Effect of adaptation to high light intensity on the kinetics of energy transfer from phycobilisomes to photosystem II in Anabaena cylindrica

Transfer efficiencies between phycobilisomes and photosystem II antenna chlorophylls were determined on membrane fragments isolated from low and high light adapted Anabaena cells. The observed increase in energy transfer in high light adapted cells is a consequence of shorter interchromophore distances and a decrease in the number of jumps of the exciting photons. Calculation of the rates of energy transfer and the coupling energies indicate that the weak interaction inferred for energy transfer between phycobilisome and photosystem II in low light adapted cells is replaced by an intermediate interaction in high light adapted cells.Abbreviations LLA low light adapted - HLA high light adapted - PBS phycobilisome - PS photosystem     

Effects of proteinase K on the energy transfer between phycobiliproteins in phycobilisomes

Spectral changes in fluorescence of phycobilisomes (PBS) of A. variabilis treated with proteinase K were studied at room and liquid nitrogen temperature. In control PBS, the relative yield of 77 K fluorescence of F686 was very high, and those of F655 and F666 were low. In PBS treated with proteinase K for less than 1 h, F686 decreased, and F655 and F666 increased. In PBS treated with proteinase K for 2 h, F655 was the main peak of fluorescence emission, F686 was the second peak, the fluorescence emission peak of F666 disappeared. In PBS treated with proteinase K for more than 8 h, F655 showed only one fluorescence emission peak.We suggested that phycobiliporteins in the PBS of A. variabilis constitute an energy transfer chain, shown as follows:{fx91-1}The linkages between APC and APC-B, C-PC and APC, and C-PC and APC-B had different sensitivity towards proteinase K.     

The Supramolecular Structure of Photosystem II — Phycobilisome-Complexes of Porphyridium cruentum

The structure and arrangement of phycobilisomes of the unicellular red alga Porphyridium cruentum is compared with the organization of the thylakoid freeze-fracture particles in order to determine the relationship between phycobilisomes and photosystem II. The hemi-ellipsoidal phycobilisomes, 20 nm thick, are predominantly organized into rows; their centre to centre periodicity is 30–40 nm, so that they are well separated by a gap of 10–20 nm. The phycobilisomes are cleaved by a central faint furrow, parallel to the long axis from top to base. The organization of the exoplasmic particles in rows is similar to the arrangement of the phycobilisomes so that a structural relationship between both systems, previously demonstrated in cyanobacteria, is evident. Within the rows, the 10 nm EF-particles are grouped in tetrameric complexes separated by distances similar to those observed for phycobilisomes. We propose that the tetrameric EF-particle complexes correspond to tetrameric photosystem II complexes which bind one hemi-ellipsoidal phycobilisome on the stroma exposed surface of the thylakoid. A hypothetical model of this photosystem II-phycobilisome complex is presented.     

Recombinant phycobiliproteins. Recombinant C-phycocyanins equipped with affinity tags, oligomerization, and biospecific recognition domains

A family of specific cloning vectors was constructed to express in the cyanobacterium Anabaena sp. PCC7120 recombinant C-phycocyanin subunits with one or more different tags, including the 6xHis tag, oligomerization domains, and the streptavidin-binding Strep2 tag. Such tagged alpha or beta subunits of Anabaena sp. PCC7120 C-phycocyanin formed stoichiometric complexes in vivo with appropriate wild-type subunits to give constructs with the appropriate oligomerization state and normal posttranslational modifications and with spectroscopic properties very similar to those of unmodified phycocyanin. All of these constructs were incorporated in vivo into the rod substructures of the light-harvesting complex, the phycobilisome. The C-terminal 114-residue portion of the Anabaena sp. PCC7120 biotin carboxyl carrier protein (BCCP114) was cloned and overexpressed and was biotinylated up to 20% in Escherichia coli and 40% in wild-type Anabaena sp. His-tagged phycocyanin beta--BCCP114 constructs expressed in Anabaena sp. were >30% biotinylated. In such recombinant phycocyanins equipped with stable trimerization domains, >75% of the fusion protein was specifically bound to streptavidin- or avidin-coated beads. Thus, the methods described here achieve in vivo production of stable oligomeric phycobiliprotein constructs equipped with affinity purification tags and biospecific recognition domains usable as fluorescent labels without further chemical manipulation.     

Light Harvesting and State Transitions in Cyanobacteria

Cyanobacteria are oxygenic phototrophic prokaryotes and are considered to be the ancestors of chloroplasts. Their photosynthetic machinery is functionally equivalent in terms of primary photochemistry and photosynthetic electron transport. Fluorescence measurements and other techniques indicate that cyanobacteria, like plants, are capable of redirecting pathways of excitation energy transfer from light harvesting antennae to both photosystems. Cyanobacterial cells can reach two energetically different states, which are defined as “State 1” (obtained after preferential excitation of photosystem I) and “State 2” (preferential excitation of photosystem II). These states can be distinguished by static and time resolved fluorescence techniques. One of the most important conclusions reached so far is that the presence of both photosystems, as well as certain antenna components, are necessary for state transitions to occur. Spectroscopic evidence suggests that changes in the coupling state of the light harvesting antenna complexes (the phycobilisomes) to both photosystems occur during state transitions. The finding that the phycobilisome complexes are highly mobile on the surface of the thylakoid membrane (the mode of interaction with the thylakoid membrane is essentially unknown), has led to the proposal that they are in dynamic equilibrium with both photosystems and regulation of energy transfer is mediated by changes in affinity for either photosystem.     

