Christopher D. P. Duffy

The photoprotective molecular switch in the photosystem II antenna.

We have reviewed the current state of multidisciplinary knowledge of the photoprotective mechanism in the photosystem II antenna underlying non-photochemical chlorophyll fluorescence quenching (NPQ). The physiological need for photoprotection of photosystem II and the concept of feed-back control of excess light energy are described. The outline of the major component of nonphotochemical quenching, qE, is suggested to comprise four key elements: trigger (ΔpH), site (antenna), mechanics (antenna dynamics) and quencher(s). The current understanding of the identity and role of these qE components is presented. Existing opinions on the involvement of protons, different LHCII antenna complexes, the PsbS protein and different xanthophylls are reviewed. The evidence for LHCII aggregation and macrostructural reorganization of photosystem II and their role in qE are also discussed. The models describing the qE locus in LHCII complexes, the pigments involved and the evidence for structural dynamics within single monomeric antenna complexes are reviewed. We suggest how PsbS and xanthophylls may exert control over qE by controlling the affinity of LHCII complexes for protons with reference to the concepts of hydrophobicity, allostery and hysteresis. Finally, the physics of the proposed chlorophyll-chlorophyll and chlorophyll-xanthophyll mechanisms of energy quenching is explained and discussed. This article is part of a Special Issue entitled: Photosystem II.

Photoprotective Energy Dissipation Involves the Reorganization of Photosystem II Light-Harvesting Complexes in the Grana Membranes of Spinach Chloroplasts

The rapidly reversible macrostructural changes in higher-plant chloroplast thylakoid membrane organization accompanying photoprotective energy dissipation (qE) are studied using freeze-fracture electron and laser confocal microscopy. qE is shown to involve the aggregation of light-harvesting complexes and their segregation from photosystem II. Plants must regulate their use of absorbed light energy on a minute-by-minute basis to maximize the efficiency of photosynthesis and to protect photosystem II (PSII) reaction centers from photooxidative damage. The regulation of light harvesting involves the photoprotective dissipation of excess absorbed light energy in the light-harvesting antenna complexes (LHCs) as heat. Here, we report an investigation into the structural basis of light-harvesting regulation in intact spinach (Spinacia oleracea) chloroplasts using freeze-fracture electron microscopy, combined with laser confocal microscopy employing the fluorescence recovery after photobleaching technique. The results demonstrate that formation of the photoprotective state requires a structural reorganization of the photosynthetic membrane involving dissociation of LHCII from PSII and its aggregation. The structural changes are manifested by a reduced mobility of LHC antenna chlorophyll proteins. It is demonstrated that these changes occur rapidly and reversibly within 5 min of illumination and dark relaxation, are dependent on ΔpH, and are enhanced by the deepoxidation of violaxanthin to zeaxanthin.

Light-harvesting antenna composition controls the macrostructure and dynamics of thylakoid membranes in Arabidopsis

We characterized a set of Arabidopsis mutants deficient in specific light-harvesting proteins, using freeze-fracture electron microscopy to probe the organization of complexes in the membrane and confocal fluorescence recovery after photobleaching to probe the dynamics of thylakoid membranes within intact chloroplasts. The same methods were used to characterize mutants lacking or over-expressing PsbS, a protein related to light-harvesting complexes that appears to play a role in regulation of photosynthetic light harvesting. We found that changes in the complement of light-harvesting complexes and PsbS have striking effects on the photosystem II macrostructure, and that these effects correlate with changes in the mobility of chlorophyll proteins within the thylakoid membrane. The mobility of chlorophyll proteins was found to correlate with the extent of photoprotective non-photochemical quenching, consistent with the idea that non-photochemical quenching involves extensive re-organization of complexes in the membrane. We suggest that a key feature of the physiological function of PsbS is to decrease the formation of ordered semi-crystalline arrays of photosystem II in the low-light state. Thus the presence of PsbS leads to an increase in the fluidity of the membrane, accelerating the re-organization of the photosystem II macrostructure that is necessary for induction of non-photochemical quenching.

Limits on WIMP cross-sections from the NAIAD experiment at the Boulby Underground Laboratory

The NAIAD experiment (NaI Advanced Detector) for WIMP dark matter searches at the Boulby Underground Laboratory (North Yorkshire, UK) ran from 2000 until 2003. A total of 44.9 kg x years of data collected with 2 encapsulated and 4 unencapsulated NaI(Tl) crystals with high light yield were included in the analysis. We present final results of this analysis carried out using pulse shape discrimination. No signal associated with nuclear recoils from WIMP interactions was observed in any run with any crystal. This allowed us to set upper limits on the WIMP-nucleon spin-independent and WIMP-proton spin-dependent cross-sections. The NAIAD experiment has so far imposed the most stringent constraints on the spin-dependent WIMP-proton cross-section.

Economic photoprotection in photosystem II that retains a complete light-harvesting system with slow energy traps

The light-harvesting antenna of higher plant photosystem II has an intrinsic capability for self-defence against intense sunlight. The thermal dissipation of excess energy can be measured as the non-photochemical quenching of chlorophyll fluorescence. It has recently been proposed that the transition between the light-harvesting and self-defensive modes is associated with a reorganization of light-harvesting complexes. Here we show that despite structural changes, the photosystem II cross-section does not decrease. Our study reveals that the efficiency of energy trapping by the non-photochemical quencher(s) is lower than the efficiency of energy capture by the reaction centres. Consequently, the photoprotective mechanism works effectively for closed rather than open centres. This type of defence preserves the exceptional efficiency of electron transport in a broad range of light intensities, simultaneously ensuring high photosynthetic productivity and, under hazardous light conditions, sufficient photoprotection for both the reaction centre and the light-harvesting pigments of the antenna.

