness, cost-effectiveness, and posttranslational machinery make baculovirus expression system a popular choice for production of eukaryotic membrane proteins. This system can be readily adapted for high-throughput operations. This chapter outlines the techniques and procedures for cloning, transfection, small-scale production, and purification of membrane protein samples in a high-throughput manner. PMID:27485337

  13. Negative Ions Enhance Survival of Membrane Protein Complexes

    NASA Astrophysics Data System (ADS)

    Liko, Idlir; Hopper, Jonathan T. S.; Allison, Timothy M.; Benesch, Justin L. P.; Robinson, Carol V.

    2016-06-01

    Membrane protein complexes are commonly introduced to the mass spectrometer solubilized in detergent micelles. The collisional activation used to remove the detergent, however, often causes protein unfolding and dissociation. As in the case for soluble proteins, electrospray in the positive ion mode is most commonly used for the study of membrane proteins. Here we show several distinct advantages of employing the negative ion mode. Negative polarity can yield lower average charge states for membrane proteins solubilized in saccharide detergents, with enhanced peak resolution and reduced adduct formation. Most importantly, we demonstrate that negative ion mode electrospray ionization (ESI) minimizes subunit dissociation in the gas phase, allowing access to biologically relevant oligomeric states. Together, these properties mean that intact membrane protein ions can be generated in a greater range of solubilizing detergents. The formation of negative ions, therefore, greatly expands the possibilities of using mass spectrometry on this intractable class of protein.

  14. Protein quality control at the inner nuclear membrane.

    PubMed

    Khmelinskii, Anton; Blaszczak, Ewa; Pantazopoulou, Marina; Fischer, Bernd; Omnus, Deike J; Le Dez, Gaëlle; Brossard, Audrey; Gunnarsson, Alexander; Barry, Joseph D; Meurer, Matthias; Kirrmaier, Daniel; Boone, Charles; Huber, Wolfgang; Rabut, Gwenaël; Ljungdahl, Per O; Knop, Michael

    2014-12-18

    The nuclear envelope is a double membrane that separates the nucleus from the cytoplasm. The inner nuclear membrane (INM) functions in essential nuclear processes including chromatin organization and regulation of gene expression. The outer nuclear membrane is continuous with the endoplasmic reticulum and is the site of membrane protein synthesis. Protein homeostasis in this compartment is ensured by endoplasmic-reticulum-associated protein degradation (ERAD) pathways that in yeast involve the integral membrane E3 ubiquitin ligases Hrd1 and Doa10 operating with the E2 ubiquitin-conjugating enzymes Ubc6 and Ubc7 (refs 2, 3). However, little is known about protein quality control at the INM. Here we describe a protein degradation pathway at the INM in yeast (Saccharomyces cerevisiae) mediated by the Asi complex consisting of the RING domain proteins Asi1 and Asi3 (ref. 4). We report that the Asi complex functions together with the ubiquitin-conjugating enzymes Ubc6 and Ubc7 to degrade soluble and integral membrane proteins. Genetic evidence suggests that the Asi ubiquitin ligase defines a pathway distinct from, but complementary to, ERAD. Using unbiased screening with a novel genome-wide yeast library based on a tandem fluorescent protein timer, we identify more than 50 substrates of the Asi, Hrd1 and Doa10 E3 ubiquitin ligases. We show that the Asi ubiquitin ligase is involved in degradation of mislocalized integral membrane proteins, thus acting to maintain and safeguard the identity of the INM. PMID:25519137

  15. Protein quality control at the inner nuclear membrane

    PubMed Central

    Khmelinskii, Anton; Blaszczak, Ewa; Pantazopoulou, Marina; Fischer, Bernd; Omnus, Deike J.; Le Dez, Gaëlle; Brossard, Audrey; Gunnarsson, Alexander; Barry, Joseph D.; Meurer, Matthias; Kirrmaier, Daniel; Boone, Charles; Huber, Wolfgang; Rabut, Gwenaël; Ljungdahl, Per O.; Knop, Michael

