Carotenoid Excited States Research Articles

Dynamical Simulations of Carotenoid Photoexcited States Using Density Matrix Renormalization Group Techniques.

We present a dynamical simulation scheme to model the highly correlated excited state dynamics of linear polyenes. We apply it to investigate the internal conversion processes of carotenoids following their photoexcitation. We use the extended Hubbard-Peierls model, , to describe the π-electronic system coupled to nuclear degrees of freedom. This is supplemented by a Hamiltonian, , that explicitly breaks both the particle-hole and two-fold rotation symmetries of idealized carotenoid structures. The electronic degrees of freedom are treated quantum mechanically by solving the time-dependent Schrödinger equation using the adaptive time-dependent DMRG (tDMRG) method, while nuclear dynamics are treated via the Ehrenfest equations of motion. By defining adiabatic excited states as the eigenstates of the full Hamiltonian, , and diabatic excited states as eigenstates of , we present a computational framework to monitor the internal conversion process from the initial photoexcited 11Bu+ state to the singlet triplet-pair states of carotenoids. We further incorporate Lanczos-DMRG to the tDMRG-Ehrenfest method to calculate transient absorption spectra from the evolving photoexcited state. We describe in detail the accuracy and convergence criteria for DMRG, and show that this method accurately describes the dynamical processes of carotenoid excited states. We also discuss the effect of the symmetry-breaking term, , on the internal conversion process, and show that its effect on the extent of internal conversion can be described by a Landau-Zener-type transition. This methodological paper is a companion to our more explanatory discussion of carotenoid excited state dynamics in Manawadu, D.; Georges, T. N.; Barford, W. Photoexcited State Dynamics and Singlet Fission in Carotenoids. J. Phys. Chem. A 2023, 127, 1342.

Low-Lying Excited States of Natural Carotenoids Viewed by Ab Initio Methods.

Low-lying excited states of carotenoids (the optically dark 2Ag- and bright 1Bu+) are deeply involved in energy transfer processes in photosynthetic antennas, such as light harvesting and non-photochemical quenching. Though any ab initio modeling of these phenomena has to rely on precise energies of the carotenoid electronic states, the accurate evaluation of these states remains a challenging problem due to their different natures. The paper aims to study the accuracy of the excitation energies of the low-lying excited states of certain open- and closed-chain carotenoids obtained by a state-of-the-art multireference approach for electronic structure calculation. Here, density matrix renormalization group SCF (DMRGSCF) and a perturbative approach based on driven similarity renormalization group second-order multireference perturbation theory (DSRG-MRPT2) were used to treat the static and dynamic correlation, respectively. Nuclear geometries of the electronic states were optimized with DFT-based approaches. It is demonstrated that spin-flip TD-DFT can replace multiconfigurational methods for the geometry optimization of the 2Ag- state but not for the calculation of the excitation energy. Adiabatic excitation energies to the 1Bu+ state were shown to be within a margin of 1000 cm-1 with an appropriate flow parameter value. Adiabatic excitation energies to the 2Ag- state for the open-chain carotenoids lie within a range of experimental values (taking into account the broad range of experimental estimates); for the closed-chain ones, the error does not exceed 2000 cm-1. Ab initio stationary (1Ag- → 1Bu+) and transient (2Ag- → 1Bu+) absorption spectra were modeled for violaxanthin and lycopene, and these spectra showed good agreement with the experimental ones both in terms of the vibronic structure and the transition energies.

Ultrafast laser spectroscopic studies on carotenoids in solution and on those bound to photosynthetic pigment-protein complexes.

Carotenoid excited singlet states, in particular, are typically very short lived. Therefore, time-resolved absorption spectroscopy in the time regime from femtoseconds to sub-milliseconds are required to unravel and understand the complicated relaxation and excitation energy-transfer pathways of carotenoids in solution and in photosynthetic pigment-protein complexes. The focus of this chapter is to explain how to use ultrafast time-resolved absorption spectroscopy in carotenoid research. The importance of a systematic approach to understanding the various carotenoid excited states by using a series of carotenoids with different conjugation lengths and the isomers of carotenoids is also emphasized.

Excited state geometry of β-carotene influenced by environments: the nature and decisive role of solvent revealing by two-dimensional resonance Raman correlation spectroscopy.

