Lanthanide cations (La3+ and Tb3+) bind to the Ca-binding site of the oxygen-evolving complex in Ca-depleted PSII membranes and irreversibly inhibit the oxygen evolution. Оn the other hand, EPR measurement of Mn2+ concentration in buffer revealed that lanthanide cations inhibit the light-dependent oxidation of Mn2+ cations via the high-affinity Mn-binding site in Mn-depleted PSII membranes, which suggests that they bind to and inhibit the high-affinity Mn-binding site of the oxygen-evolving complex. The inhibition is irreversible, bound Ln3+ cation could not be washed out from the sample. Calcium ion inhibits oxidation of Mn2+ (5 μM) at very high concentration (tens mM) and the inhibition is reversible. In this work we measured the reduction rate of exogenic electron acceptor 2,6-dichlorophenolindophenol during the oxidation of Mn2+ cations in the Ca-depleted PSII and in the Ca-depleted PSII treated with lanthanides after extraction of Mn cluster from these preparations. We found that irreversible binding of the lanthanide cation to the Ca-binding site in the Ca-depleted PSII membranes leads to a partial inhibition of the high-affinity Mn-binding site.

С помощью спектроскопии комбинационного рассеяния исследовали изменения конформация гема и глобина гемоглобина при варьировании рО2 в среде инкубации эритроцитов и гемоглобина. При изменении pO2, в спектрах комбинационного рассеяния гемоглобина в эритроците и в растворе наблюдаются изменения ряда характерных полос как в области 1000-1700 см-1 (валентные колебания гема), так и в высокочастотной области спектра (2800-3000 см-1, валентные колебания аминокислот глобина). Установлено, что зависимость степени оксигенации гемоглобина (sO2), рассчитанной из параметров спектра комбинационного рассеяния, характеризуется S-образной кривой только для гемоглобина в эритроците. Установлено, что различия в конформация гема при изменении рО2 меняют его способность образовывать и комплексы с NO. При увеличении рО2 изменения конформация гема гемоглобина в клетке сопровождаются увеличением вклада боковых -СН3-групп колебаний полуколец пиррола и групповых колебания связей полуколец пиррола, а конформации глобина гемоглобина обусловлено увеличением вклада колебаний Н-метиновых групп и симметричных концевых метиленовых групп аминокислот.

The variation in the conformations of hemoglobin heme and globin in response to different oxygen partial pressures ( p O 2 ) has been studied by Raman spectroscopy inside erythrocytes and in hemoglobin dissolved in incubation medium. The spectra of hemoglobin inside erythrocytes and in the medium show that several characteristic bands in the range from 1000 to 1700 cm –1 (stretching vibrations in the heme group) and in the high-frequency range from 2800 to 3000 cm –1 (stretching vibrations in globin amino acids) vary with p O 2 levels. The curve of hemoglobin saturation with oxygen ( s O 2 ) obtained from the Raman-scattering data displays sigmoidal behavior only in hemoglobin inside erythrocytes. It has been shown that differences in heme conformation at different p O 2 levels affect the ability of hemoglobin to form complexes with nitric oxide. As the p O 2 level increases, the conformational changes of hemoglobin heme in the cell entail an increase in the contribution of vibrations of lateral –CH 3 groups in pyrrole half-rings and group vibrations of bonds inside pyrrole half-rings, whereas the globin conformation is associated with a higher contribution of vibrations from H-methine groups and symmetrical terminal methylene groups in amino acids.

Abstract—Using Raman spectroscopy (RS) approach in a spectral range of 1000–3000 cm–1 were used to study the conformational and structural changes that arise in the heme group and globin moiety of hemoglobin in human red blood cells at various temperatures and oxygen contents. In hypoxia, the hemoglobin conformation was shown to change as a result of the increasing contribution of hematoporphyrin pyrrole rings and vibrational motions of vinyl groups. Modifications were additionally detected in the contributions of symmetric and asymmetric vibrations of the CH2 and CH3 radicals of histidine (2850, 2860, and 2900 cm–1) and lysine (2880 and 2860 cm–1) residues. The mechanisms of oxygen binding are discussed for hemoglobin located in the submembrane region and cytoplasm of the cell.

Red blood cells are involved not only in transportation of oxygen and carbon dioxide but also in autoregulation of vascular tone by ATP release in hypoxic conditions. Molecular mechanisms of the ATP release from red blood cells in response to a decrease in partial oxygen pressure still remain to be elucidated. In this work we have studied effects of hypoxia on red blood cell hemolysis in humans and rats and compared the effects of inhibitors of ecto-ATPase and pannexin on the release of ATP and hemoglobin from rat erythrocytes. The 20-min hypoxia at 37°C increased hemolysis of red blood cells in humans and rats 1.5- and 2.5-fold, respectively. In rat erythrocytes a significant increase in hypoxia-induced extracellular ATP level was found only in the presence of ecto-ATPase inhibitor ARL 67156. In these conditions we observed a positive correlation ( R 2 = 0.5003) between the increase in free hemoglobin concentration and the ATP release. Neither carbenoxolon nor probenecid, the inhibitors of low-selectivity pannexin channels, altered the hypoxia-induced ATP release from rat erythrocytes. The obtained results indicate a key role of hemolysis in the ATP release from red blood cells.