    2015-01-01

    The nuclear envelope is a double membrane that separates the nucleus from the cytoplasm. The inner nuclear membrane (INM) functions in essential nuclear processes including chromatin organization and regulation of gene expression1. The outer nuclear membrane is continuous with the endoplasmic reticulum (ER) and is the site of membrane protein synthesis. Protein homeostasis in this compartment is ensured by ER-associated protein degradation (ERAD) pathways that in yeast involve the integral membrane E3 ubiquitin ligases Hrd1 and Doa10 operating with the E2 ubiquitin-conjugating enzymes Ubc6 and Ubc72,3. However, little is known regarding protein quality control at the INM. Here we describe a protein degradation pathway at the INM mediated by the Asi complex consisting of the RING domain proteins Asi1 and Asi34. We report that the As complex functions together with the ubiquitin conjugating enzymes Ubc6andUbc7to degrade soluble and integral membrane proteins. Genetic evidence suggest that the Asi ubiquitin ligase defines a pathway distinct from but complementary to ERAD. Using unbiased screening with a novel genome-wide yeast library based on a tandem fluorescent protein timer (tFT)5, we identify more than 50 substrates of the Asi, Hrd1 and Doa10 E3 ubiquity ligases. We show that the Asi ubiquitin ligase is involved in degradation of mislocalised integral membrane proteins, thus acting to maintain and safeguard the identity of the INM. PMID:25519137

  16. BPROMPT: A consensus server for membrane protein prediction.

    PubMed

    Taylor, Paul D; Attwood, Teresa K; Flower, Darren R

    2003-07-01

    Protein structure prediction is a cornerstone of bioinformatics research. Membrane proteins require their own prediction methods due to their intrinsically different composition. A variety of tools exist for topology prediction of membrane proteins, many of them available on the Internet. The server described in this paper, BPROMPT (Bayesian PRediction Of Membrane Protein Topology), uses a Bayesian Belief Network to combine the results of other prediction methods, providing a more accurate consensus prediction. Topology predictions with accuracies of 70% for prokaryotes and 53% for eukaryotes were achieved. BPROMPT can be accessed at http://www.jenner.ac.uk/BPROMPT. PMID:12824397

  17. Prediction of lipid-binding regions in cytoplasmic and extracellular loops of membrane proteins as exemplified by protein translocation membrane proteins.

    PubMed

    Keller, Rob C A

    2013-01-01

    The presence of possible lipid-binding regions in the cytoplasmic or extracellular loops of membrane proteins with an emphasis on protein translocation membrane proteins was investigated in this study using bioinformatics. Recent developments in approaches recognizing lipid-binding regions in proteins were found to be promising. In this study a total bioinformatics approach specialized in identifying lipid-binding helical regions in proteins was explored. Two features of the protein translocation membrane proteins, the position of the transmembrane regions and the identification of additional lipid-binding regions, were analyzed. A number of well-studied protein translocation membrane protein structures were checked in order to demonstrate the predictive value of the bioinformatics approach. Furthermore, the results demonstrated that lipid-binding regions in the cytoplasmic and extracellular loops in protein translocation membrane proteins can be predicted, and it is proposed that the interaction of these regions with phospholipids is important for proper functioning during protein translocation. PMID:22961045

  18. Charge transfer and quantum coherence in solar cells and artificial light harvesting system

    NASA Astrophysics Data System (ADS)

    Lienau, Christoph

    2014-03-01

    In artificial light harvesting systems the conversion of light into electrical or chemical energy happens on the femtosecond time scale, and is thought to involve the incoherent jump of an electron from the optical absorber to an electron acceptor. Here we investigate the primary dynamics of the photoinduced electronic charge transfer process in two prototypical structures: (i) a carotene-porphyrin-fullerene triad, a prototypical elementary component for an artificial light harvesting system and (ii) a polymer:fullerene blend as a model system for an organic solar cell. Our approach combines coherent femtosecond spectroscopy and first-principles quantum dynamics simulations. Our experimental and theoretical results provide strong evidence that the driving mechanism of the primary step within the current generation cycle is a quantum-correlated wavelike motion of electrons and nuclei on a timescale of few tens of femtoseconds. We furthermore highlight the fundamental role played by the flexible interface between the light-absorbing chromophore and the charge acceptor in triggering the coherent wavelike electron-hole splitting.