Resonance Raman scattering can be used to investigate the ground and excited state information of carotenoid. It is known that the Dushinsky rotation can significantly influence the resonant Raman intensity of β-carotene (β-car). The excited state geometry revealed by the double components feature of the C = C stretching vibrational modes and the environmental dependence of the Raman intensity for each component remain unknown. We explore the influence of environmental factors on the relative intensity of these two C = C stretching vibration modes and perform two-dimensional resonance Raman correlation analysis to reveal the changes on β-car excited state geometry. The results show that the relative wavelength difference between the 0-0 absorption and the excitation is the key factor that decides the intensity ratio of the two components and that the intensity of each mode is modulated by environmental factors. This modulation is closely related to the excited state geometry and dynamics, effective conjugation length, and electron-phonon coupling constant. It also shows that the asynchronous cross-peaks in the two-dimensional resonance Raman correlation spectrum (2DRRCOS) can effectively characterize the degree of the varied electron-phonon coupling with the changing conditions. These results are not only complementary to the research on the excited states of carotenoids but also applicable to investigate the environmental dependence of Raman intensity for a lot of π-conjugated molecules.

A Protein Environment-Modulated Energy Dissipation Channel in LHCII Antenna Complex.

SummaryThe major light-harvesting complex of photosystem II (LHCII) is the main contributor to sunlight energy harvesting in plants. The flexible design of LHCII underlies a photoprotective mechanism whereby this complex switches to a dissipative state in response to high light stress, allowing the rapid dissipation of excess excitation energy (non-photochemical quenching, NPQ). In this work, we locked single LHCII trimers in a quenched conformation after immobilization of the complexes in polyacrylamide gels to impede protein interactions. A comparison of their pigment excited-state dynamics with quenched LHCII aggregates in buffer revealed the presence of a new spectral band at 515 nm arising after chlorophyll excitation. This is suggested to be the signature of a carotenoid excited state, linked to the quenching of chlorophyll singlet excited states. Our data highlight the marked sensitivity of pigment excited-state dynamics in LHCII to structural changes induced by the environment.

Excitation quenching in chlorophyll\u2013carotenoid antenna systems: \u2018coherent\u2019 or \u2018incoherent\u2019

Plants possess an essential ability to rapidly down-regulate light-harvesting in response to high light. This photoprotective process involves the formation of energy-quenching interactions between the chlorophyll and carotenoid pigments within the antenna of Photosystem II (PSII). The nature of these interactions is currently debated, with, among others, ‘incoherent’ or ‘coherent’ quenching models (or a combination of the two) suggested by a range of time-resolved spectroscopic measurements. In ‘incoherent quenching’, energy is transferred from a chlorophyll to a carotenoid and is dissipated due to the intrinsically short excitation lifetime of the latter. ‘Coherent quenching’ would arise from the quantum mechanical mixing of chlorophyll and carotenoid excited state properties, leading to a reduction in chlorophyll excitation lifetime. The key parameters are the energy gap, Updelta varepsilon =varepsilon_{rm Car}-varepsilon_{rm Chl}, and the resonance coupling, J, between the two excited states. Coherent quenching will be the dominant process when -J<Updelta varepsilon <J, i.e., when the two molecules are resonant, while the quenching will be largely incoherent when varepsilon_{rm Chl}>(varepsilon_{rm Car}+J). One would expect quenching to be energetically unfavorable for varepsilon_{rm Chl}<(varepsilon_{rm Car}-J). The actual dynamics of quenching lie somewhere between these limiting regimes and have non-trivial dependencies of both J and Updelta varepsilon. Using the Hierarchical Equation of Motion (HEOM) formalism we present a detailed theoretical examination of these excitation dynamics and their dependence on slow variations in J and Updelta varepsilon. We first consider an isolated chlorophyll–carotenoid dimer before embedding it within a PSII antenna sub-unit (LHCII). We show that neither energy transfer, nor the mixing of excited state lifetimes represent unique or necessary pathways for quenching and in fact discussing them as distinct quenching mechanisms is misleading. However, we do show that quenching cannot be switched ‘on’ and ‘off’ by fine tuning of Updelta varepsilon around the resonance point, Updelta varepsilon =0. Due to the large reorganization energy of the carotenoid excited state, we find that the presence (or absence) of coherent interactions have almost no impact of the dynamics of quenching. Counter-intuitively significant quenching is present even when the carotenoid excited state lies above that of the chlorophyll. We also show that, above a rather small threshold value of J>10,mathrm{cm}^{-1}quenching becomes less and less sensitive to J (since in the window -J<Updelta varepsilon <J the overall lifetime is independent of it). The requirement for quenching appear to be only that J>0. Although the coherent/incoherent character of the quenching can vary, the overall kinetics are likely robust with respect to fluctuations in J and Updelta varepsilon. This may be the basis for previous observations of NPQ with both coherent and incoherent features.

