Y.V. Bolychevtseva

Protein–protein interactions in carotenoid triggered quenching of phycobilisome fluorescence in Synechocystis sp. PCC 6803

An inquiry into the effect of temperature on carotenoid triggered quenching of phycobilisome (PBS) fluorescence in a photosystem II‐deficient mutant of Synechocystis sp. results in identification of two temperature‐dependent processes: one is responsible for the quenching rate, and one determines the yield of PBS fluorescence. Non‐Arrhenius behavior of the light‐on quenching rate suggests that carotenoid‐absorbed light triggers a process that bears a strong resemblance to soluble protein folding, showing temperature‐dependent enthalpy of activated complex formation. The response of PBS fluorescence yield to hydration changing additives and to passing of the membrane lipid phase transition point indicates that the pool size of PBSs subject to quenching depends on the state of some membrane component.

Photosynthetic and respiratory electron transport in the alkaliphilic cyanobacterium Arthrospira (Spirulina) platensis

Photosynthetic and respiratory electron transport and their interplay with ion transport have been studied in Arthrospira platensis, a filamentous alkaliphilic cyanobacterium living in hypersaline lakes. As typical for alkaliphiles, A. platensis apparently does not maintain an outward positive pH gradient at its plasma membrane. Accordingly, sodium extrusion occurs via an ATP-dependent primary sodium pump, in contrast to the Na+/H+ antiport in most cyanobacteria. A. platensis is strongly dependent on sodium/bicarbonate symport for the uptake of inorganic carbon. Sodium extrusion in the presence of the Photosystem II inhibitor diuron indicates that a significant amount of ATP is supplied by cyclic electron transport around Photosystem I, the content of which in A. platensis is exceptionally high. Plastoquinol is oxidized by two parallel pathways, via the cytochrome b6f complex and a putative cytochrome bd complex, both of which are active in the light and in the dark.

The development of carotenoid-deficient membranes in plastids of barley seedlings treated with norflurazon☆

The effect of carotenoid (Car) deficiency on the formation of thylakoid membranes in barley seedlings grown with norflurazon (NF) was investigated. To exclude photodestruction during the growth of Car-deficient seedlings, the etiolated seedlings were illuminated for 24 h with 2.5 ms flashes every 12 min. The morphometric analysis of seven main ultrastructural parameters of plastids control and NF-treated seedlings was carried out. The length of one partition and the number of partitions per plastids section, which characterize the initial stages of stacking, showed no difference in etiolated NF-treated and control seedlings. Compared with the control, the number of partitions was considerably lower in those plastids of Car-deficient seedlings which, after flash illumination, were kept for 1.5 h in continuous light of low intensity. The photobleaching of chlorophyll (Chl) in post-illuminated seedlings provides an indication of the photodestructive processes in Car-deficient seedlings exposed to low-intensity light. Chl bleaching did not appear in Car-deficient seedlings illuminated only with flashes; however, the stacking of the membranes was disturbed: the partition lengths and number of partitions per plastid section were higher in flashed, non-treated seedlings than in Car-deficient seedlings. Although the flashed control and NF-treated seedlings had an equal amount of Chl and the same polypeptide composition, no activity of photosystem II (PSII) and no PSII reaction centre complex were found in NF-treated seedlings. Car-deficient seedlings mainly contained photosystem I (PSI) with a ratio of Chl to P700 of 60 compared with 150 in the control. It is therefore suggested that Car deficiency in flashed leaves, which may accumulate only a small amount of light-harvesting Chl ab protein, prevents the assembly of the PSII complex and membrane stacking.

