In bright sunlight, the amount of energy harvested by plants exceeds the electron transport capacity of Photosystem II in the chloroplasts. The excess energy can lead to severe damage of the photosynthetic apparatus and to avoid this, part of the energy is thermally dissipated via a mechanism called non-photochemical quenching (NPQ). It has been found that LHCII, the major antenna complex of Photosystem II, is involved in this mechanism and it was proposed that its quenching site is formed by the cluster of strongly interacting pigments: chlorophylls 611 and 612 and lutein 620 [A.V. Ruban, R. Berera, C. Ilioaia, I.H.M. van Stokkum, J.T.M. Kennis, A.A. Pascal, H. van Amerongen, B. Robert, P. Horton and R. van Grondelle, Identification of a mechanism of photoprotective energy dissipation in higher plants, Nature 450 (2007) 575-578.]. In the present work we have investigated the interactions between the pigments in this cluster not only for LHCII, but also for the homologous minor antenna complexes CP24, CP26 and CP29. Use was made of wild-type and mutated reconstituted complexes that were analyzed with (low-temperature) absorption and circular-dichroism spectroscopy as well as by biochemical methods. The pigments show strong interactions that lead to highly specific spectroscopic properties that appear to be identical for LHCII, CP26 and CP29. The interactions are similar but not identical for CP24. It is concluded that if the 611/612/620 domain is responsible for the quenching in LHCII, then all these antenna complexes are prepared to act as a quencher. This can explain the finding that none of the Lhcb complexes seems to be strictly required for NPQ while, in the absence of all of them, NPQ is abolished.

Lhcb5 is an antenna protein that is highly conserved in plants and green algae. It is part of the inner layer of photosystem II antenna system retained in high light acclimated plants. To study the structure-function relation and the role of individual pigments in this complex, we (i) "knocked out" each of the chromophores bound to multiple (nine total) chlorophyll sites and (ii) exchanged the xanthophylls bound to the three xanthophyll sites. The occupancy and associated energy of the pigment binding sites were determined. The role of the individual pigments in protein folding, stability, energy transfer, and dissipation was studied in vitro. The results indicate that lutein has a primary role in the folding and stability of the complex, whereas violaxanthin and zeaxanthin have a negative effect on folding yield and stability, respectively. The data showed a distinct function for the L1 and L2 carotenoid binding sites, the former preferentially involved in gathering the excitation energy to chlorophyll a (Chl a), whereas the latter modulates the concentration of chlorophyll singlet excited states dependent on the xanthophylls bound to it, likely via an interaction with Chl-603. Our results also underscored the role of zeaxanthin and lutein in quenching the excitation energy, whereas violaxanthin was shown to be very effective in energy transfer. The characteristics of the isolated proteins were consistent with the observed role of Lhcb5 in vivo in catalyzing fluorescence quenching upon zeaxanthin binding.Sunlight energy is absorbed and converted into chemical energy by pigments bound to photosystems I and II (PSI 2 and PSII). Photosystems are composed of two moieties: (i) a reaction center devoted to the conversion of light energy into chemical energy and (ii) an antenna system that increases the capacity of light absorption and contributes to photoprotection (1). In photosynthetic eukaryotes, the antenna system is composed of members of a multigenic family called light-harvesting complexes (Lhc). In higher plants, 10 different subunits are associated with the photosystems, Lhca1-4 with PSI and Lhcb1-6 with PSII (2). All of these proteins share the same evolutionary origin and a common structural organization (3). Only the structure of LHCII, a trimer composed mainly of Lhcb1 (4), has been resolved at the atomic level, revealing the presence of three transmembrane and two amphipathic helices named, respectively, A-C and D-E (5). Each Lhcb1 polypeptide has been shown to coordinate four carotenoids and 14 Chl molecules. Two carotenoid binding sites, L1 and L2, 3 are close to helices A and B, respectively (6), and a third site, N1, specific for neoxanthin, is located in proximity to helix C (7). Finally, a peripheral and less stable binding site, V1, has been shown to accommodate violaxanthin and zeaxanthin (5,8,9).Other members of the Lhc protein family coordinate a variable number of pigments (8 -14 Chl and 2-4 carotenoid molecules, depending on the polypeptide (9 -14)) implying that some of the binding sites id...

In this work the photoprotective role of all xanthophylls in LHCII, Lhcb4, and Lhcb5 is investigated by laser-induced Triplet-minus-Singlet (TmS) spectroscopy. The comparison of native LHCII trimeric complexes with different carotenoid composition shows that the xanthophylls in sites V1 and N1 do not directly contribute to the chlorophyll triplet quenching. The largest part of the triplets is quenched by the lutein bound in site L1, which is located in close proximity to the chlorophylls responsible for the low energy state of the complex. The lutein in the L2 site is also active in triplet quenching, and it shows a longer triplet lifetime than the lutein in the L1 site. This lifetime difference depends on the occupancy of the N1 binding site, where neoxanthin acts as an oxygen barrier, limiting the access of O 2 to the inner domain of the Lhc complex, thereby strongly contributing to the photostability. The carotenoid triplet decay of monomeric Lhcb1, Lhcb4, and Lhcb5 is mono-exponential, with shorter lifetimes than observed for trimeric LHCII, suggesting that their inner domains are more accessible for O 2 . As for trimeric LHCII, only the xanthophylls in sites L1 and L2 are active in triplet quenching. Although the chlorophyll to carotenoid triplet transfer is efficient (95%) in all complexes, it is not perfect, leaving 5% of the chlorophyll triplets unquenched. This effect appears to be intrinsically related to the molecular organization of the Lhcb proteins.

The peripheral light-harvesting complex of photosystem I contains red chlorophylls (Chls) that, unlike the typical antenna Chls, absorb at lower energy than the primary electron donor P700. It has been shown that the red-most absorption band arises from two excitonically coupled Chls, although this interaction alone cannot explain the extreme red-shifted emission (25 nm, approximately 480 cm(-1) for Lhca4 at 4 K) that the red Chls present. Here, we report the electric field-induced absorption changes (Stark effect) on the Q(y) region of the Lhca4 complex. Two spectral forms, centered around 690 nm and 710 nm, were necessary to describe the absorption and Stark spectra. The analysis of the lowest energy transition yields a high value for the change in dipole moment, Deltamu(710nm) approximately 8 Df(-1), between the ground and excited states as compared with monomeric, Deltamu = 1 D, or dimeric, Deltamu = 5 D, Chl a in solution. The high value of the Deltamu demonstrates that the origin of the red-shifted emission is the mixing of the lowest exciton state with a charge-transfer state of the dimer. This energetic configuration, an excited state with charge-transfer character, is very favorable for the trapping and dissipation of excitations and could be involved in the photoprotective mechanism(s) of the photosystem I complex.