A Flexible LHCII Structure Allows for Fine-Tuning of Excitation Energy Dissipation

The major light-harvesting complex of photosystem II (LHCII) is the main contributor to sunlight energy harvesting in plants. The flexible design of LHCII underlies a photoprotective mechanism whereby this complex switches to a dissipative state in response to high light stress, allowing the rapid dissipation of excess excitation energy (NPQ). In this work, we stabilized the quenched conformation of single LHCII trimers after immobilization of the complexes in polyacrylamide gels to impede protein interactions. A comparison of their pigment excited-state dynamics with quenched LHCII aggregates in buffer revealed the presence of a new spectral band at 515 nm arising after chlorophyll excitation. This is suggested to be the signature of a carotenoid excited state, linked to the quenching of chlorophyll singlet excited states. Our data highlight the marked sensitivity of pigment excited-state dynamics in LHCII to structural changes induced by the environment.

Carotenoid Nuclear Reorganization and Interplay of Bright and Dark Excited States.

We report quantum chemical calculations using multireference perturbation theory (MRPT) with the density matrix renormalization group (DMRG) plus photothermal deflection spectroscopy measurements to investigate the manifold of carotenoid excited states and establish their energies relative to the bright state (S2) as a function of nuclear reorganization. We conclude that the primary photophysics and function of carotenoids are determined by interplay of only the bright (S2) and lowest-energy dark (S1) states. The lowest-lying dark state, far from being energetically distinguishable from the lowest-lying bright state along the entire excited-state nuclear reorganization pathway, is instead computed to be either the second or first excited state depending on what equilibrium geometry is considered. This result suggests that, rather than there being a dark intermediate excited state bridging a non-negligible energy gap from the lowest-lying dark state to the lowest-lying bright state, there is in fact no appreciable energy gap to bridge following photoexcitation. Instead, excited-state nuclear reorganization constitutes the bridge from S2 to S1, in the sense that these two states attain energetic degeneracy along this pathway.

Photoactivation Mechanism, Timing of Protein Secondary Structure Dynamics and Carotenoid Translocation in the Orange Carotenoid Protein.

The orange carotenoid protein (OCP) is a two-domain photoactive protein that noncovalently binds an echinenone (ECN) carotenoid and mediates photoprotection in cyanobacteria. In the dark, OCP assumes an orange, inactive state known as OCPO; blue light illumination results in the red active state, known as OCPR. The OCPR state is characterized by large-scale structural changes that involve dissociation and separation of C-terminal and N-terminal domains accompanied by carotenoid translocation into the N-terminal domain. The mechanistic and dynamic-structural relations between photon absorption and formation of the OCPR state have remained largely unknown. Here, we employ a combination of time-resolved UV–visible and (polarized) mid-infrared spectroscopy to assess the electronic and structural dynamics of the carotenoid and the protein secondary structure, from femtoseconds to 0.5 ms. We identify a hereto unidentified carotenoid excited state in OCP, the so-called S* state, which we propose to play a key role in breaking conserved hydrogen-bond interactions between carotenoid and aromatic amino acids in the binding pocket. We arrive at a comprehensive reaction model where the hydrogen-bond rupture with conserved aromatic side chains at the carotenoid β1-ring in picoseconds occurs at a low yield of <1%, whereby the β1-ring retains a trans configuration with respect to the conjugated π-electron chain. This event initiates structural changes at the N-terminal domain in 1 μs, which allow the carotenoid to translocate into the N-terminal domain in 10 μs. We identified infrared signatures of helical elements that dock on the C-terminal domain β-sheet in the dark and unfold in the light to allow domain separation. These helical elements do not move within the experimental range of 0.5 ms, indicating that domain separation occurs on longer time scales, lagging carotenoid translocation by at least 2 decades of time.