  19. Realizing omnidirectional light harvesting by employing hierarchical architecture for dye sensitized solar cells.

    PubMed

    Hsieh, Ming-Yang; Lai, Fang-I; Chen, Wei-Chun; Hsieh, Min-Chi; Hu, Hsiang-Yi; Yu, Peichen; Kuo, Hao-Chung; Kuo, Shou-Yi

    2016-03-14

    To improve the omnidirectional light-harvesting in dye-sensitized solar cells (DSSCs), here we present a dandelion-like structure composed of ZnO hemispherical shells and nanorods. Uniformly distributed hemispherical shells effectively suppress the reflection over the broadband region at incident angles up to 60°, greatly improving the optical absorption of the DSSCs. In addition, modulating the length of the ZnO nanorods controls the omnidirectional characteristics of DSSCs. This phenomenon is attributed to the degree of periodicity of the ZnO dandelion-like structures. Cells with shorter rods exhibit a high degree of periodicity, thus the conversion efficiencies of the cells show specific angle-independent features. On the other hand, the cells with longer lengths reveal angle-dependent photovoltaic performance. Along with the simulation, the cells with dandelion-like ZnO structures can couple incident photons efficiently to achieve excellent broadband and omnidirectional light-harvesting performances experimentally, and the DSSCs enhanced the conversion efficiency by 48% at large incident angles. All these findings not only provide further insight into the light-trapping mechanism in these complex three-dimensional nanostructures but also offer efficient omnidirectional and broadband nanostructured photovoltaics for advanced applications. PMID:26899775

  20. Realizing omnidirectional light harvesting by employing hierarchical architecture for dye sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Hsieh, Ming-Yang; Lai, Fang-I.; Chen, Wei-Chun; Hsieh, Min-Chi; Hu, Hsiang-Yi; Yu, Peichen; Kuo, Hao-Chung; Kuo, Shou-Yi

    2016-03-01

    To improve the omnidirectional light-harvesting in dye-sensitized solar cells (DSSCs), here we present a dandelion-like structure composed of ZnO hemispherical shells and nanorods. Uniformly distributed hemispherical shells effectively suppress the reflection over the broadband region at incident angles up to 60°, greatly improving the optical absorption of the DSSCs. In addition, modulating the length of the ZnO nanorods controls the omnidirectional characteristics of DSSCs. This phenomenon is attributed to the degree of periodicity of the ZnO dandelion-like structures. Cells with shorter rods exhibit a high degree of periodicity, thus the conversion efficiencies of the cells show specific angle-independent features. On the other hand, the cells with longer lengths reveal angle-dependent photovoltaic performance. Along with the simulation, the cells with dandelion-like ZnO structures can couple incident photons efficiently to achieve excellent broadband and omnidirectional light-harvesting performances experimentally, and the DSSCs enhanced the conversion efficiency by 48% at large incident angles. All these findings not only provide further insight into the light-trapping mechanism in these complex three-dimensional nanostructures but also offer efficient omnidirectional and broadband nanostructured photovoltaics for advanced applications.

