Photosynthetic light-harvesting antennae are pigment-binding proteins that perform one of the most fundamental tasks on Earth, capturing light and transferring energy that enables life in our biosphere. Adaptation to different light environments led to the evolution of an astonishing diversity of light-harvesting systems. At the same time, several strategies have been developed to optimize the light energy input into photosynthetic membranes in response to fluctuating conditions. The basic feature of these prompt responses is the dynamic nature of antenna complexes, whose function readily adapts to the light available. High-resolution microscopy and spectroscopic studies on membrane dynamics demonstrate the crosstalk between antennae and other thylakoid membrane components. With the increased understanding of light-harvesting mechanisms and their regulation, efforts are focusing on the development of sustainable processes for effective conversion of sunlight into functional bio-products. The major challenge in this approach lies in the application of fundamental discoveries in light-harvesting systems for the improvement of plant or algal photosynthesis. Here, we underline some of the latest fundamental discoveries on the molecular mechanisms and regulation of light harvesting that can potentially be exploited for the optimization of photosynthesis.

Low-temperature fluorescence measurements are frequently used in photosynthesis research to assess photosynthetic processes. Upon illumination of photosystem II (PSII) frozen to 77K, fluorescence quenching is observed. In this work, we studied the light-induced quenching in intact cells of Chlamydomonas reinhardtii at 77K using time-resolved fluorescence spectroscopy with a streak camera setup. In agreement with previous studies, global analysis of the data shows that prolonged illumination of the sample affects the nanosecond decay component of the PSII emission. Using target analysis, we resolved the quenching on the PSII-684 compartment which describes bulk chlorophyll molecules of the PSII core antenna. Further, we quantified the quenching rate constant and observed that as the illumination proceeds the accumulation of the quencher leads to a speed up of the fluorescence decay of the PSII-684 compartment as the decay rate constant increases from about 3 to 4ns(-1). The quenching on PSII-684 leads to indirect quenching of the compartments PSII-690 and PSII-695 which represent the red chlorophyll of the PSII core. These results explain past and current observations of light-induced quenching in 77K steady-state and time-resolved fluorescence spectra.

The efficient use of excitation energy in photosynthetic membranes is achieved by a dense network of pigment–protein complexes. These complexes fulfill specific functions and interact dynamically with each other in response to rapidly changing environmental conditions. Here, we studied how in the intact cells of Chlamydomonas reinhardtii (C.r.) the lack of the photosystem I (PSI) core or the photosystem II (PSII) core affects these interactions. To that end the mutants F15 and M18 (both PSI-deficient) and FUD7 (PSII-deficient) were incubated under conditions known to promote state transitions in wild-type. The intact cells were then instantly frozen to 77 K and the full-spectrum time-resolved fluorescence emission of the cells was measured by means of streak camera. In the PSI-deficient mutants excitation energy transfer (EET) towards light-harvesting complexes of PSI (Lhca) occurs in less than 0.5 ns, and fluorescence from Lhca decays in 3.1 ns. Decreased trapping by PSII and increased fluorescence of Lhca upon state 1 (S1) → state 2 (S2) transition appears in the F15 and less in the M18 mutant. In the PSII-deficient mutant FUD7, quenched (0.5 ns) and unquenched (2 ns) light-harvesting complexes of PSII (LHCII) are present in both states, with the quenched form more abundant in S2 than in S1. Moreover, EET of 0.4 ns from the remaining LHCII to PSI increases upon S1 → S2 transition. We relate the excitation energy kinetics observed in F15, M18 and FUD7 to the remodeling of the photosynthetic apparatus in these mutants under S1 and S2 conditions.

The regulatory mechanism of state transitions was studied in Chlamydomonas reinhardtii (C.r.) wild type (WT) as well as mutant strains deficient in the photosystem I (PSI) or the photosystem II (PSII) core. Time-resolved fluorescence measurements were obtained on instantly frozen cells incubated beforehand in the dark in aerobic or anaerobic conditions which leads to state 1 (S1) or state 2 (S2). WT data contains information on the light-harvesting complex (LHC) connected to PSI and PSII. The mutants' data contain information on either LHCII-LHCI-PSI or LHCII-PSII, plus information on LHC antennas devoid of a PS core. In a simultaneous analysis of the data from all strains under S1 or S2 conditions a unified model for the excited state dynamics at 77K was created. This yielded the completely resolved LHCII-LHCI-PSI and LHCII-PSII dynamics and quantified the state transitions. In WT cells the fraction of light absorbed by LHCII connected to PSII decreases from 45% in S1 to 29% in S2, while it increases from 0% to 16% for LHCII connected to PSI. Thus (16/45=) 36% of all LHCII is involved in the state transition. In the mutant strains deficient in the PSI core, the red most species peaking at 716nm disappears completely, indicating that this far red Chl pigment is located in the PSI core. In the mutant strain deficient in the PSII core, red shifted species with maxima at 684 and 686nm appear in the LHCII antenna. LHCII-684 is quenched and decays with a rate of (310ps)−1.

