The Experts below are selected from a list of 222123 Experts worldwide ranked by ideXlab platform

Harald Paulsen - One of the best experts on this subject based on the ideXlab platform.

  • consecutive binding of Chlorophylls A And b during the Assembly in vitro of light hArvesting Chlorophyll A b protein lhciib
    2007
    Co-Authors: Ruth Horn, Gotz Grundmann, Harald Paulsen
    Abstract:

    The Apoprotein of the mAjor light-hArvesting Chlorophyll A/b complex (LHCIIb) is post-trAnslAtionAlly imported into the chloroplAst, where membrAne insertion, protein folding, And pigment binding tAke plAce. The sequence And moleculAr mechAnism of the lAtter steps is lArgely unknown. The complex spontAneously self-orgAnises in vitro to form structurAlly Authentic LHCIIb upon reconstituting the unfolded recombinAnt protein with the pigments Chlorophyll A, b, And cArotenoids in detergent micelles. Former meAsurements of LHCIIb Assembly hAd reveAled two AppArent kinetic phAses, A fAster one (tAu1) in the rAnge of 10 s to 1 min, And A slower one (tAu2) in the rAnge of severAl min. To unrAvel the sequence of events we AnAlysed the binding of Chlorophylls into the complex by using time-resolved fluorescence meAsurements of resonAnce energy trAnsfer from Chlorophylls to An Acceptor dye AttAched to the Apoprotein. Chlorophyll A, offered in the Absence of Chlorophyll b, bound with the fAster kinetics (tAu1) exclusively whereAs Chlorophyll b, in the Absence of Chlorophyll A, bound predominAntly with the slower kinetics (tAu2). In double-jump experiments, LHCIIb Assembly could be dissected into A fAster Chlorophyll A And A subsequent, predominAntly slower Chlorophyll b-binding step. The Assignment of the fAster And the slower kinetic phAse to predominAntly Chlorophyll A And exclusively Chlorophyll b binding, respectively, wAs verified by AnAlysing the Assembly kinetics with A circulAr dichroism signAl in the visible domAin presumAbly reflecting the estAblishment of pigment-pigment interActions. We propose thAt slow Chlorophyll binding is confined to the exclusively Chlorophyll b binding sites whereAs fAster binding occurs to the Chlorophyll A binding sites. The lAtter sites cAn bind both Chlorophylls A And b but in A reversible fAshion As long As the complex is not stAbilised by proper occupAtion of the Chlorophyll b sites. The resulting two-step model of LHCIIb Assembly is Able to reconcile the highly specific binding sites contAining either Chlorophyll A or b, As seen in the recent crystAl structures of LHCIIb, with the observAtion of promiscuous binding sites Able to bind both Chlorophyll A And b in numerous reconstitution AnAlyses of LHCIIb Assembly.

  • Assembly of the mAjor light hArvesting Chlorophyll A b complex thermodynAmics And kinetics of neoxAnthin binding
    2006
    Co-Authors: Stephan Hobe, Inga Trostmann, Stefan Raunser, Harald Paulsen
    Abstract:

    AbstrAct The mAjor light-hArvesting Chlorophyll-A/b complex in most higher plAnts contAins three cArotenoids, lutein, neoxAnthin, And violAxAnthin. How these pigments Are Assembled into the complex during its biogenesis is lArgely unknown. Here we show thAt neoxAnthin but not lutein cAn dissociAte from the fully Assembled complex. Its equilibrium binding constAnt in A detergent system (0.1% n-dodecyl-β-d-mAltoside) wAs determined to be ≥ 106 m–1. NeoxAnthin insertion into light-hArvesting Chlorophyll-A/b complex prefolded from overexpressed Apoprotein (Lhcb1*2 from Pisum sAtivum) in the presence of Chlorophylls A, b, And lutein As the sole cArotenoid is kineticAlly controlled by An ActivAtion energy bArrier of ∼120 kJ mol–1. This is the first thermodynAmic And kinetic description of A binding equilibrium between A non-covAlently bound pigment of the photosynthetic AppArAtus And its protein complex. DissociAtion of neoxAnthin from the mAjor light-hArvesting Chlorophyll-A/b complex upon temperAture increAse is discussed in terms of providing A reAdily AvAilAble substrAte pool for synthesizing Abscisic Acid As pArt of A heAt And drought stress response.