  1. Exploiting Collective Effects to Direct Light Absorption in Natural and Artificial Light-Harvesters

    NASA Astrophysics Data System (ADS)

    Schroeder, Christopher

    Photosynthesis---the conversion of sunlight to chemical energy---is fundamental for supporting life on our planet. Despite its importance, the physical principles that underpin the primary steps of photosynthesis, from photon absorption to electronic charge separation, remain to be understood in full. Electronic coherence within tightly-packed light-harvesting (LH) units or within individual reaction centers (RCs) has been recognized as an important ingredient for a complete understanding of the excitation energy transfer (EET) dynamics. However, the electronic coherence across units---RC and LH or LH and LH---has been consistently neglected as it does not play a significant role during these relatively slow transfer processes. Here, we turn our attention to the absorption process, which, as we will show, has a much shorter built-in timescale. We demonstrate that the---often overlooked---spatially extended but short-lived excitonic delocalization plays a relevant role in general photosynthetic systems. Most strikingly, we find that absorption intensity is, quite generally, redistributed from LH units to the RC, increasing the number of excitations which can effect charge separation without further transfer steps. A biomemetic nano-system is proposed which is predicted to funnel excitation to the RC-analogue, and hence is the first step towards exploiting these new design principles for efficient artificial light-harvesting.

  2. Structural insights into energy regulation of light-harvesting complex CP29 from spinach.

    PubMed

    Pan, Xiaowei; Li, Mei; Wan, Tao; Wang, Longfei; Jia, Chenjun; Hou, Zhiqiang; Zhao, Xuelin; Zhang, Jiping; Chang, Wenrui

    2011-03-01

    CP29, one of the minor light-harvesting complexes of higher-plant photosystem II, absorbs and transfers solar energy for photosynthesis and also has important roles in photoprotection. We have solved the crystal structure of spinach CP29 at 2.80-Å resolution. Each CP29 monomer contains 13 chlorophyll and 3 carotenoid molecules, which differs considerably from the major light-harvesting complex LHCII and the previously proposed CP29 model. The 13 chlorophyll-binding sites are assigned as eight chlorophyll a sites, four chlorophyll b and one putative mixed site occupied by both chlorophylls a and b. Based on the present X-ray structure, an integrated pigment network in CP29 is constructed. Two special clusters of pigment molecules, namely a615-a611-a612-Lut and Vio(Zea)-a603-a609, have been identified and might function as potential energy-quenching centers and as the exit or entrance in energy-transfer pathways.

  3. Light-harvesting cross-linked polymers for efficient heterogeneous photocatalysis.

    PubMed

    Wang, Cheng; Xie, Zhigang; deKrafft, Kathryn E; Lin, Wenbin

    2012-04-01

    Nonporous, phosphorescent cross-linked polymers (Ru-CP and Ir-CP) were synthesized via Pd-catalyzed Sonogashira cross-coupling reactions between tetra(p-ethynylphenyl)methane and dibrominated Ru(bpy)(3)(2+) or Ir(ppy)(2)(bpy)(+), respectively. The resultant particulate cross-linked polymer (CP) materials have very high catalyst loadings (76.3 wt % for Ru-CP and 71.6 wt % for Ir-CP), and are nonporous with negligibly small surface areas (2.9 m(2)/g for Ru-CP and 2.7 m(2)/g for Ir-CP). Despite their nonporous nature, the insoluble CP materials serve as highly active and recyclable heterogeneous photocatalysts for a range of organic transformations such as aza-Henry reaction, aerobic amine coupling, and dehalogenation of benzyl bromoacetate. An efficient light-harvesting mechanism, which involves collection of photons by exciting the (3)MLCT states of the phosphors and migration of the excited states to the particle surface, is proposed to account for the very high catalytic activities of these nonporous CPs. Steady-state and time-resolved emission data, as well as the reduced catalytic activity of Os(bpy)(3)(2+)-doped Ru-CPs supports efficient excited state migration for the CP frameworks. This work uncovers a new strategy in designing highly efficient photocatalysts based on light-harvesting cross-linked polymers.

  4. Light-Harvesting Antenna System from the Phototrophic Bacterium Roseiflexus castenholzii

    SciTech Connect

    Collins, Aaron M.; Qian, Pu; Tang, Qun; Bocian, David F; Hunter, C. Neil; Blankenship, Robert E.

