Biochemistry (Moscow)最新文献

Thermal burns of the skin are associated not only with local tissue alterations but also with the development of systemic disorders, which promote generalization of inflammatory processes. In particular, burn injury leads to an overproduction of reactive oxygen species, activation of free-radical oxidation, and lipid peroxidation. This study investigated the protective role of plastoquinone, a natural plant antioxidant, on the morphological condition of the skin and on the shape and aggregation of erythrocytes in rats with second-degree thermal burns. Thermal burn resulted in the decrease in epidermis thickness, increase in the number of hyperemic vessels, damaged hair follicles and sebaceous glands. Application of plastoquinone, incorporated into liposomes, onto the damaged skin had a protective effect on the skin structures; in the case of liposomes applied without plastoquinone, the protective effect was less pronounced. In addition, thermal burn altered the state of erythrocytes, leading to their deformation and aggregation. Plastoquinone in liposomes applied topically or administered intravenously showed a protective effect on erythrocytes comparable to that of ubiquinone, preventing the development of burn-induced erythrocyte shape alterations. However, only plastoquinone administered intravenously completely prevented erythrocyte aggregation, thus eliminating negative effects of the burn injury on the functional activity of erythrocytes, indicating the potential of plant-derived plastoquinone as an effective agent in burn injury management.

Klaus Möbius, Professor at the Free University of Berlin, was an outstanding physical chemist and biophysicist. He was a pioneer in the development of high-field/high-frequency EPR spectroscopy methods and their application in the study of photosynthesis. Among the most essential are the applications in studying the charge transfer kinetics and properties of the ion-radical pairs in photosynthetic reaction centers (RC). Under his leadership and with his direct participation a unique setup allowing registration of the kinetics of the electron transfer between the (bacterio)chlorophyll dimer and quinone in the bacterial photosynthetic RC and plant photosystem I (PSI) was created. This setup also allowed precise determining of the distance between separated charges based on measuring the frequencies of the Electron Spin Echo Envelope Modulation (ESEEM). This setup made it possible to prove that electron transfer in PSI occurs mainly along the A branch of redox cofactors. The kinetics of backward electron transfer reaction (reoxidation of the phyllosemiquinone anion A₁⁻ and reduction of the photooxidized chlorophyll dimer P₇₀₀⁺) in PSI were measured under the same conditions. The essential data on the bioprotective effect of the disaccharide trehalose on the kinetics of forward and backward electron transfer in PSI complexes were obtained. A significant slowdown in the kinetics of electron transfer due to the restriction of protein conformational mobility, as well as long-term maintaining of functional activity of PSI dried in a vitreous trehalose matrix at room temperature (i.e., subjected to a reversible anhydrobiosis) was demonstrated. These results obtained in collaboration with Prof. Möbius and Prof. Venturoli (Bologna) allowed elucidating the role of hydrogen bond network and the conformational mobility of the protein subunits in facilitating electron transfer in the photosynthetic RC.

One of the adaptive mechanisms used by photosynthetic organisms in response to changing light conditions is redistribution of antenna complexes between the photosystems, a process known as state transitions (ST). This mechanism allows to regulate the amount of light energy absorbed by the photosystems. Numerous studies have reported inhibition of ST at the elevated light intensity; however, the mechanism underlying this process is still debated. We evaluated the effect of H₂O₂ at various concentrations on the ST process in functionally active thylakoids isolated from Arabidopsis thaliana leaves and investigated which stage of this process is affected by H₂O₂. To assess the extent of ST, we measured low-temperature chlorophyll a fluorescence spectra (650-780 nm) and calculated the F745/F685 ratio, whose changes can serve as an indicator of ST progression. H₂O₂ inhibited ST under the low-intensity light conditions and, furthermore, led to a decrease in the accumulation of phosphorylated Lhcb1 and Lhcb2 proteins involved in ST. This suggests that the observed ST inhibition resulted from the suppression of STN7 kinase activity. Importantly, H₂O₂ in the tested concentrations did not affect the electron transport rate, indicating that the inhibition of STN7 kinase activity was not associated with suppression of the photosynthetic electron transport chain (PETC) activity. The treatment with H₂O₂ did not reduce the level of phosphorylated D1 protein (a product of phosphorylation by the thylakoid STN8 kinase). Taken together, these results demonstrate for the first time the mechanism by which H₂O₂ inhibits STN7 kinase activity and, consequently, the process of ST.