Structural organization of an intact phycobilisome and its association with photosystem II

Phycobilisomes (PBSs) are light-harvesting antennae that transfer energy to photosynthetic reaction centers in cyanobacteria and red algae. PBSs are supermolecular complexes composed of phycobiliproteins (PBPs) that bear chromophores for energy absorption and linker proteins. Although the structures of some individual components have been determined using crystallography, the three-dimensional structure of an entire PBS complex, which is critical for understanding the energy transfer mechanism, remains unknown. Here, we report the structures of an intact PBS and a PBS in complex with photosystem II (PSII) from Anabaena sp. strain PCC 7120 using single-particle electron microscopy in combination with biochemical and molecular analyses. In the PBS structure, all PBP trimers and the conserved linker protein domains were unambiguously located, and the global distribution of all chromophores was determined. We provide evidence that ApcE and ApcF are critical for the formation of a protrusion at the bottom of PBS, which plays an important role in mediating PBS interaction with PSII. Our results provide insights into the molecular architecture of an intact PBS at different assembly levels and provide the basis for understanding how the light energy absorbed by PBS is transferred to PSII.     

Fluorescence induction in the phycobilisome-containing cyanobacterium Synechococcus sp PCC 7942: Analysis of the slow fluorescence transient

At room temperature, the chlorophyll (Chl) a fluorescence induction (FI) kinetics of plants, algae and cyanobacteria go through two maxima, P at ∼ 0.2-1 and M at ∼ 100-500 s, with a minimum S at ∼ 2-10 s in between. Thus, the whole FI kinetic pattern comprises a fast OPS transient (with O denoting origin) and a slower SMT transient (with T denoting terminal state). Here, we examined the phenomenology and the etiology of the SMT transient of the phycobilisome (PBS)-containing cyanobacterium Synechococcus sp PCC 7942 by modifying PBS → Photosystem (PS) II excitation transfer indirectly, either by blocking or by maximizing the PBS → PS I excitation transfer. Blocking the PBS → PS I excitation transfer route with N-ethyl-maleimide [NEM; A. N. Glazer, Y. Gindt, C. F. Chan, and K.Sauer, Photosynth. Research 40 (1994) 167-173] increases both the PBS excitation share of PS II and Chl a fluorescence. Maximizing it, on the other hand, by suspending cyanobactrial cells in hyper-osmotic media [G. C. Papageorgiou, A. Alygizaki-Zorba, Biochim. Biophys. Acta 1335 (1997) 1-4] diminishes both the PBS excitation share of PS II and Chl a fluorescence. Here, we show for the first time that, in either case, the slow SMT transient of FI disappears and is replaced by continuous P → T fluorescence decay, reminiscent of the typical P → T fluorescence decay of higher plants and algae. A similar P → T decay was also displayed by DCMU-treated Synechococcus cells at 2 °C. To interpret this phenomenology, we assume that after dark adaptation cyanobacteria exist in a low fluorescence state (state 2) and transit to a high fluorescence state (state 1) when, upon light acclimation, PS I is forced to run faster than PS II. In these organisms, a state 2 → 1 fluorescence increase plus electron transport-dependent dequenching processes dominate the SM rise and maximal fluorescence output is at M which lies above the P maximum of the fast FI transient. In contrast, dark-adapted plants and algae exist in state 1 and upon illumination they display an extended P → T decay that sometimes is interrupted by a shallow SMT transient, with M below P. This decay is dominated by a state 1 → 2 fluorescence lowering, as well as by electron transport-dependent quenching processes. When the regulation of the PBS → PS I electronic excitation transfer is eliminated (as for example in hyper-osmotic suspensions, after NEM treatment and at low temperature), the FI pattern of Synechococcus becomes plant-like.     

Role of inter-domain cavity in the attachment of the orange carotenoid protein to the phycobilisome core and to the fluorescence recovery protein

Using molecular modeling and known spatial structure of proteins, we have derived a universal 3D model of the orange carotenoid protein (OCP) and phycobilisome (PBS) interaction in the process of non-photochemical PBS quenching. The characteristic tip of the phycobilin domain of the core-membrane linker polypeptide (L_CM) forms the attachment site on the PBS core surface for interaction with the central inter-domain cavity of the OCP molecule. This spatial arrangement has to be the most advantageous one because the L_CM, as the major terminal PBS-fluorescence emitter, accumulates energy from the most other phycobiliproteins within the PBS before quenching by OCP. In agreement with the constructed model, in cyanobacteria, the small fluorescence recovery protein is wedged in the OCP’s central cavity, weakening the PBS and OCP interaction. The presence of another one protein, the red carotenoid protein, in some cyanobacterial species, which also can interact with the PBS, also corresponds to this model.     

Non-photochemical quenching of fluorescence in cyanobacteria

The pathways of energy dissipation of excessive absorbed energy in cyanobacteria in comparison with that in higher plants are discussed. Two mechanisms of non-photochemical quenching in cyanobacteria are described. In one case this quenching occurs as light-induced decrease of the fluorescence yield of long-wavelength chlorophylls of the photosystem I trimers induced by inactive reaction centers: P700 cation-radical or P700 in triplet state. In the other case, non-photochemical quenching in cyanobacteria takes place with contribution of water-soluble protein OCP (containing 3′-hydroxyechinenone) that induces reversible quenching of allophycocyanin fluorescence in phycobilisomes. The possible evolutionary pathways of the involvement of carotenoid-binding proteins in non-photochemical quenching are discussed comparing the cyanobacterial OCP and plant PsbS protein. Published in Russian in Biokhimiya, 2007, Vol. 72, No. 10, pp. 1385–1395.