Photosystem activity and state transitions of the photosynthetic apparatus in cyanobacterium Synechocystis PCC 6803 mutants with different redox state of the plastoquinone pool

To better understand how photosystem (PS) activity is regulated during state transitions in cyanobacteria, we studied photosynthetic parameters of photosystem II (PSII) and photosystem I (PSI) in Synechocystis PCC 6803 wild type (WT) and its mutants deficient in oxidases (Ox−) or succinate dehydrogenase (SDH−). Dark-adapted Ox− mutant, lacking the oxidation agents, is expected to have a reduced PQ pool, while in SDH− mutant the PQ pool after dark adaptation will be more oxidized due to partial inhibition of the respiratory chain electron carriers. In this work, we tested the hypothesis that control of balance between linear and cyclic electron transport by the redox state of the PQ pool will affect PSII photosynthetic activity during state transition. We found that the PQ pool was reduced in Ox− mutant, but oxidized in SDH− mutant after prolonged dark adaptation, indicating different states of the photosynthetic apparatus in these mutants. Analysis of variable fluorescence and 77K fluorescence spectra revealed that the WT and SDH− mutant were in State 1 after dark adaptation, while the Ox− mutant was in State 2. State 2 was characterized by ∼1.5 time lower photochemical activity of PSII, as well as high rate of P700 reduction and the low level of P700 oxidation, indicating high activity of cyclic electron transfer around PSI. Illumination with continuous light 1 (440 nm) along with flashes of light 2 (620 nm) allowed oxidation of the PQ pool in the Ox− mutant, thus promoting it to State 1, but it did not affect PSII activity in dark adapted WT and SDH− mutant. State 1 in the Ox− mutant was characterized by high variable fluorescence and P700+ levels typical for WT and the SDH− mutant, indicating acceleration of linear electron transport. Thus, we show that PSII of cyanobacteria has a higher photosynthetic activity in State 1, while it is partially inactivated in State 2. This process is controlled by the redox state of PQ in cyanobacteria through enhancement/inhibition of electron transport on the acceptor side of PSII.

Association of high light-inducible HliA/HliB stress proteins with photosystem 1 trimers and monomers of the cyanobacterium Synechocystis PCC 6803

Hlip (high light-inducible proteins) are important for protection of the photosynthetic apparatus of cyanobacteria from light stress. However, the interaction of these proteins with chlorophyll–protein complexes of thylakoids remains unclear. The association of HliA/HliB stress proteins with photosystem 1 (PS1) complexes of the cyanobacterium Synechocystis PCC 6803 was studied to understand their function. Western blotting demonstrated that stress-induced HliA/HliB proteins are associated with PS1 trimers in wild-type cells grown under moderate light condition (40 µmol photons/m2 per sec). The content of these proteins increased 1.7-fold after light stress (150 µmol photons/m2 per sec) for 1 h. In the absence of PS1 trimers (ΔpsaL mutant), the HliA/HliB proteins are associated with PS1 monomers and the PS2 complex. HliA/HliB proteins are associated with PS1 monomers but not with PS1 trimers in Synechocystis PS2-deficient mutant grown at 5 µmol photons/m2 per sec; the content of Hli proteins associated with PS1 monomers increased 1.2-fold after light stress. The HliA/HliB proteins were not detected in wild-type cells of cyanobacteria grown in glucose-supplemented medium at 5 µmol photons/m2 per sec, but light stress induces the synthesis of stress proteins associated with PS1 trimers. Thus, for the first time, the association of HliA/HliB proteins not only with PS1 trimers, but also with PS1 monomers is shown, which suggests a universal role of these proteins in the protection of the photosynthetic apparatus from excess light.

Thermostability of photosystem I trimers and monomers from the cyanobacterium Thermosynechococcus elongatus