Some microalgae are capable of light-dependent hydrogen production after a period of anaerobic adaptation, thus performing biophotolysis of water. The rate of hydrogen production the start of illumination has the rate equal to the maximum rate of photosynthesis. However, this process is short-lived: oxygen produced during photosynthesis quickly inactivates the key enzyme of biophotolysis, hydrogenase, and inhibits its expression. To date, approaches have been developed to achieve sustained hydrogen production by microalgae. The most studied are those based on transferring microalgae to nutrient-deficient conditions. However, it is known that hydrogen production under nutrient deficiency is always accompanied by the decrease in activity of photosystem II (PSII). Several mechanisms for suppression of PSII activity have been described in the literature, and there is no consensus on which mechanism is the determining one. The aim of this work was to test the hypothesis that realization of a particular mechanism of PSII suppression depends not only on the type of stress but also on the growth conditions. For this purpose, the photoautotrophic culture of the microalga Chlamydomonas reinhardtii was grown under nitrogen or sulfur deficiency under different light regimes, and realization of the following mechanisms of PSII activity suppression was analyzed: over-reduction of the plastoquinone pool (coupled with over-reduction of the entire photosynthetic electron transport chain), decoupling of PSII (based on the kinetics of ascorbate accumulation and the JIP test) with water-oxidizing complex, violaxanthin cycle, anaerobic stress associated with the creation of a reducing redox potential of the culture suspension. It was found that the key mechanism determining hydrogen production is the over-reduction of the plastoquinone pool. Other mechanisms are also realized under various conditions but do not show clear correlation with hydrogen production. The obtained results indicate that induction of stress through starvation of cultures is a convenient approach for studying hydrogen production by microalgae, but due to the low activity of PSII, it is impractical. New approaches are required to create industrial systems based on microalgae, allowing full realization of their photosynthetic potential.

The knockout of either At3g01500 or At3g52720 gene encoding Arabidopsis thaliana βCA1 and αCA1 carbonic anhydrase (CA), respectively, led to a lower CA activity of the chloroplast stroma preparations from the knockout mutant plants (αCA1-KO and βCA1-KO) compared to such preparations from the wild-type (WT) plants. To identify the differences in the photosynthetic characteristics of mutant and WT plants, they were grown in low light (LL; 50-70 µmol quanta∙m⁻²∙s⁻¹, natural conditions) and high light (HL; 400 µmol quanta∙m⁻²∙s⁻¹, stressful conditions). The rate of CO₂ assimilation measured at 400 µmol quanta∙m⁻²∙s⁻¹ in plants grown under LL was lower in αCA1-KO and βCA1-KO mutants compared to WT plants. The rate of photosynthetic electron transport was lower in αCA1-KO plants and higher in βCA1-KO plants than in WT plants; the content of CO₂ in chloroplasts was lower in βCA1-KO plants than in both WT and αCA1-KO plants, where it differed little. The value of the proton-motive force was higher in βCA1-KO plants and lower in αCA1-KO plants than in WT plants due to changes in ΔpH value. The obtained results suggest that βCA1 facilitates the intake of inorganic carbon into chloroplasts, while αCA1 ensures the conversion of bicarbonate into CO₂ in the chloroplast stroma for its use in the reaction catalyzed by Ribulose bisphosphate carboxylase/oxygenase (RuBisCO). In both αCA1-KO and βCA1-KO mutants, the expression levels of genes encoding other chloroplast CAs differed markedly from those in WT plants; the pattern of the changes in the genes expression depended on the light intensity during cultivation. The content of hydrogen peroxide in the leaves of both αCA1-KO and βCA1-KO mutants was higher in LL and lower in HL than in WT plants. The expression levels of stress marker genes changed similarly in both types of mutant plants. A possible involvement of the chloroplast stroma CAs in the transmission of stress signals in higher plants is discussed.

This review addresses photosynthetic control as a protective mechanism that prevents photoinhibition of photosystem I under conditions of imbalance between CO₂ assimilation during the Calvin–Benson–Bassham cycle and light reactions in the thylakoid photosynthetic apparatus. We discuss the pathways of photosystem I photoinhibition and describe protective mechanisms that prevent photodamage of photosystem I. We propose a hypothesis regarding the influence of photosynthetic control on formation of reactive oxygen species in photosystem I. pH-sensitivity of plastoquinol oxidation at the quinol-oxidizing (Qo) site of the cytochrome b₆f complex is analyzed, and function of two proton-conducting channels that release protons into the thylakoid lumen from the cytochrome b₆f complex is described. We examine impact of photosynthetic control on the functioning of the cytochrome b₆f complex itself, and propose a hypothesis regarding the preferential activation of photosynthetic control in the thylakoid grana, which ensures operation of the cyclic electron transport around photosystem I as a main protective mechanism.

