Structural insights into light-driven anion pumping in cyanobacteria.

Transmembrane ion transport is a key process in living cells. Active transport of ions is carried out by various ion transporters including microbial rhodopsins (MRs). MRs perform diverse functions such as active and passive ion transport, photo-sensing, and others. In particular, MRs can pump various monovalent ions like Na+, K+, Cl-, I-, NO3-. The only characterized MR proposed to pump sulfate in addition to halides belongs to the cyanobacterium Synechocystis sp. PCC 7509 and is named Synechocystis halorhodopsin (SyHR). The structural study of SyHR may help to understand what makes an MR pump divalent ions. Here we present the crystal structure of SyHR in the ground state, the structure of its sulfate-bound form as well as two photoreaction intermediates, the K and O states. These data reveal the molecular origin of the unique properties of the protein (exceptionally strong chloride binding and proposed pumping of divalent anions) and sheds light on the mechanism of anion release and uptake in cyanobacterial halorhodopsins. The unique properties of SyHR highlight its potential as an optogenetics tool and may help engineer different types of anion pumps with applications in optogenetics.

Principal component analysis highlights the influence of temperature, curvature and cholesterol on conformational dynamics of lipids.

Membrane lipids are inherently highly dynamic molecules. Currently, it is difficult to probe the structures of individual lipids experimentally at the timescales corresponding to atomic motions, and consequently molecular dynamics simulations are used widely. In our previous work, we have introduced the principal component analysis (PCA) as a convenient framework for comprehensive quantitative description of lipid motions. Here, we present a newly developed open source script, PCAlipids, which automates the analysis and allows us to refine the approach and test its limitations. We use PCAlipids to determine the influence of temperature, cholesterol and curvature on individual lipids, and show that the most prominent lipid tail scissoring motion is strongly affected by these factors and allows tracking of phase transition. Addition of cholesterol affects the conformations and selectively changes the dynamics of lipid molecules, impacting the large-amplitude motions. Introduction of curvature biases the conformational ensembles towards more extended structures. We hope that the developed approach will be useful for understanding the molecular basis of different processes occurring in lipid membrane systems and will stimulate development of complementary experimental techniques probing the conformations of individual lipid molecules.

High-resolution structural insights into the heliorhodopsin family.

Rhodopsins are the most abundant light-harvesting proteins. A new family of rhodopsins, heliorhodopsins (HeRs), has recently been discovered. Unlike in the known rhodopsins, in HeRs the N termini face the cytoplasm. The function of HeRs remains unknown. We present the structures of the bacterial HeR-48C12 in two states at the resolution of 1.5 Å, which highlight its remarkable difference from all known rhodopsins. The interior of HeR's extracellular part is completely hydrophobic, while the cytoplasmic part comprises a cavity (Schiff base cavity [SBC]) surrounded by charged amino acids and containing a cluster of water molecules, presumably being a primary proton acceptor from the Schiff base. At acidic pH, a planar triangular molecule (acetate) is present in the SBC. Structure-based bioinformatic analysis identified 10 subfamilies of HeRs, suggesting their diverse biological functions. The structures and available data suggest an enzymatic activity of HeR-48C12 subfamily and their possible involvement in fundamental redox biological processes.

Unusual features of the c-ring of F1FO ATP synthases.

Membrane integral ATP synthases produce adenosine triphosphate, the universal "energy currency" of most organisms. However, important details of proton driven energy conversion are still unknown. We present the first high-resolution structure (2.3 Å) of the in meso crystallized c-ring of 14 subunits from spinach chloroplasts. The structure reveals molecular mechanisms of intersubunit contacts in the c14-ring, and it shows additional electron densities inside the c-ring which form circles parallel to the membrane plane. Similar densities were found in all known high-resolution structures of c-rings of F1FO ATP synthases from archaea and bacteria to eukaryotes. The densities might originate from isoprenoid quinones (such as coenzyme Q in mitochondria and plastoquinone in chloroplasts) that is consistent with differential UV-Vis spectroscopy of the c-ring samples, unusually large distance between polar/apolar interfaces inside the c-ring and universality among different species. Although additional experiments are required to verify this hypothesis, coenzyme Q and its analogues known as electron carriers of bioenergetic chains may be universal cofactors of ATP synthases, stabilizing c-ring and prevent ion leakage through it.