The performance of solar energy conversion into alternative energy sources in artificial systems highly depends on the thermostability of photosystem I (PSI) complexes Terasaki et al. (2007), Iwuchukwu et al. (2010), Kothe et al. (2013) . To assess the thermostability of PSI complexes from the thermophilic cyanobacterium Thermosynechococcus elongatus heating induced perturbations on the level of secondary structure of the proteins were studied. Changes were monitored by Fourier transform infrared (FT-IR) spectra in the mid-IR region upon slow heating (1°C per minute) of samples in D2O phosphate buffer (pD 7.4) from 20°C to 100°C. These spectra showed distinct changes in the Amide I region of PSI complexes as a function of the rising temperature. Absorbance at the Amide I maximum of PSI monomers (centered around 1653cm-1), gradually dropped in two temperature intervals, i.e. 60-75 and 80-90°C. In contrast, absorbance at the Amide I maximum of PSI trimers (around 1656cm-1) dropped only in one temperature interval 80-95°C. The thermal profile of the spectral shift of α-helices bands in the region 1656-1642cm-1 confirms the same two temperature intervals for PSI monomers and only one interval for trimers. Apparently, the observed absorbance changes at the Amide I maximum during heating of PSI monomers and trimers are caused by deformation and unfolding of α-helices. The absence of absorbance changes in the interval of 20-65°C in PSI trimers is probably caused by a greater stability of protein secondary structure as compared to that in monomers. Upon heating above 80°C a large part of α-helices both in trimers and monomers converts to unordered and aggregated structures. Spectral changes of PSI trimers and monomers heated up to 100°C are irreversible due to protein denaturation and non-specific aggregation of complexes leading to new absorption bands at 1618-1620cm-1. We propose that monomers shield the denaturation sensitive sides at the monomer/monomer interface within a trimer, making the oligomeric structure more stable against thermal stress.

Photosystem II activity of wild type Synechocystis PCC 6803 and its mutants with different plastoquinone pool redox states

To assess the role of redox state of photosystem II (PSII) acceptor side electron carriers in PSII photochemical activity, we studied sub-millisecond fluorescence kinetics of the wild type Synechocystis PCC 6803 and its mutants with natural variability in the redox state of the plastoquinone (PQ) pool. In cyanobacteria, dark adaptation tends to reduce PQ pool and induce a shift of the cyanobacterial photosynthetic apparatus to State 2, whereas illumination oxidizes PQ pool, leading to State 1 (Mullineaux, C. W., and Holzwarth, A. R. (1990) FEBS Lett., 260, 245-248). We show here that dark-adapted Ox– mutant with naturally reduced PQ is characterized by slower QA– reoxidation and O2 evolution rates, as well as lower quantum yield of PSII primary photochemical reactions (Fv/Fm) as compared to the wild type and SDH–mutant, in which the PQ pool remains oxidized in the dark. These results indicate a large portion of photochemically inactive PSII reaction centers in the Ox– mutant after dark adaptation. While light adaptation increases Fv/Fm in all tested strains, indicating PSII activation, by far the greatest increase in Fv/Fm and O2 evolution rates is observed in the Ox– mutant. Continuous illumination of Ox– mutant cells with low-intensity blue light, that accelerates QA– reoxidation, also increases Fv/Fm and PSII functional absorption cross-section (590 nm); this effect is almost absent in the wild type and SDH–mutant. We believe that these changes are caused by the reorganization of the photosynthetic apparatus during transition from State 2 to State 1. We propose that two processes affect the PSII activity during changes of light conditions: 1) reversible inactivation of PSII, which is associated with the reduction of electron carriers on the PSII acceptor side in the dark, and 2) PSII activation under low light related to the increase in functional absorption cross-section at 590 nm.

Carotenoid-Induced Non-photochemical Fluorescence Quenching in Phycobilisomes of the Cyanobacterium Synechocystis sp. PCC 6803

A mechanism of carotenoid triggered quenching of phycobilisome fluorescence in a photosystem II-deficient mutant of Synechocystis sp. has been studied. The effect of temperature on quenching reveals two temperature-dependent processes: one is responsible for the quenching rate, and one determines the yield of phycobilisome fluorescence. The non-Arrhenius behavior of the light-on quenching rate suggests that carotenoidabsorbed light triggers a process that bears a strong resemblance to soluble protein folding, showing temperature-dependent enthalpy of activated complex formation. The response of phycobilisome fluorescence yield to hydration changes, and to membrane lipid phase transitions, indicates that the pool size of phycobilisome quenching depends on the state of some membrane component.