Direct measurement of the excited states of carotenoids in LHCII trimers using fs-stimulated Raman Spectroscopy

Direct measurement of the excited states of carotenoids in LHCII trimers using fs-stimulated Raman Spectroscopy

A new energy transfer channel from carotenoids to chlorophylls in purple bacteria

It is unclear whether there is an intermediate dark state between the S2 and S1 states of carotenoids. Previous two-dimensional electronic spectroscopy measurements support its existence and its involvement in the energy transfer from carotenoids to chlorophylls, but there is still considerable debate on the origin of this dark state and how it regulates the energy transfer process. Here we use ab initio calculations on excited-state dynamics and simulated two-dimensional electronic spectrum of carotenoids from purple bacteria to provide evidence supporting that the dark state may be assigned to a new Ag+ state. Our calculations also indicate that groups on the conjugation backbone of carotenoids may substantially affect the excited-state levels and the energy transfer process. These results contribute to a better understanding of carotenoid excited states.

The nature of self-regulation in photosynthetic light-harvesting antenna.

The photosynthetic apparatus of green plants is well known for its extremely high efficiency that allows them to operate under dim light conditions. On the other hand, intense sunlight may result in overexcitation of the light-harvesting antenna and the formation of reactive compounds capable of 'burning out' the whole photosynthetic unit. Non-photochemical quenching is a self-regulatory mechanism utilized by green plants on a molecular level that allows them to safely dissipate the detrimental excess excitation energy as heat. Although it is believed to take place in the plant's major light-harvesting complexes (LHC) II, there is still no consensus regarding its molecular nature. To get more insight into its physical origin, we performed high-resolution time-resolved fluorescence measurements of LHCII trimers and their aggregates across a wide temperature range. Based on simulations of the excitation energy transfer in the LHCII aggregate, we associate the red-emitting state, having fluorescence maximum at ∼700 nm, with the partial mixing of excitonic and chlorophyll-chlorophyll charge transfer states. On the other hand, the quenched state has a totally different nature and is related to the incoherent excitation transfer to the short-lived carotenoid excited states. Our results also show that the required level of photoprotection in vivo can be achieved by a very subtle change in the number of LHCIIs switched to the quenched state.

Vibronic coupling in the excited-states of carotenoids.

The ultrafast femtochemistry of carotenoids is governed by the interaction between electronic excited states, which has been explained by the relaxation dynamics within a few hundred femtoseconds from the lowest optically allowed excited state S2 to the optically dark state S1. Extending this picture, some additional dark states (3A(g)(-) and 1B(u)(-)) and their interaction with the S2 state have also been suggested to play a major role in the ultrafast deactivation of carotenoids and their properties. Here, we investigate the interaction between such dark and bright electronic excited states of open chain carotenoids, particularly its dependence on the number of conjugated double bonds (N). We focus on the ultrafast wave packet motion on the excited potential surface, which is modified by the interaction between bright and dark electronic states. Such a coupling between electronic states leads to a shift of the vibrational frequency during the excited-state evolution. In this regard, pump-degenerate four-wave mixing (pump-DFWM) is applied to a series of carotenoids with different numbers of conjugated double bonds N = 9, 10, 11 and 13 (neurosporene, spheroidene, lycopene and spirilloxanthin, respectively). Moreover, we demonstrate in a closed-chain carotenoid (lutein) that the coupling strength and therefore the vibrational shift can be tailored by changing the energy degeneracy between the 1B(u)(+) and 1B(u)(-) states via solvent interaction.

Mechanistic aspects of carotenoid biosynthesis.