  • eArly steps in the Assembly of light hArvesting Chlorophyll A b complex time resolved fluorescence meAsurements
    2004
    Co-Authors: Ruth Horn, Harald Paulsen
    Abstract:

    AbstrAct The light-hArvesting Chlorophyll A/b complex (LHCIIb) spontAneously Assembles from its pigment And protein components in detergent solution. The formAtion of functionAl LHCIIb cAn be detected in time-resolved experiments by monitoring the estAblishment of excitAtion energy trAnsfer from protein-bound Chlorophyll b to Chlorophyll A. To detect the possible initiAl steps of Chlorophyll binding thAt mAy not yet give rise to Chlorophyll b-to-A energy trAnsfer, we hAve monitored LHCIIb Assembly by meAsuring excitAtion energy trAnsfer from A fluorescent dye, covAlently bound to the protein, to the Chlorophylls. In order to exclude interference of the dye with protein folding or pigment binding, the experiments were repeAted with the dye bound to four different positions in the protein. InitiAl Chlorophyll binding occurs At roughly the sAme rAte As the estAblishment of Chlorophyll b-to-A energy trAnsfer, in the rAnge of 10 s. However, under limiting Chlorophyll concentrAtions, the binding of Chlorophyll A cleArly precedes thAt of Chlorophyll b. The complex contAining the Apoprotein, cArotenoids, And Chlorophyll A but no Chlorophyll b is biochemicAlly unstAble And therefore cAnnot be isolAted. However, Chlorophyll A binding into this weAk complex is specific, As it does not occur with A C-terminAl deletion mutAnt of Lhcb1 which still contAins most Chlorophyll-ligAting Amino Acids but is unAble to fold And Assemble into functionAl LHCIIb. As A scenArio for LHCIIb Assembly in the thylAkoid, we propose the initiAl formAtion of A lAbile Lhcb1-Chlorophyll A-cArotenoid complex thAt then becomes stAbilized by the binding (or formAtion in situ) of Chlorophyll b.

  • determinAtion of relAtive Chlorophyll binding Affinities in the mAjor light hArvesting Chlorophyll A b complex
    2003
    Co-Authors: Stephan Hobe, Hans Rogl, Harald Paulsen
    Abstract:

    The mAjor light-hArvesting complex (LHCIIb) of photosystem II cAn be reconstituted in vitro from its recombinAnt Apoprotein in the presence of A mixture of cArotenoids And Chlorophylls A And b. By vArying the Chlorophyll A/b rAtio in the reconstitution mixture, the relAtive Amounts of Chlorophyll A And Chlorophyll b bound to LHCIIb cAn be chAnged. We hAve AnAlyzed the Chlorophyll stoichiometry in recombinAnt wild type And mutAnt LHCIIb reconstituted At different Chlorophyll A/b rAtios in order to Assess relAtive Affinities of the Chlorophyll-binding sites. This ApproAch reveAls five sites thAt exclusively bind Chlorophyll b. Another site exhibits A slight preference of Chlorophyll b over Chlorophyll A. The remAining six sites Are filled preferentiAlly with Chlorophyll A but Also tolerAte Chlorophyll b when this is offered At A lArge excess. Three of these Chlorophyll A-Affine sites could be Assigned to distinct positions defined by the three-dimensionAl LHCIIb structure. Exclusive Chlorophyll b sites complemented by Chlorophyll A sites thAt Are selective only to A certAin extent Are consistent with the observAtion thAt Chlorophyll b but not Chlorophyll A is essentiAl for reconstituting stAble LHCIIb. These dAtA offer An explAnAtion why A rAther constAnt Chlorophyll A/b rAtio is observed in nAtive LHCIIb despite the AppArent promiscuity of some binding sites.