This article is concerned with the emergence of the science–business (SB) profession and the role of the SB professional (SBP) in both academia and business. Transferring science to markets can take a variety of routes depending on the stage of development and the people and skill sets involved. An SBP may co-develop opportunities for exploitation as part of a team in a university or in the business world in a spin-off or a mature company. In both settings, the SBP connects with scientists and co-develops knowledge or technology into an exploitable commodity. The need for this type of professional was the starting point for a new curriculum: a 3-year bachelor’s programme in science, business and innovation that integrates fundamental sciences (physics, chemistry, mathematics), life and health sciences, energy and sustainability sciences, business/social sciences, innovation and entrepreneurship. The authors provide the context and rationale for the development of the programme and discuss how it meets its objective to address a critical skills need in both academic and business sectors.

State transitions in the green alga Chlamydomonas reinhardtii serve to balance excitation energy transfer to photosystem I (PSI) and to photosystem II (PSII) and possibly play a role as a photoprotective mechanism. Thus, light-harvesting complex II (LHCII) can switch between the photosystems consequently transferring more excitation energy to PSII (state 1) or to PSI (state 2) or can end up in LHCII-only domains. In this study, low-temperature (77 K) steady-state and time-resolved fluorescence measured on intact cells of Chlamydomonas reinhardtii shows that independently of the state excitation energy transfer from LHCII to PSI or to PSII occurs on two main timescales of <15 ps and ∼ 100 ps. Moreover, in state 1 almost all LHCIIs are functionally connected to PSII, whereas the transition from state 1 to a state 2 chemically locked by 0.1 M sodium fluoride leads to an almost complete functional release of LHCIIs from PSII. About 2/3 of the released LHCIIs transfer energy to PSI and ∼ 1/3 of the released LHCIIs form a component designated X-685 peaking at 685 nm that decays with time constants of 0.28 and 5.8 ns and does not transfer energy to PSI or to PSII. A less complete state 2 was obtained in cells incubated under anaerobic conditions without chemical locking. In this state about half of all LHCIIs remained functionally connected to PSII, whereas the remaining half became functionally connected to PSI or formed X-685 in similar amounts as with chemical locking. We demonstrate that X-685 originates from LHCII domains not connected to a photosystem and that its presence introduces a change in the interpretation of 77 K steady-state fluorescence emission measured upon state transitions in Chalamydomonas reinhardtii.

A characteristic feature of the active Photosystem II (PSII) complex is a red-shifted low temperature fluorescence emission at about 693nm. The origin of this emission has been attributed to a monomeric 'red' chlorophyll molecule located in the CP47 subunit. However, the identity and function of this chlorophyll remain uncertain. In our previous work, we could not detect the red PSII emission in a mutant of the cyanobacterium Synechocystis sp. PCC 6803 lacking PsbH, a small transmembrane subunit bound to CP47. However, it has not been clear whether the PsbH is structurally essential for the red emission or the observed effect of mutation has been indirectly caused by compromised PSII stability and function. In the present work we performed a detailed spectroscopic characterization of PSII in cells of a mutant lacking PsbH and Photosystem I and we also characterized PSII core complexes isolated from this mutant. In addition, we purified and characterized the CP47 assembly modules containing and lacking PsbH. The results clearly confirm an essential role of PsbH in the origin of the PSII red emission and also demonstrate that PsbH stabilizes the binding of one β-carotene molecule in PSII. Crystal structures of the cyanobacterial PSII show that PsbH directly interacts with a single monomeric chlorophyll ligated by the histidine 114 residue of CP47 and we conclude that this peripheral chlorophyll hydrogen-bonded to PsbH is responsible for the red fluorescence state of CP47. Given the proximity of β-carotene this state could participate in the dissipation of excessive light energy.

When cyanobacteria are grown under iron-limited or other oxidative stress conditions the iron stress inducible pigment-protein IsiA is synthesized in variable amounts. IsiA accumulates in aggregates inside the photosynthetic membrane that strongly dissipate chlorophyll excited state energy. In this paper we applied Stark fluorescence (SF) spectroscopy at 77K to IsiA aggregates to gain insight into the nature of the emitting and energy dissipating state(s). Our study shows that two emitting states are present in the system, one emitting at 684 nm and the other emitting at about 730 nm. The new 730 nm state exhibits strongly reduced fluorescence (F) together with a large charge transfer character. We discuss these findings in the light of the energy dissipation mechanisms involved in the regulation of photosynthesis in plants, cyanobacteria and diatoms. Our results suggest that photosynthetic organisms have adopted common mechanisms to cope with the deleterious effects of excess light under unfavorable growth conditions.