This work demonstrates, for the first time, capacity of the Chlorella sorokiniana immobilized in alginate to produce hydrogen (H₂) over an extended period of time when cultivated under strictly photoautotropic conditions on complete mineral medium. In order to reduce photosynthetic activity, immobilized cells were subjected to a 30-minute pre-incubation period at high light intensity of 1000 μmol photons m⁻²∙s⁻¹. The ability to produce H₂ was evaluated under illumination of 40 μmol/(m²∙s). The culture not bubbled with argon produced H₂ for 9 days; total gas yield was 0.1 mol H₂/m². In the culture under argon atmosphere, the release of H₂ continued for 51 days, resulting in a total yield of 0.55 mol H₂/m². The immobilized culture was capable of H₂ production at 16% O₂ in the gas phase, which may be due to the effects of photoinhibition and activation of oxygen uptake pathways in mitochondria and chloroplast. Analysis of the functioning of electron-transport chain in the microalgae cells revealed decrease in the rate of electron transport, increase in the size of the PSII antenna, and development of non-photochemical quenching processes, while activity of PSII remained moderately high (Fv/Fm = 0.4-0.6). Inhibitor analysis using 10⁻⁵ M DCMU demonstrated that contribution of PSII to hydrogenase reaction increased from 30% on the first day of the experiment to 50% by the fourth day. Addition of 10⁻⁵ M DBMIB led to the 90% reduction in the rate of H₂ formation on both day 1 and day 4.

Methods of site-directed mutagenesis are successfully used in structural and functional studies of photosynthetic reaction centers (RCs). It has been noted that many mutations near electron transfer cofactors reduce temperature stability of the Cereibacter sphaeroides RCs and affect amount of RCs in the membranes. We previously reported [Selikhanov et al. (2023) Membranes, 25, 154] that introduction of inter-subunit disulfide bridges on the periplasmic or cytoplasmic surface of the complex promotes increase in thermal stability of the C. sphaeroides RCs. In this work, an attempt was made to increase thermal stability of the mutant RC with the Ile M206 – Gln substitution by introducing inter-subunit disulfide bonds. This RC is of considerable interest for studying mechanisms of early electron transfer processes in RCs. The effect of mutations on the amount of RCs in chromatophores was analyzed and it was found that the I(M206)Q mutation leads to twofold decrease in the RC content in chromatophores, introduction of disulfide bonds on the cytoplasmic or periplasmic sides of the complex reduces the amount of RCs in membranes by one third, the triple substitution I(M206)Q/G(M19)C/T(L214)C reduces the amount of RCs in membranes almost 4-fold, and the substitutions I(M206)Q/V(M84)C/G(L278)C lead to disruption of RC assembly in the membrane. It was shown that introduction of the inter-subunit S-S bond on the cytoplasmic surface of the complex did not have a significant effect on thermal stability of the I(M206)Q RC. Our own and literature data on the factors influencing assembly processes and ensuring stability of the structure of integral membrane complexes are discussed.

Using electrometric technique, the cationic antiseptic octenidine was revealed to reduce generation of transmembrane electrical potential difference in the chromatophores of photosynthetic bacterium Cereibacter sphaeroides. This is also confirmed by measurements of electrochromic shifts of carotenoid absorption bands in chromatophores. In reaction centers (RCs), isolated from chromatophores in the absence of external electron donors and acceptors, the rate of recombination between photooxidized bacteriochlorophyll P₈₇₀ and reduced secondary quinone acceptor Q_B, as measured by absorption changes in the near infrared region, was very weakly dependent on the presence of antiseptics, in contrast to the kinetics in the 400-600 nm spectral range, where absorption changes associated with the oxidation of P₈₇₀ and the formation of semiquinone radicals Q_A⁻ and Q_B⁻, as well as electrochromic shifts of the carotenoid and bacteriopheophytin RC absorption bands, were observed. The addition of cationic antiseptics modified the flash-induced absorbance changes in this region with the formation time of ~100-200 ms and a decay time of ~3 s. In the series: picloxydine – chlorhexidine – octenidine – miramistin, the last one was the most effective. The maximum amplitude of such changes was observed in the absorption region of the semiquinone radical around 460 nm. When electron transfer from Q_A⁻ to Q_B was blocked by o-phenanthroline, the effect disappeared. Cationic antiseptics are suggested to stimulate protonation of Q_B⁻ with the formation of a neutral Q_B⁻H⁺ complex.