Carotenoids represent a large class of terpenoids characterized by an extensively conjugated polyene chain. The conjugation system imparts carotenoids with excellent light absorbing properties in the blue-green (450–550 nm) range of the visible spectrum. The light-absorbing properties of carotenoids have been exploited by photosynthetic organisms to extend the range of light absorption by the photosynthetic apparatus beyond that of chlorophyll. 1 Following light absorption the carotenoid excited state undergoes excitation energy transfer to chlorophyll. 2 In addition to their role as accessory pigments, carotenoids protect against excess light by quenching both singlet and triplet state chlorophylls. In plants oxygenated carotenoids, known as xanthophylls, provide additional photoprotective functions by non-photochemical quenching (NPQ) of chlorophyll fluorescence. 2a,3 Carotenoids carry out light independent functions in scavenging peroxyl radicals and preventing oxidative damage particularly against singlet oxygen (1O2).4 Since many organisms and specific animal tissues, such as the macula lutea and the corpus luteum, accumulate large amounts of carotenoids, it was proposed that carotenoids may protect against the damaging effects of oxidative stress in such tissues. 5a,b These properties have prompted much research in the chemopreventative potential of carotenoids. At high oxygen pressures, however, some carotenoids display prooxidative activity4b and some β-carotene formulations have even shown adverse effects in supplementation trials aimed at preventing lung cancer in smokers. 6 The length of the carotenoid polyene chain corresponds to the width of the phospholipid bilayer, which led to the proposal that carotenoids act as “molecular rivets” to stabilize and add rigidity to the phospholipid membrane. 2b,7 The membrane spanning topology could also allow carotenoids to counteract oxidative damage on either side of the membrane. 4b It has also been proposed that membrane-spanning carotenoids can mediate proton transfer across the membrane or serve as transmembrane radical channels. 8 Due to their striking and rich color carotenoids are important floral pigments serving to attract pollinators and seed dispersers. 9 In birds and fish, carotenoids are an important signal of good nutritional condition and are used in ornamental displays as a sign of fitness and to increase sexual attractiveness. 10a,b,c,10d–f Following oxidative cleavage, carotenoids generate apocarotenoid metabolites which serve important signaling and photoreceptive functions. The 11-cis isomers, 11-cis-retinaldehyde, 3,4-didehydro-11-cis-retinaldehyde or 3-hydroxy-11-cis-retinal are used by most animals as a photosensitive moiety coupled to the opsin protein, rhodopsin, cone opsin or melanopsin. 11 These photoreceptor molecules mediate vision and circadian photoentrainment. 12 Bacteria use the light-sensitive carotenoid cleavage product, retinaldehyde, coupled to bacteriorhodopsin and related proteins to transport protons and other ions across the membrane. This ion transport function allows the cell to generate energy, regulate ion balance or sense light. 13a,b,c The acidic forms of several apocarotenoids act as signaling molecules in fungi, plants and vertebrates. The apocarotenoid, trisporic acid, signals mating type in fungi. 14 Plants cleave carotenoids such as 9-cis-neoxanthin to generate the hormone abscisic acid, which plays important roles in inducing seed dormancy, and allowing the plant to adapt to abiotic stress. 15a,b,c Other plant apocarotenoid metabolites, such as strigolactones trigger seed germination of parasitic weeds and inhibit shoot branching. 16 Finally, vertebrates use retinoic acid, a ligand for nuclear receptors to regulate gene transcription in physiological processes that include embryonic development, cell differentiation, and immunity. 17a–c,17d,e Carotenogenesis occurs in all photosynthetic organisms and in some non-photosynthetic bacteria, archaea, protozoa and fungi. Reflecting their ubiquitous presence and pleiotropic roles there are well over 700 different types of carotenoids generated through variations of their pathways of synthesis. 18 There is even recent evidence for the acquisition of carotenogenic enzymes by metazoans through lateral gene transfer from endosymbiotic fungi. 19 Many excellent reviews have focused on the later steps of the carotenoid synthetic pathways and their regulation in bacteria, and plants. 20,21 In this review we concentrate on the mechanisms of carotenoid synthesis by examining the structure and enzymology of enzymes involved in the production of carotenoids starting from the production of isoprenoid precursors.