To investigate the dynamics of photosynthetic pigment-protein complexes in vascular plants at high resolution in an aqueous environment, membrane-protruding oxygen-evolving complexes (OECs) associated with photosystem II (PSII) on spinach (Spinacia oleracea) grana membranes were examined using contact mode atomic force microscopy. This study represents, to our knowledge, the first use of atomic force microscopy to distinguish the putative large extrinsic loop of Photosystem II CP47 reaction center protein (CP47) from the putative oxygen-evolving enhancer proteins 1, 2, and 3 (PsbO, PsbP, and PsbQ) and large extrinsic loop of Photosystem II CP43 reaction center protein (CP43) in the PSII-OEC extrinsic domains of grana membranes under conditions resulting in the disordered arrangement of PSII-OEC particles. Moreover, we observed uncharacterized membrane particles that, based on their physical characteristics and electrophoretic analysis of the polypeptides associated with the grana samples, are hypothesized to be a domain of photosystem I that protrudes from the stromal face of single thylakoid bilayers. Our results are interpreted in the context of the results of others that were obtained using cryo-electron microscopy (and single particle analysis), negative staining and freeze-fracture electron microscopy, as well as previous atomic force microscopy studies.

In purple bacteria of the genus Rhodobacter (Rba.), an LH1 antenna complex surrounds the photochemical reaction centre (RC) with a PufX protein preventing the LH1 complex from completely encircling the RC. In membranes of Rba. sphaeroides, RC–LH1 complexes associate as dimers which in turn assemble into longer range ordered arrays. The present work uses linear dichroism (LD) and dark-minus-light difference LD (ΔLD) to probe the organisation of genetically altered RC–LH1 complexes in intact membranes. The data support previous proposals that Rba. capsulatus, and Rba. sphaeroides heterologously expressing the PufX protein from Rba. capsulatus, produce monomeric core complexes in membranes that lack long-range order. Similarly, Rba. sphaeroides with a point mutation in the Gly 51 residue of PufX, which is located on the membrane-periplasm interface, assembles mainly non-ordered RC–LH1 complexes that are most likely monomeric. All the Rba. sphaeroides membranes in their ΔLD spectra exhibited a spectral fingerprint of small degree of organisation implying the possibility of ordering influence of LH1, and leading to an important conclusion that PufX itself has no influence on ordering RC–LH1 complexes, as long-range order appears to be induced only through its role of configuring RC–LH1 complexes into dimers.

We report for the first time steady-state and time-resolved emission properties of photosystem I (PSI) complexes isolated from the cyanobacterial strain Synechococcus WH 7803. The PSI complexes from this strain display an extremely small fluorescence emission yield at 77 K, which we attribute to the absence of so-called red antenna chlorophylls, chlorophylls with absorption maxima at wavelengths longer than those of the primary electron donor P700. Emission measurements at room temperature with picosecond time resolution resulted in two main decay components with lifetimes of about 7.5 and 18 ps and spectra peaking at about 685 nm. Especially in the red flanks, these spectra show consistent differences, which means that earlier proposed models for the primary charge separation reactions based on ultrafast (∼1 ps) excitation equilibration processes cannot describe the data. We show target analyses of a number of alternative models and conclude that a simple model (Ant2)* ↔ (Ant1/RC)* → RP2 can explain the time-resolved emission data very well. In this model, (Ant2)* represents chlorophylls that spectrally equilibrate in about 7.5 ps and in which RP2 represents the "final" radical pair P700(+)A0(-). Adding an equilibrium (Ant1/RC)* ↔ RP1, in which RP1 represents an "intermediate" radical pair A(+)A0(-), resulted in the same fit quality. We show that the simple model without RP1 can easily be extended to PSI complexes from cyanobacteria with one or more pools of red antenna chlorophylls and also that the model provides a straightforward explanation of steady-state emission properties observed at cryogenic temperatures.

Time-resolved fluorescence spectroscopy measurements at 77 K on thylakoid membrane preparations and isolated photosynthetic complexes thereof were investigated using target analysis with the aim of building functional compartmental models for the photosystems in the thylakoid membrane. Combining kinetic schemes with different spectral constraints enabled us to resolve the energy transfer pathways and decay characteristics of the different emissive species. We determined the spectral and energetic properties of the red Chl pools in both photosystems and quantified the formation of LHCII-LHCI-PSI supercomplexes in the transition from native to unstacked thylakoid membranes.