    2010-08-12

    Photosynthetic organisms have evolved diverse light-harvesting complexes to harness light of various qualities and intensities. Photosynthetic bacteria can have (bacterio)chlorophyll Qy antenna absorption bands ranging from ~650 to ~1100 nm. This broad range of wavelengths has allowed many organisms to thrive in unique light environments. Roseiflexus castenholzii is a niche-adapted, filamentous anoxygenic phototroph (FAP) that lacks chlorosomes, the dominant antenna found in most green bacteria, and here we describe the purification of a full complement of photosynthetic complexes: the light-harvesting (LH) antenna, reaction center (RC), and core complex (RC-LH). By high-performance liquid chromatography separation of bacteriochlorophyll and bacteriopheophytin pigments extracted from the core complex and the RC, the number of subunits that comprise the antenna was determined to be 15 ± 1. Resonance Raman spectroscopy of the carbonyl stretching region displayed modes indicating that 3C-acetyl groups of BChl a are all involved in molecular interactions probably similar to those found in LH1 complexes from purple photosynthetic bacteria. Finally, two-dimensional projections of negatively stained core complexes and the LH antenna revealed a closed, slightly elliptical LH ring with an average diameter of 130 ± 10 Å surrounding a single RC that lacks an H-subunit but is associated with a tetraheme c-type cytochrome.

  5. Photothermal light harvesting and light-gated molecular release by nanoporous gold disks

    NASA Astrophysics Data System (ADS)

    Santos, Greggy M.; Zhao, Fusheng; Zeng, Jianbo; Shih, Wei-Chuan

    2015-03-01

    Photothermal heating has been an effective mechanism for harvesting light energy by plasmonic resonance. Photothermally generated hyperthermia can alter cell behavior, change cell microenvironment, and promote or suppress cell growth. In the past, colloidal nanoparticles such as gold nanospheres, nanoshells, nanorods, and nanocages have been developed for various applications. Here, we show that nanoporous gold disks (NPGDs) with 400 nm diameter, 75 nm thickness, and 13 nm pores exhibit large specific surface area and effective photothermal light harvesting capability. Another potential application is demonstrated by light-gated, multi-step molecular release of pre-adsorbed R6G fluorescent dye on arrayed NPGDs. Through the use of time-resolved temperature mapping, the spatial and temporal characteristics of photothermal heating in NPGD arrays is successfully demonstrated for both aqueous and air ambient environments. By applying a thermodynamic model to our experimental data, we determined the photothermal conversion efficiency at 56% for NPGD arrays. As a potential application, light-gated, multi-stage release of pre-adsorbed R6G dye molecules from NPGD arrays has been demonstrated. The results establish the foundation that NPGDs can be employed for photothermal light harvesting and light-gated molecular release.

  6. Overcoming bottlenecks in the membrane protein structural biology pipeline.

    PubMed

    Hardy, David; Bill, Roslyn M; Jawhari, Anass; Rothnie, Alice J

    2016-06-15

    Membrane proteins account for a third of the eukaryotic proteome, but are greatly under-represented in the Protein Data Bank. Unfortunately, recent technological advances in X-ray crystallography and EM cannot account for the poor solubility and stability of membrane protein samples. A limitation of conventional detergent-based methods is that detergent molecules destabilize membrane proteins, leading to their aggregation. The use of orthologues, mutants and fusion tags has helped improve protein stability, but at the expense of not working with the sequence of interest. Novel detergents such as glucose neopentyl glycol (GNG), maltose neopentyl glycol (MNG) and calixarene-based detergents can improve protein stability without compromising their solubilizing properties. Styrene maleic acid lipid particles (SMALPs) focus on retaining the native lipid bilayer of a membrane protein during purification and biophysical analysis. Overcoming bottlenecks in the membrane protein structural biology pipeline, primarily by maintaining protein stability, will facilitate the elucidation of many more membrane protein structures in the near future. PMID:27284049