Excited States of the Inactive and Active Forms of the Orange Carotenoid Protein

The orange carotenoid protein (OCP) is a crucial player in the process of nonphotochemical quenching in a large number of cyanobacteria. This water-soluble protein binds one pigment only, the keto carotenoid 3'-hydroxyechinenone, and needs to be photoactivated by strong (blue-green) light in order to induce energy dissipation within or from the phycobilisome, the main light harvesting system of these organisms. We performed transient-absorption spectroscopy on OCP samples frozen in the inactive and active forms at 77 K. By making use of target analysis we determined the excited state properties of the active form. Our results show that OCP photoactivation modifies the carotenoid excited state energy landscape. More specifically the photoactivated OCP is characterized by one state with predominantly ICT character (ICT/S1) and a lifetime of 2.3 ps, and another state with mainly S1 character (S1/ICT) with a lifetime of 7.6 ps. We also show that the kinetic model is fully consistent with the RT data obtained earlier (Berera et al., J. Phys. Chem. B 2012, 116, 2568-2574). We propose that this ICT/S1 state acts as the quencher in the OCP mediated nonphotochemical quenching.

The Photophysics of the Orange Carotenoid Protein, a Light-Powered Molecular Switch

To cope with the deleterious effects of excess illumination, photosynthetic organisms have developed photoprotective mechanisms that dissipate the absorbed excess energy as heat from the antenna system. In cyanobacteria, a crucial step in the process is the activation, by blue-green light, of a soluble protein, known as orange carotenoid protein (OCP), which binds the carotenoid 3'-hydroxyechinenone as its only pigment. While the spectroscopic properties of the inactive form of OCP have been described, the nature of the excited states in the active form still awaits elucidation. We applied transient absorption spectroscopy to the dark and the light activated forms of OCP to study and compare the excited state dynamics of both forms. We show that excitation of the photoactivated OCP leads to the population of new carotenoid excited states. One of these states populated shortly after excitation is characterized by a very pronounced charge transfer character and a lifetime of about 0.6 ps. When the illuminated sample is exposed to a dark relaxation period, it responds to excitation as the original dark sample, showing that photoactivation and decay of the photoactivated state are fully reversible. Thus OCP functions as a light-powered molecular switch that modulates its spectroscopic properties as a response to specific changes in light environment. We discuss the importance of this switch in cyanobacteria photoprotection and propose a mechanism wherein the red state of OCP echinenone acts as an energy dissipator via its charge transfer state.

Spectroscopic Studies of Carotenoid-to-Bacteriochlorophyll Energy Transfer in LHRC Photosynthetic Complex from Roseiflexus castenholzii 1 Resubmitted to J Phys Chem B.

Carotenoids present in the photosynthetic light-harvesting reaction center (LHRC) complex from chlorosome lacking filamentous anoxygenic phototroph, Roseiflexus castenholzii were purified and characterized for their photochemical properties. The LHRC from anaerobically grown cells contains five different carotenoids, methoxy-keto-myxocoxanthin, gamma-carotene, and its three derivatives, whereas the LHRC from aerobically grown cells contains only three carotenoid pigments with methoxy-keto-myxocoxanthin being the dominant one. The spectroscopic properties and dynamics of excited singlet states of the carotenoids were studied by steady-state absorption, fluorescence and ultrafast time-resolved optical spectroscopy in organic solvent and in the intact LHRC complex. Time-resolved transient absorption spectroscopy performed in the near-infrared (NIR) on purified carotenoids combined with steady-state absorption spectroscopy led to the precise determination of values of the energies of the S(1)(2(1)A(g)(-)) excited state. Global and single wavelength fitting of the ultrafast spectral and temporal data sets of the carotenoids in solvents and in the LHRC revealed the pathways of de-excitation of the carotenoid excited states.

A Mechanism of Energy Dissipation in Cyanobacteria

When grown under a variety of stress conditions, cyanobacteria express the isiA gene, which encodes the IsiA pigment-protein complex. Overexpression of the isiA gene under iron-depletion stress conditions leads to the formation of large IsiA aggregates, which display remarkably short fluorescence lifetimes and thus a strong capacity to dissipate energy. In this work we investigate the underlying molecular mechanism responsible for chlorophyll fluorescence quenching. Femtosecond transient absorption spectroscopy allowed us to follow the process of energy dissipation in real time. The light energy harvested by chlorophyll pigments migrated within the system and eventually reaches a quenching site where the energy is transferred to a carotenoid-excited state, which dissipates it by decaying to the ground state. We compare these findings with those obtained for the main light-harvesting complex in green plants (light-harvesting complex II) and artificial light-harvesting antennas, and conclude that all of these systems show the same mechanism of energy dissipation, i.e., one or more carotenoids act as energy dissipators by accepting energy via low-lying singlet-excited S 1 states and dissipating it as heat.