We present a thorough spectroscopic investigation of the role of carotenoids in energy transfer and chlorophyll fluorescence quenching in aggregates of the IsiA pigment-protein complex of the cyanobacterium Synechocystis PCC 6803. The non-selective excitation of the carotenoids S0→S2 transition at 475 nm yields a carotenoid to chlorophyll energy transfer efficiency of ~20%. When the carotenoids are excited in the red wing of the absorption spectrum of the S0→S2 transition at 530 nm the energy transfer efficiency increases to ~30 %, demonstrating that a ‘red’ carotenoid is active in energy transfer to the chlorophyll(s). In both cases energy transfer takes place on the sub 100 fs timescale from the carotenoids S2 state only. Results from excitation of chlorophyll a at 675 nm strongly suggest the involvement of a carotenoid molecule in the quenching of chlorophyll excited state energy. We propose that one of the carotenoids acts as an energy sink by accepting energy from the excited chlorophyll via a low-lying dark excited state. These findings are discussed in the light of recent results obtained in other natural and artificial light-harvesting antennas and point to a general mechanism of photoprotection in photosynthesis.

We investigated the electronic structure of the photosystem II reaction center (PSII RC) in relation to the light-induced charge separation process using Stark spectroscopy on a series of site-directed PSII RC mutants from the cyanobacterium Synechocystis sp. PCC 6803. The site-directed mutations modify the protein environment of the cofactors involved in charge separation (PD1, PD2, ChlD1, and PheD1). The results demonstrate that at least two different exciton states are mixed with charge-transfer (CT) states, yielding exciton states with CT character: (PD2δ+PD1δ−ChlD1)∗673nm and (ChlD1δ+PheD1δ−)∗681nm (where the subscript indicates the wavelength of the electronic transition). Moreover, the CT state PD2+PD1− acquires excited-state character due to its mixing with an exciton state, producing (PD2+PD1−)δ∗684nm. We conclude that the states that initiate charge separation are mixed exciton-CT states, and that the degree of mixing between exciton and CT states determines the efficiency of charge separation. In addition, the results reveal that the pigment-protein interactions fine-tune the energy of the exciton and CT states, and hence the mixing between these states. This mixing ultimately controls the selection and efficiency of a specific charge separation pathway, and highlights the capacity of the protein environment to control the functionality of the PSII RC complex.

The thylakoid membrane system is a complex membrane system that organizes and reorganizes itself to provide plants optimal chemical energy from sunlight under different and varying environmental conditions. Grana membranes are part of this system and contain the light-driven water-splitting enzyme Photosystem II (PSII) and light-harvesting antenna complexes. Here, we present a direct visualization of PSII complexes within grana membranes from spinach. By means of jumping mode atomic force microscopy in liquid, minimal forces were applied between the scanning tip and membrane or protein, allowing complexes to be imaged with high detail. We observed four different packing arrangements of PSII complexes, which occur primarily as dimers: co-linear crystalline rows, nanometric domains of straight or skewed rows, and disordered domains. Upon storing surface-adhered membranes at low temperature prior to imaging, large-scale reorganizations of supercomplexes between PSII and light-harvesting complex II could be induced. The highest resolution images show the existence of membrane domains without obvious topography extending beyond supercomplexes. These observations illustrate the possibility for diffusion of proteins and smaller molecules within these densely packed membranes.

We explain the transient absorption kinetics (E. Romero, I. H. M. van Stokkum, V. I. Novoderezhkin, J. P. Dekker, R. van Grondelle, Biochemistry 2010, 49, 4300) measured for isolated reaction centers of photosystem II at 77 K upon excitation of the primary donor band (680 nm). The excited-state dynamics is modeled on the basis of the exciton states of 6 cofactors coupled to 4 charge-transfer (CT) states. One CT state (corresponding to charge separation within the special pair) is supposed to be strongly coupled with the excited states, whereas the other radical pairs are supposed to be localized. Relaxation within the strongly coupled manifold and transfer to localized CT's are described by the modified Redfield and generalized Förster theories, respectively. A simultaneous and quantitative fit of the 680, 545, and 460 nm kinetics (corresponding to respectively the Q(y) transitions of the red-most cofactors, Q(x) transition of pheophytin, and pheophytin anion absorption) enables us to define the pathways and time scales of primary electron transfer. A consistent modeling of the data is only possible with a Scheme where charge separation occurs from both the accessory chlorophyll and from the special pair, giving rise to fast and slow components of the pheophytin anion formation, respectively.