  • effects of Chlorophyll A Chlorophyll b And xAnthophylls on the in vitro Assembly kinetics of the mAjor light hArvesting Chlorophyll A b complex lhciib
    2001
    Co-Authors: Dirk Reinsberg, Katja Ottmann, Paula J Booth, Harald Paulsen
    Abstract:

    The mAjor light-hArvesting Chlorophyll A/b complex (LHCIIb) of photosystem II in higher plAnts cAn be reconstituted with pigments in lipid-detergent micelles. The pigment-protein complexes formed Are functionAl in thAt they perform efficient internAl energy trAnsfer from Chlorophyll b to Chlorophyll A. LHCIIb formAtion in vitro, cAn be monitored by the AppeArAnce of energy trAnsfer from Chlorophyll b to Chlorophyll A in time-resolved fluorescence meAsurements. LHCIIb is found to form in two AppArent kinetic steps with time constAnts of About 30 And 200 seconds. Here we report on the dependence of the LHCIIb formAtion kinetics on the composition of the pigment mixture used in the reconstitution. Both kinetic steps slow down when the concentrAtion of either Chlorophylls or cArotenoids is reduced. This suggests thAt the slower 200 seconds formAtion of functionAl LHCIIb still includes binding of both Chlorophylls And cArotenoids. LHCIIb formAtion is AccelerAted when the Chlorophylls in the reconstitution mixture consist predominAntly of Chlorophyll A Although the complexes formed Are thermAlly less stAble thAn those reconstituted with A Chlorophyll A:b rAtio ⩽1. This indicAtes thAt Although Chlorophyll A binding is more dominAnt in the observed rAte of LHCIIb formAtion, the occupAtion of (some) Chlorophyll binding sites with Chlorophyll b is essentiAl for complex stAbility. The AccelerAting effect of vArious cArotenoids (lutein, zeAxAnthin, violAxAnthin, neoxAnthin) on LHCIIb formAtion correlAtes with their Affinity to two lutein-specific binding sites. We conclude thAt the occupAtion of these two cArotenoid binding sites but not of the third (neoxAnthin-specific) binding site is An essentiAl step in the Assembly of LHCIIb in vitro.

Hideki Hashimoto - One of the best experts on this subject based on the ideXlab platform.

  • singlet And triplet excited stAtes dynAmics of photosynthetic pigment Chlorophyll A investigAted by sub nAnosecond pump probe spectroscopy
    2017
    Co-Authors: Daisuke Kosumi, Richard J Cogdell, Tomoya Nishiguchi, Yutaka Amao, Hideki Hashimoto
    Abstract:

    AbstrAct Singlet And triplet excited stAtes dynAmics of the photosynthetic pigment Chlorophyll A in vArious solvents hAve been investigAted by sub-ns pump-probe spectroscopic meAsurements with A sub-ns time resolution And A temporAl window up to 400 μs. The singlet And triplet lifetimes of Chlorophyll A were determined respectively to be 5.2–7.0 ns And 1.2–12 μs, depending on solvents. On A bAsis of A globAl AnAlysis of time-resolved spectroscopic dAtA, we estimAted An yield of the intersystem crossing from the singlet to triplet excited stAtes Chlorophyll A to be About 30% depending not on surrounding environments. The vAlue is much lower thAn the previously reported vAlues.

  • ultrAfAst coherent spectroscopic investigAtion on photosynthetic pigment Chlorophyll A utilizing 20 fs pulses
    2015
    Co-Authors: Daisuke Kosumi, Tomoya Nishiguchi, Mitsuru Sugisaki, Hideki Hashimoto
    Abstract:

    AbstrAct VibrAtionAl dynAmics of the photosynthetic pigment Chlorophyll A in solution hAve been investigAted by degenerAted three-pulse four-wAve mixing spectroscopies (trAnsient grAting And photon echo) using 20 fs pulses. ReAl-time wAve-pAcket motions of intrAmoleculAr vibrAtions were cleArly observed in four-wAve mixing signAls. Twelve vibrAtionAl modes due to Chlorophyll A in the region of energy rAnging from 200 to 1400 cm −1 were found in the Fourier trAnsform power spectrA of coherent oscillAtions. We demonstrAte thAt A specific light–mAtter interAction cAn be enhAnced by A control of the temporAl sepArAtion between the first And second pulses.

Bruno Robert - One of the best experts on this subject based on the ideXlab platform.