Ultrafast time-resolved carotenoid to-bacteriochlorophyll energy transfer in LH2 complexes from photosynthetic bacteria.

Steady-state and ultrafast time-resolved optical spectroscopic investigations have been carried out at 293 and 10 K on LH2 pigment-protein complexes isolated from three different strains of photosynthetic bacteria: Rhodobacter (Rb.) sphaeroides G1C, Rb. sphaeroides 2.4.1 (anaerobically and aerobically grown), and Rps. acidophila 10050. The LH2 complexes obtained from these strains contain the carotenoids, neurosporene, spheroidene, spheroidenone, and rhodopin glucoside, respectively. These molecules have a systematically increasing number of pi-electron conjugated carbon-carbon double bonds. Steady-state absorption and fluorescence excitation experiments have revealed that the total efficiency of energy transfer from the carotenoids to bacteriochlorophyll is independent of temperature and nearly constant at approximately 90% for the LH2 complexes containing neurosporene, spheroidene, spheroidenone, but drops to approximately 53% for the complex containing rhodopin glucoside. Ultrafast transient absorption spectra in the near-infrared (NIR) region of the purified carotenoids in solution have revealed the energies of the S1 (2(1)Ag-)-->S2 (1(1)Bu+) excited-state transitions which, when subtracted from the energies of the S0 (1(1)Ag-)-->S2 (1(1)Bu+) transitions determined by steady-state absorption measurements, give precise values for the positions of the S1 (2(1)Ag-) states of the carotenoids. Global fitting of the ultrafast spectral and temporal data sets have revealed the dynamics of the pathways of de-excitation of the carotenoid excited states. The pathways include energy transfer to bacteriochlorophyll, population of the so-called S* state of the carotenoids, and formation of carotenoid radical cations (Car*+). The investigation has found that excitation energy transfer to bacteriochlorophyll is partitioned through the S1 (1(1)Ag-), S2 (1(1)Bu+), and S* states of the different carotenoids to varying degrees. This is understood through a consideration of the energies of the states and the spectral profiles of the molecules. A significant finding is that, due to the low S1 (2(1)Ag-) energy of rhodopin glucoside, energy transfer from this state to the bacteriochlorophylls is significantly less probable compared to the other complexes. This work resolves a long-standing question regarding the cause of the precipitous drop in energy transfer efficiency when the extent of pi-electron conjugation of the carotenoid is extended from ten to eleven conjugated carbon-carbon double bonds in LH2 complexes from purple photosynthetic bacteria.

Identification of a mechanism of photoprotective energy dissipation in higher plants

Under conditions of excess sunlight the efficient light-harvesting antenna found in the chloroplast membranes of plants is rapidly and reversibly switched into a photoprotected quenched state in which potentially harmful absorbed energy is dissipated as heat, a process measured as the non-photochemical quenching of chlorophyll fluorescence or qE. Although the biological significance of qE is established, the molecular mechanisms involved are not. LHCII, the main light-harvesting complex, has an inbuilt capability to undergo transformation into a dissipative state by conformational change and it was suggested that this provides a molecular basis for qE, but it is not known if such events occur in vivo or how energy is dissipated in this state. The transition into the dissipative state is associated with a twist in the configuration of the LHCII-bound carotenoid neoxanthin, identified using resonance Raman spectroscopy. Applying this technique to study isolated chloroplasts and whole leaves, we show here that the same change in neoxanthin configuration occurs in vivo, to an extent consistent with the magnitude of energy dissipation. Femtosecond transient absorption spectroscopy, performed on purified LHCII in the dissipative state, shows that energy is transferred from chlorophyll a to a low-lying carotenoid excited state, identified as one of the two luteins (lutein 1) in LHCII. Hence, it is experimentally demonstrated that a change in conformation of LHCII occurs in vivo, which opens a channel for energy dissipation by transfer to a bound carotenoid. We suggest that this is the principal mechanism of photoprotection.