  7. Concentrating membrane proteins using asymmetric traps and AC electric fields.

    PubMed

    Cheetham, Matthew R; Bramble, Jonathan P; McMillan, Duncan G G; Krzeminski, Lukasz; Han, Xiaojun; Johnson, Benjamin R G; Bushby, Richard J; Olmsted, Peter D; Jeuken, Lars J C; Marritt, Sophie J; Butt, Julea N; Evans, Stephen D

    2011-05-01

    Membrane proteins are key components of the plasma membrane and are responsible for control of chemical ionic gradients, metabolite and nutrient transfer, and signal transduction between the interior of cells and the external environment. Of the genes in the human genome, 30% code for membrane proteins (Krogh et al. J. Mol. Biol.2001, 305, 567). Furthermore, many FDA-approved drugs target such proteins (Overington et al. Nat. Rev. Drug Discovery 2006, 5, 993). However, the structure-function relationships of these are notably sparse because of difficulties in their purification and handling outside of their membranous environment. Methods that permit the manipulation of membrane components while they are still in the membrane would find widespread application in separation, purification, and eventual structure-function determination of these species (Poo et al. Nature 1977, 265, 602). Here we show that asymmetrically patterned supported lipid bilayers in combination with AC electric fields can lead to efficient manipulation of charged components. We demonstrate the concentration and trapping of such components through the use of a "nested trap" and show that this method is capable of yielding an approximately 30-fold increase in the average protein concentration. Upon removal of the field, the material remains trapped for several hours as a result of topographically restricted diffusion. Our results indicate that this method can be used for concentrating and trapping charged membrane components while they are still within their membranous environment. We anticipate that our approach could find widespread application in the manipulation and study of membrane proteins.

  8. Concentrating membrane proteins using asymmetric traps and AC electric fields.

    PubMed

    Cheetham, Matthew R; Bramble, Jonathan P; McMillan, Duncan G G; Krzeminski, Lukasz; Han, Xiaojun; Johnson, Benjamin R G; Bushby, Richard J; Olmsted, Peter D; Jeuken, Lars J C; Marritt, Sophie J; Butt, Julea N; Evans, Stephen D

    2011-05-01

    Membrane proteins are key components of the plasma membrane and are responsible for control of chemical ionic gradients, metabolite and nutrient transfer, and signal transduction between the interior of cells and the external environment. Of the genes in the human genome, 30% code for membrane proteins (Krogh et al. J. Mol. Biol.2001, 305, 567). Furthermore, many FDA-approved drugs target such proteins (Overington et al. Nat. Rev. Drug Discovery 2006, 5, 993). However, the structure-function relationships of these are notably sparse because of difficulties in their purification and handling outside of their membranous environment. Methods that permit the manipulation of membrane components while they are still in the membrane would find widespread application in separation, purification, and eventual structure-function determination of these species (Poo et al. Nature 1977, 265, 602). Here we show that asymmetrically patterned supported lipid bilayers in combination with AC electric fields can lead to efficient manipulation of charged components. We demonstrate the concentration and trapping of such components through the use of a "nested trap" and show that this method is capable of yielding an approximately 30-fold increase in the average protein concentration. Upon removal of the field, the material remains trapped for several hours as a result of topographically restricted diffusion. Our results indicate that this method can be used for concentrating and trapping charged membrane components while they are still within their membranous environment. We anticipate that our approach could find widespread application in the manipulation and study of membrane proteins. PMID:21476549

  9. Phenotypic effects of membrane protein overexpression in Saccharomyces cerevisiae.

    PubMed

    Osterberg, Marie; Kim, Hyun; Warringer, Jonas; Melén, Karin; Blomberg, Anders; von Heijne, Gunnar