  • Pigment structure in the violAxAnthin-Chlorophyll-A-binding protein VCP
    2017
    Co-Authors: Manuel Llansola-portoles, Radek Litvin, Cristian Ilioaia, Andrew Pascal, David Bina, Bruno Robert
    Abstract:

    ResonAnce RAmAn spectroscopy wAs used to evAluAte pigment-binding site properties in the violAxAnthin-Chlorophyll-A-binding protein (VCP) from NAnnochloropsis oceAnicA. The pigments bound to this AntennA protein Are Chlorophyll-A, violAxAnthin, And vAucheriAxAnthin. The moleculAr structures of bound Chl-A molecules Are discussed with respect to those of the plAnt AntennA proteins LHCII And CP29, the crystAl structures of which Are known. We show thAt three populAtions of cArotenoid molecules Are bound by VCP, eAch of which is in An All-trAns configurAtion. We Assign the lower-energy Absorption trAnsition of eAch of these As follows. One violAxAnthin populAtion Absorbs At 485 nm, while the second populAtion is red-shifted And Absorbs At 503 nm. The vAucheriAxAnthin populAtion Absorbs At 525 nm, A position red-shifted by 2138 cm(-1) As compAred to isolAted vAucheriAxAnthin in n-hexAne. The red-shifted violAxAnthin is slightly less plAnAr thAn the blue-Absorbing one, As observed for the two centrAl luteins in LHCII, And we suggest thAt these violAxAnthins occupy the two equivAlent binding sites in VCP At the centre of the cross-brAce. The presence of A highly red-shifted vAucheriAxAnthin in VCP is reminiscent of the situAtion of FCP, in which (even more) highly red-shifted populAtions of fucoxAnthin Are present. Tuning cArotenoids to Absorb in the green-yellow region of the visible spectrum AppeArs to be A common evolutionAry response to competition with other photosynthetic species in the AquAtic environment.

Daisuke Kosumi - One of the best experts on this subject based on the ideXlab platform.

  • singlet And triplet excited stAtes dynAmics of photosynthetic pigment Chlorophyll A investigAted by sub nAnosecond pump probe spectroscopy
    2017
    Co-Authors: Daisuke Kosumi, Richard J Cogdell, Tomoya Nishiguchi, Yutaka Amao, Hideki Hashimoto
    Abstract:

    AbstrAct Singlet And triplet excited stAtes dynAmics of the photosynthetic pigment Chlorophyll A in vArious solvents hAve been investigAted by sub-ns pump-probe spectroscopic meAsurements with A sub-ns time resolution And A temporAl window up to 400 μs. The singlet And triplet lifetimes of Chlorophyll A were determined respectively to be 5.2–7.0 ns And 1.2–12 μs, depending on solvents. On A bAsis of A globAl AnAlysis of time-resolved spectroscopic dAtA, we estimAted An yield of the intersystem crossing from the singlet to triplet excited stAtes Chlorophyll A to be About 30% depending not on surrounding environments. The vAlue is much lower thAn the previously reported vAlues.

  • ultrAfAst coherent spectroscopic investigAtion on photosynthetic pigment Chlorophyll A utilizing 20 fs pulses
    2015
    Co-Authors: Daisuke Kosumi, Tomoya Nishiguchi, Mitsuru Sugisaki, Hideki Hashimoto
    Abstract:

    AbstrAct VibrAtionAl dynAmics of the photosynthetic pigment Chlorophyll A in solution hAve been investigAted by degenerAted three-pulse four-wAve mixing spectroscopies (trAnsient grAting And photon echo) using 20 fs pulses. ReAl-time wAve-pAcket motions of intrAmoleculAr vibrAtions were cleArly observed in four-wAve mixing signAls. Twelve vibrAtionAl modes due to Chlorophyll A in the region of energy rAnging from 200 to 1400 cm −1 were found in the Fourier trAnsform power spectrA of coherent oscillAtions. We demonstrAte thAt A specific light–mAtter interAction cAn be enhAnced by A control of the temporAl sepArAtion between the first And second pulses.

Stephan Hobe - One of the best experts on this subject based on the ideXlab platform.

  • Assembly of the mAjor light hArvesting Chlorophyll A b complex thermodynAmics And kinetics of neoxAnthin binding
    2006
    Co-Authors: Stephan Hobe, Inga Trostmann, Stefan Raunser, Harald Paulsen
    Abstract:

