A cluster of carboxylic groups in PsbO protein is involved in proton transfer from the water oxidizing complex of Photosystem II

The hypothesis presented here for proton transfer away from the water oxidation complex of Photosystem II (PSII) is supported by biochemical experiments on the isolated PsbO protein in solution, theoretical analyses of better understood proton transfer systems like bacteriorhodopsin and cytochrome oxidase, and the recently published 3D structure of PS II (Pdb entry 1S5L). We propose that a cluster of conserved glutamic and aspartic acid residues in the PsbO protein acts as a buffering network providing efficient acceptors of protons derived from substrate water molecules. The charge delocalization of the cluster ensures readiness to promptly accept the protons liberated from substrate water. Therefore protons generated at the catalytic centre of PSII need not be released into the thylakoid lumen as generally thought. The cluster is the beginning of a localized, fast proton transfer conduit on the lumenal side of the thylakoid membrane. Proton-dependent conformational changes of PsbO may play a role in the regulation of both supply of substrate water to the water oxidizing complex and the resultant proton transfer.     

Quantitative assessment of intrinsic carbonic anhydrase activity and the capacity for bicarbonate oxidation in photosystem II

On the basis of equilibrium isotopic distribution experiments using (18)O-labeled water, it is generally accepted that water is the sole substrate for O(2) production by photosystem II (PSII). Nevertheless, recent studies indicating a direct interaction between bicarbonate and the donor side of PSII have been used to hypothesize that bicarbonate may have been a physiologically important substrate for O(2) production during the evolution of PSII [Dismukes, G. C., Klimov, V. V., Baranov, S. V., Kozlov, Y. N., DasGupta, J., and Tyryshikin, A. (2001) Proc. Natl. Acad. Sci. U.S.A. 98, 2170-2175]. To test out this hypothesis and to determine whether contemporary oxygenic organisms have the capacity to oxidize bicarbonate, we employed special rapid-mixing isotopic experiments using (18)O/(13)C-labeled bicarbonate to quantify the inherent carbonic anhydrase activity in PSII samples and the potential flux of oxygen from bicarbonate into the photosynthetically produced O(2). The measurements were made on PSII samples prepared from spinach, Thermosynechococcus elongatus, and Arthrospira maxima. For the latter organism, a strain was used that grows naturally in an alkaline, high (bi)carbonate soda lake in Africa. The results reveal that bicarbonate is not the substrate for O(2) production in these contemporary oxygenic photoautotrophs when assayed under single turnover conditions.     

Evaluation of new Cu(II) complexes as a novel class of inhibitors against plant carbonic anhydrase,glutathione reductase,and photosynthetic activity in photosystem II

Increasing inefficiency of production of important agricultural plants raises one of the biggest problems in the modern world. Herbicide application is still the best method of weed management. Traditional herbicides blocking only one of the plant metabolic pathways is ineffective due to the rapid growth of herbicide-resistant weeds. The synthesis of novel compounds effectively suppressing several metabolic processes, and therefore achieving the synergism effect would serve as the alternative approach to weed problem. For this reason, recently, we synthesized a series of nine novel Cu(II) complexes and four ligands, characterized them with different analyses techniques, and carried out their primary evaluation as inhibitors of photosynthetic electron transfer in spinach thylakoids (design, synthesis, and evaluation of a series of Cu(II) based metal–organic complexes as possible inhibitors of photosynthesis, J Photochem Photobiol B, submitted). Here, we evaluated in vitro inhibitory potency of these agents against: photochemistry and carbonic anhydrase activity of photosystem II (PSII); α-carbonic anhydrase from bovine erythrocytes; as well as glutathione reductase from chloroplast and baker’s yeast. Our results show that all Cu(II) complexes excellently inhibit glutathione reductase and PSII carbonic anhydrase activity. Some of them also decently inhibit PSII photosynthetic activity.     

Isolation of polymorphic tetranucleotide microsatellite markers for the brown anole (Anolis sagrei)

Eleven polymorphic microsatellite markers have been developed for the brown anole, Anolis sagrei. The number of alleles range from five to 14 per locus with the observed heterozygosity ranging from 0.46 to 0.92. These markers will be useful for analysis of questions concerning population genetic structure and reproductive behaviour.     

Quantification of the Blood Platelet Reactivity in the ADP-Induced Model of Non-Lethal Pulmonary Thromboembolism in Mice with the Use of Laser Doppler Flowmetry

IntroductionThe paper describes an alternative method for quantification of in vivo ADP-induced thromboembolism. The aim of the studies was to develop a method of quantification which would not require either extravasation or labelling of platelets. Our proposed approach is based on the monitoring of changes of blood flow with the use of laser Doppler flowmetry.ResultsThe injection of ADP resulted in a dose-dependent reduction of the blood flow in the mesentery. These responses were fully attributable to blood platelet aggregation, as shown by the lack of the effect in platelet-depleted mice, and significantly reduced responses in mice pretreated with cangrelor and eptifibatide. No platelet aggregate formation in mesenteric vessels was revealed by intravital microscopy, while ex vivo imaging showed accumulation of fluorescent labelled platelets in the lung.ConclusionsInjection of ADP to the venous system results in the formation of platelet aggregates predominantly in the lung. This results in reversible blood flow cessation in peripheral blood vessels. The measurement of this blood flow cessation in the mesentery allows indirect measurement of ADP-induced pulmonary thromboembolism. We suggest that this approach can be useful for in vivo screening for antiplatelet drug candidates.     

Galectin-3 interacts with membrane lipids and penetrates the lipid bilayer

The precise mechanism by which galectin-3 and other cytosolic proteins that lack signal peptides are secreted is yet to be elucidated. In the present analyses, we determined that galectin-3, a beta-galactoside binding protein, can interact directly with membrane lipids in solid phase binding assays. More interestingly, we determined by spectrophotometric methods that it can spontaneously penetrate the lipid bilayer of liposomes in either direction. These findings suggest that galectin-3 on its own has the capacity to traverse the lipid bilayer. Whereas the situation is rather simplified in liposomes, the interaction of galectin-3 with the plasma membrane may involve cholesterol-rich membrane domains where galectin-3 can be concentrated and form multimers or interact covalently with other proteins.