    2006-07-25

    Large-scale protein overexpression phenotype screens provide an important complement to the more common gene knockout screens. Here, we have targeted the so far poorly understood Saccharomyces cerevisiae membrane proteome and report growth phenotypes for a strain collection overexpressing approximately 600 C-terminally tagged integral membrane proteins grown both under normal and three different stress conditions. Although overexpression of most membrane proteins reduce the growth rate in synthetic defined medium, we identify a large number of proteins that, when overexpressed, confer specific resistance to various stress conditions. Our data suggest that regulation of glycosylphosphatidylinositol anchor biosynthesis and the Na(+)/K(+) homeostasis system constitute major downstream targets of the yeast PKA/RAS pathway and point to a possible connection between the early secretory pathway and the cells' response to oxidative stress. We also have quantified the expression levels for >550 membrane proteins, facilitating the choice of well expressing proteins for future functional and structural studies.

  10. Membrane proteins of Mycoplasma bovis and their role in pathogenesis.

    PubMed

    Adamu, James Y; Wawegama, Nadeeka K; Browning, Glenn F; Markham, Philip F

    2013-10-01

    Mycoplasma membrane proteins influence cell shape, cell division, motility and adhesion to host cells, and are thought to be integrally involved in the pathogenesis of mycoplasmoses. Many of the membrane proteins predicted from mycoplasma genome sequences remain hypothetical, as their presence in cellular protein preparations is yet to be established experimentally. Recent genome sequences of several strains of Mycoplasma bovis have provided further insight into the potential role of the membrane proteins of this pathogen in colonisation and infection. This review highlights recent advances in knowledge about the influence of M. bovis membrane proteins on the pathogenesis of infection with this species and identifies future research directions for enhancing our understanding of the role of these proteins. PMID:23810376

  11. Spectral heterogeneity and carotenoid-to-bacteriochlorophyll energy transfer in LH2 light-harvesting complexes from Allochromatium vinosum.

    PubMed

    Magdaong, Nikki M; LaFountain, Amy M; Hacking, Kirsty; Niedzwiedzki, Dariusz M; Gibson, George N; Cogdell, Richard J; Frank, Harry A

    2016-02-01

    Photosynthetic organisms produce a vast array of spectral forms of antenna pigment-protein complexes to harvest solar energy and also to adapt to growth under the variable environmental conditions of light intensity, temperature, and nutrient availability. This behavior is exemplified by Allochromatium (Alc.) vinosum, a photosynthetic purple sulfur bacterium that produces different types of LH2 light-harvesting complexes in response to variations in growth conditions. In the present work, three different spectral forms of LH2 from Alc. vinosum, B800-820, B800-840, and B800-850, were isolated, purified, and examined using steady-state absorption and fluorescence spectroscopy, and ultrafast time-resolved absorption spectroscopy. The pigment composition of the LH2 complexes was analyzed by high-performance liquid chromatography, and all were found to contain five carotenoids: lycopene, anhydrorhodovibrin, spirilloxanthin, rhodopin, and rhodovibrin. Spectral reconstructions of the absorption and fluorescence excitation spectra based on the pigment composition revealed significantly more spectral heterogeneity in these systems compared to LH2 complexes isolated from other species of purple bacteria. The data also revealed the individual carotenoid-to-bacteriochlorophyll energy transfer efficiencies which were correlated with the kinetic data from the ultrafast transient absorption spectroscopic experiments. This series of LH2 complexes allows a systematic exploration of the factors that determine the spectral properties of the bound pigments and control the rate and efficiency of carotenoid-to-bacteriochlorophyll energy transfer. PMID:26048106

  12. Spectral heterogeneity and carotenoid-to-bacteriochlorophyll energy transfer in LH2 light-harvesting complexes from Allochromatium vinosum.