Диаметр кровеносных сосудов увеличивается в ответ на снижение парциального давления кислорода. Установлено, что эта уникальная функция изолированных сосудов реализуется только в условиях, когда перфузируемая жидкость содержит эритроциты. Получены данные о том, что сенсором кислорода является гемоглобин, который в деоксигенированном состоянии взаимодействует с белком полосы 3, запуская не идентифицированные сигнальные каскады, влияющие на продукцию оксида азота и высвобождение из эритроцитов ATP, взаимодействующего с Р2Y-пуринергическими рецепторами эндотелия. В обзоре рассмотрены системы, принимающие участие в высвобождении ATP из эритроцитов, а также физиологическое и патофизиологическое значение этого явления.

A drop in oxygen partial pressure results in elevation of blood vessel diameter. It has been demonstrated that isolated vessels exhibit this unique feature only when they are perfused in the presence of erythrocytes. More recently, it was shown that haemoglobin plays a key role in oxygen sensing. Its deoxygenated form interacts with band 3 protein, triggering the cascade of non-identified intracellular signals involved in nitric oxide production and release of ATP interacting with P2Y purinergic receptors in endothelial cells. In this review, we summarize the data on mechanisms of ATP release from erythrocytes, as well as on its physiological and pathophysiological implications.

Morphology of erythrocytes and conformation of hemoglobin-derived hematoporphyrin were studied in patients with coronary heart disease (CHD) and patients with circulatory failure using laser interference microscopy and Raman spectroscopy. Correlation was revealed ( r =0.81) between hemoglobin oxygen saturation and oxyhemoglobin fraction in erythrocytes evaluated by Raman spectroscopy. Patients with CHD and patients with circulatory failure showed reduced oxygen-releasing capacity of hemoglobin and hemoglobin content and increased oxygen-binding capacity of hemoglobin, and hemoglobin affinity for oxygen. Significant differences from the control were observed only in patients with circulatory failure. It was found that hemoglobin content, hematocrit, and the shape of erythrocytes during CHD and circulatory failure did not differ from the control, whereas the area of erythrocytes was increased.

Surface-enhanced Raman spectroscopy (SERS) of living cells has rapidly become a powerful trend in biomedical diagnostics. It is a common belief that highly ordered, artificially engineered substrates are the best future decision in this field. This paper, however, describes an alternative successful solution, a new effortless chemical approach to the design of nanostructured silver and heterometallic continuous coatings with a stochastic "coffee ring'' morphology. The coatings are formed from an ultrasonic mist of aqueous diamminesilver hydroxide, free of reducing agents and nonvolatile pollutants, under mild conditions, at about 200-270 degrees C in air. They consist of 30-100 micrometer wide and 100-400 nm high silver rings composed, in turn, of a porous silver matrix with 10-50 nm silver grains decorating the sponge. This hierarchic structure originates from ultrasonic droplet evaporation, contact-line motion, silver(I) oxide decomposition and evolution of a growing ensemble of silver rings. The fabricated substrates are a remarkable example of a new scalable and low cost material suitable for SERS analyses of living cells. They evoke no hemolysis and reduce erythrocyte lateral mobility due to suitable "coffee ring'' sizes and a tight contact with the silver nanostructure. A high SERS enhancement, characteristic of pure silver rings, made it possible to record Raman scattering spectra from submembrane hemoglobin in its natural cellular environment inside single living erythrocytes, thus making the substrates promising for various biosensor chips.

We studied microfluidity and selective ion permeability of plasma membranes and O2-binding properties of erythrocytes of cosmonauts during early rehabilitation after a long-term space flight (LTSF). Microfluidity of plasma membranes in surface regions was found to undergo a reversible decrease during 13–15 days following LTSF, which was accompanied by a reversible increase in relative cholesterol content. Cosmonauts’ erythrocytes revealed an increased activity of Na/H-exchanger and KCa-channel as well as a decrease in number of discocytes and increase in number of echinocytes, stomatocytes and knizocytes. Total hemoglobin content as well as oxyhemoglobin content were lowered after the LTSF, while the affinity of hemoglobin to O2 was advanced. It is suggested that the changes in Hb properties, microfluidity and selective permeability of plasma membranes following the elevated cholesterol content in the membranes can decrease tissue supply with O2.