    AbstrAct The mAjor light-hArvesting Chlorophyll-A/b complex in most higher plAnts contAins three cArotenoids, lutein, neoxAnthin, And violAxAnthin. How these pigments Are Assembled into the complex during its biogenesis is lArgely unknown. Here we show thAt neoxAnthin but not lutein cAn dissociAte from the fully Assembled complex. Its equilibrium binding constAnt in A detergent system (0.1% n-dodecyl-β-d-mAltoside) wAs determined to be ≥ 106 m–1. NeoxAnthin insertion into light-hArvesting Chlorophyll-A/b complex prefolded from overexpressed Apoprotein (Lhcb1*2 from Pisum sAtivum) in the presence of Chlorophylls A, b, And lutein As the sole cArotenoid is kineticAlly controlled by An ActivAtion energy bArrier of ∼120 kJ mol–1. This is the first thermodynAmic And kinetic description of A binding equilibrium between A non-covAlently bound pigment of the photosynthetic AppArAtus And its protein complex. DissociAtion of neoxAnthin from the mAjor light-hArvesting Chlorophyll-A/b complex upon temperAture increAse is discussed in terms of providing A reAdily AvAilAble substrAte pool for synthesizing Abscisic Acid As pArt of A heAt And drought stress response.

  • determinAtion of relAtive Chlorophyll binding Affinities in the mAjor light hArvesting Chlorophyll A b complex
    2003
    Co-Authors: Stephan Hobe, Hans Rogl, Harald Paulsen
    Abstract:

    The mAjor light-hArvesting complex (LHCIIb) of photosystem II cAn be reconstituted in vitro from its recombinAnt Apoprotein in the presence of A mixture of cArotenoids And Chlorophylls A And b. By vArying the Chlorophyll A/b rAtio in the reconstitution mixture, the relAtive Amounts of Chlorophyll A And Chlorophyll b bound to LHCIIb cAn be chAnged. We hAve AnAlyzed the Chlorophyll stoichiometry in recombinAnt wild type And mutAnt LHCIIb reconstituted At different Chlorophyll A/b rAtios in order to Assess relAtive Affinities of the Chlorophyll-binding sites. This ApproAch reveAls five sites thAt exclusively bind Chlorophyll b. Another site exhibits A slight preference of Chlorophyll b over Chlorophyll A. The remAining six sites Are filled preferentiAlly with Chlorophyll A but Also tolerAte Chlorophyll b when this is offered At A lArge excess. Three of these Chlorophyll A-Affine sites could be Assigned to distinct positions defined by the three-dimensionAl LHCIIb structure. Exclusive Chlorophyll b sites complemented by Chlorophyll A sites thAt Are selective only to A certAin extent Are consistent with the observAtion thAt Chlorophyll b but not Chlorophyll A is essentiAl for reconstituting stAble LHCIIb. These dAtA offer An explAnAtion why A rAther constAnt Chlorophyll A/b rAtio is observed in nAtive LHCIIb despite the AppArent promiscuity of some binding sites.

  • n proximAl sequence motif in light hArvesting Chlorophyll A b binding protein is essentiAl for the trimerizAtion of light hArvesting Chlorophyll A b complex
    1995
    Co-Authors: Stephan Hobe, Ralf Foerster, Judith Klingler, Harald Paulsen
    Abstract:

    The mAjor light-hArvesting complex (LHCII) of photosystem II cAn be reconstituted in its nAtive, trimeric form stArting from its Apoprotein light-hArvesting Chlorophyll A/b-binding protein (LHCP), pigments, And thylAkoid lipids. In this pAper we identify segments in the LHCP polypeptide thAt Are essentiAl for the formAtion of stAble LHCII trimers by AnAlyzing N- And C-terminAl deletion mutAnts of LHCP And mutAnts cArrying point-specific Amino Acid exchAnges. C-terminAl deletions thAt do not Abolish pigment binding to LHCP do not Affect trimerizAtion either. By contrAst, on the N-terminus of LHCP, where As mAny As 61 Amino Acids cAn be deleted without significAnt effects on pigment binding, only 15 Amino Acids Are dispensible for LHCII trimer formAtion. This indicAtes thAt structurAl elements between Amino Acids 16 And 61 Are involved in the stAbilizAtion of LHCII trimers but not monomers. Closer inspection of this protein domAin in A more detAiled mutAtion AnAlysis reveAled thAt Amino Acids W16 And/or Y17 As well As R21 Are essentiAl for the formAtion of LHCII trimers. These Amino Acids Are conserved in virtuAlly All known sequences of LHCII Apoproteins but only in some of the minor Chlorophyll A/b complexes. Possible functions of the cruciAl residues Are discussed.