    PubMed

    Magdaong, Nikki M; LaFountain, Amy M; Hacking, Kirsty; Niedzwiedzki, Dariusz M; Gibson, George N; Cogdell, Richard J; Frank, Harry A

    2016-02-01

    Photosynthetic organisms produce a vast array of spectral forms of antenna pigment-protein complexes to harvest solar energy and also to adapt to growth under the variable environmental conditions of light intensity, temperature, and nutrient availability. This behavior is exemplified by Allochromatium (Alc.) vinosum, a photosynthetic purple sulfur bacterium that produces different types of LH2 light-harvesting complexes in response to variations in growth conditions. In the present work, three different spectral forms of LH2 from Alc. vinosum, B800-820, B800-840, and B800-850, were isolated, purified, and examined using steady-state absorption and fluorescence spectroscopy, and ultrafast time-resolved absorption spectroscopy. The pigment composition of the LH2 complexes was analyzed by high-performance liquid chromatography, and all were found to contain five carotenoids: lycopene, anhydrorhodovibrin, spirilloxanthin, rhodopin, and rhodovibrin. Spectral reconstructions of the absorption and fluorescence excitation spectra based on the pigment composition revealed significantly more spectral heterogeneity in these systems compared to LH2 complexes isolated from other species of purple bacteria. The data also revealed the individual carotenoid-to-bacteriochlorophyll energy transfer efficiencies which were correlated with the kinetic data from the ultrafast transient absorption spectroscopic experiments. This series of LH2 complexes allows a systematic exploration of the factors that determine the spectral properties of the bound pigments and control the rate and efficiency of carotenoid-to-bacteriochlorophyll energy transfer.

  13. Atomistic modeling of two-dimensional electronic spectra and excited-state dynamics for a Light Harvesting 2 complex.

    PubMed

    van der Vegte, C P; Prajapati, J D; Kleinekathöfer, U; Knoester, J; Jansen, T L C

    2015-01-29

    The Light Harvesting 2 (LH2) complex is a vital part of the photosystem of purple bacteria. It is responsible for the absorption of light and transport of the resulting excitations to the reaction center in a highly efficient manner. A general description of the chromophores and the interaction with their local environment is crucial to understand this highly efficient energy transport. Here we include this interaction in an atomistic way using mixed quantum-classical (molecular dynamics) simulations of spectra. In particular, we present the first atomistic simulation of nonlinear optical spectra for LH2 and use it to study the energy transport within the complex. We show that the frequency distributions of the pigments strongly depend on their positions with respect to the protein scaffold and dynamics of their local environment. Furthermore, we show that although the pigments are closely packed the transition frequencies of neighboring pigments are essentially uncorrelated. We present the simulated linear absorption spectra for the LH2 complex and provide a detailed explanation of the states responsible for the observed two-band structure. Finally, we discuss the energy transfer within the complex by analyzing population transfer calculations and 2D spectra for different waiting times. We conclude that the energy transfer from the B800 ring to the B850 ring is mediated by intermediate states that are delocalized over both rings, allowing for a stepwise downhill energy transport.

  14. Composition of photosystem II antenna in light-harvesting complex II antisense tobacco plants at varying irradiances.

    PubMed Central

    Flachmann, R

    1997-01-01

    Plants with genes coding for chlorophyll a/b-binding proteins of light-harvesting complex II (LHCII) in antisense orientation (Lhcb) that are characterized by severely reduced Lhcb transcript levels (below 10% of wild type) do not show a bleached phenotype due to a specific loss of the polypeptide. To produce such a phenotype, a conceptually different antisense approach was tested with a dual-functional transcript encoding the gene for hygromycin phosphotransferase and the transit sequence of Lhcb1-2 in the antisense orientation. Using increasing concentrations of hygromycin, transformants with Lhcb steady-state levels as low as 9% of wild type were regenerated and grown in a growth chamber. Together with Lhcb antisense plants obtained in an earlier study, these antisense plants were analyzed biochemically for their photosystem II (PSII) antenna composition under varying light conditions. All antisense plants showed a characteristic low-irradiance-induced increase of their PSII antenna size as determined by higher chloro