Natural light harvesting: principles and environmental trends

Light harvesting in photosynthetic organisms is largely an efficient process. The first steps of the light phase of photosynthesis, capture of light quanta and primary charge separation processes are particularly well-tuned. In plants, these primary events that take place within the photosystems possess remarkable quantum efficiency, reaching 80% and 100% in photosystems II and I respectively. This paper presents a view on the organisation of a natural light harvesting machine—the antenna of the photosystem II of higher plants. It explains the key principles of biological antenna design and the strategies of adaptation to light environment which have evolved over millions of years. This article argues that the high efficiency of the light harvesting antenna and its control are intimately interconnected owing to the molecular design of the pigment–proteins it is built of, enabling high pigment density combined with the long excited-state lifetime. The protein plays the role of a programmed solvent, accommodating high quantities of pigments, while ensuring their orientations and interaction yields are optimised to efficiently transfer energy to the reaction centres, simultaneously avoiding energy losses due to concentration quenching. The minor group of pigments, the xanthophylls, play a central role in the regulation of light harvesting, defining the antenna efficiency and thus its abilities to simultaneously provide energy to photosystem II and protect itself from excess light damage. Xanthophyll hydrophobicity was found to be a key factor controlling chlorophyll efficiency by modulating pigment–pigment and pigment–protein interactions. Xanthophylls also endow the light harvesting antenna with the remarkable ability to memorise photosystem II light exposure—a light counter principle. Indeed, this type of light harvesting regulation displays hysteretic behaviour, typically observed during electromagnetic induction of ferromagnetic materials, the polarization of ferroelectric materials and the deformation of semi-elastic materials. The photosynthetic antenna is thus a magnificent example of how nature utilises the principles of physics to achieve its goal—extremely efficient, robust, autonomic and yet flexible light harvesting.

Excitation Migration, Quenching, and Regulation of Photosynthetic Light Harvesting in Photosystem II

Excitation energy transfer and quenching in LHCII aggregates is considered in terms of a coarse-grained model. The model assumes that the excitation energy transfer within a pigment-protein complex is much faster than the intercomplex excitation energy transfer, whereas the quenching ability is attributed to a specific pigment-protein complex responsible for the nonphotochemical quenching (NPQ). It is demonstrated that the pump-probe experimental data obtained at low excitation intensities for LHCII aggregates under NPQ conditions can be equally well explained at two limiting cases, either describing the excitation kinetics in the migration-limited or in the trap-limited regime. Thus, it is concluded that low excitation conditions do not allow one to unambiguously define the relationship between the mean times of excitation migration and trapping. However, this could be achieved by using high excitation conditions when exciton-exciton annihilation is dominant. In this case it was found that in the trap-limited regime the excitation kinetics in the aggregate should be almost insensitive to the excitation density, meaning that singlet-singlet annihilation has little effect on the NPQ decay kinetics, whereas in the migration-limited case there is a clear intensity dependence. In order to account for the random distribution of the NPQ-traps within the LHCII aggregates, excitation diffusion in a continuous medium with random static traps was considered. This description demonstrates a very good correspondence to the experimental fluorescence kinetics assuming a lamellar (quasi-3D) structure of the antenna characterized by the dimension d=2.4 and therefore justifying the diffusion-limited approach on which the model is based. Using the coarse-grained model to describe the aggregate we estimate one NPQ-trap per 100 monomeric LHCII complexes. Finally we discuss the origin of the traps responsible for excitation quenching under NPQ conditions.

Dissipative pathways in the photosystem-II antenna in plants

The antenna of photosystem II in plants possesses a remarkable functional flexibility, allowing for the photoprotective regulation of light-harvesting in the face of rapid fluctuations in light intensity. Central to this adaptability is the reversible formation of dissipative energy transfer pathways within the antenna that protect the reaction centres from a potentially damaging excess of excitation energy. The exact molecular nature of these pathways and the mechanism by which they form are still open questions within the field of photosynthesis research. We present a review of current knowledge on the subject. We discuss the multi-scale nature of these pathways, how intrinsic structural and electronic changes within individual antenna proteins are coupled to large scale changes in the structure and energetic connectivity of the membrane as a whole. We review the physical properties and likely validity of current competing models of the dissipation mechanism before discussing a recently studied general property of the dissipative pathways--the slow and economic nature of the NPQ quencher. This property reflects the finely-tuned nature of the quenching pathway, i.e., its ability to offer protection to the photosynthetic machinery without compromising normal photosynthetic function.

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 plants 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.

Electronic Spectra of Structurally Deformed Lutein

Quantum chemical calculations have been employed for the investigation of the lowest excited electronic states of lutein, with particular reference to its function within light harvesting antenna complexes of higher plants. Through comparative analysis obtained by using different methods based on gas-phase calculations of the spectra, it was determined that variations in the lengths of the long C-C valence bonds and the dihedral angles of the polyene chain are the dominant factors in determining the spectral properties of Lut 1 and Lut 2 corresponding to the deformed lutein molecules taken from crystallographic data of the major pigment-protein complex of photosystem II. By MNDO-CAS-CI method, it was determined that the two singlet B(u) states of lutein (nominally 1B(u)(-)* and 1B(u)(+)) arise as a result of mixing of the canonical 1B(u)(-) and 1B(u)(+) states of the all-trans polyene due to the presence of the ending rings in lutein. The 1B(u)(-)* state of lutein is optically allowed, while the 1B(u)(-) of a pure all-trans polyene chain is optically forbidden. As demonstrated, the B(u) states are much more sensitive to minor distortions of the conjugated chain due to mixing of the canonical states, resulting in states of poorly defined particle-hole symmetry. Conversely, the A(g) states are relatively robust with respect to geometric distortion, and their respective inversion and particle-hole symmetries remain relatively well-defined.