New Insights on Signal Propagation by Sensory Rhodopsin II/Transducer Complex.

The complex of two membrane proteins, sensory rhodopsin II (NpSRII) with its cognate transducer (NpHtrII), mediates negative phototaxis in halobacteria N. pharaonis. Upon light activation NpSRII triggers a signal transduction chain homologous to the two-component system in eubacterial chemotaxis. Here we report on crystal structures of the ground and active M-state of the complex in the space group I212121. We demonstrate that the relative orientation of symmetrical parts of the dimer is parallel ("U"-shaped) contrary to the gusset-like ("V"-shaped) form of the previously reported structures of the NpSRII/NpHtrII complex in the space group P21212, although the structures of the monomers taken individually are nearly the same. Computer modeling of the HAMP domain in the obtained "V"- and "U"-shaped structures revealed that only the "U"-shaped conformation allows for tight interactions of the receptor with the HAMP domain. This is in line with existing data and supports biological relevance of the "U" shape in the ground state. We suggest that the "V"-shaped structure may correspond to the active state of the complex and transition from the "U" to the "V"-shape of the receptor-transducer complex can be involved in signal transduction from the receptor to the signaling domain of NpHtrII.

ESR - a retinal protein with unusual properties from Exiguobacterium sibiricum.

This review covers the properties of a retinal protein (ESR) from the psychrotrophic bacterium Exiguobacterium sibiricum that functions as a light-driven proton pump. The presence of a lysine residue at the position corresponding to intramolecular proton donor for the Schiff base represents a unique structural feature of ESR. We have shown that Lys96 successfully facilitates delivery of protons from the cytoplasmic surface to the Schiff base, thus acting as a proton donor in ESR. Since proton uptake during the photocycle precedes Schiff base reprotonation, we conclude that this residue is initially in the uncharged state and acquires a proton for a short time after Schiff base deprotonation and M intermediate formation. Involvement of Lys as a proton donor distinguishes ESR from the related retinal proteins - bacteriorhodopsin (BR), proteorhodopsin (PR), and xanthorhodopsin (XR), in which the donor function is performed by residues with a carboxyl side chain. Like other eubacterial proton pumps (PR and XR), ESR contains a histidine residue interacting with the proton acceptor Asp85. In contrast to PR, this interaction leads to shift of the acceptor's pKa to more acidic pH, thus providing its ability to function over a wide pH range. The presence of a strong H-bond between Asp85 and His57, the structure of the proton-conducting pathways from cytoplasmic surface to the Schiff base and to extracellular surface, and other properties of ESR were demonstrated by solving its three-dimensional structure, which revealed several differences from known structures of BR and XR. The structure of ESR, its photocycle, and proton transfer reactions are discussed in comparison with homologous retinal proteins.

[Metal content of local foods in Pechenga district of Murmansk region].

Within international project KolArctic "Food safety and health in frontier area of Russia, Finland and Norway", the study covered local food sampling in Pechenga district of Murmansk region during autumn of 2013, including fish (from 6 lakes), game, mushrooms, wild and cultivated berries, vegetables from private gardens situated at various distances from Nickel and Zapolarnyi settlements, also polling among 400 residents. Levels of 13 metals in the foods were assessed in "Taifun" laboratory. MACs for cadmium was 1.5-2 times exceeded in mushrooms (lamellate and tubular), that for mercury was up to 3 times exceeded in aspen mushrooms. Fresh-water fish appeared to contain the highest levels of mercury, close to MAC. Assessing levels of other metals that were previously normalized in USSR, the findings are 1.5 times exceeded MAC for copper in milk mushrooms, MAC for nickel was 4.5 times exceeded in wild berries, 2.5 times exceeded in cultivated berries, 2 times exceeded in potatoes and 2.5 to 30 times exceeded in mushrooms. Mushrooms have to be considered as major sorbents of total complex of the metals under study. Fresh-water fish is foodstuff mostly contaminated with mercury. Highly toxic nickel has to be considered as a major factor of exposure (and health risk) among the population under study. The data obtained help to specify recommendations on restricting some food items and reducing health risk for the residents subjected to industrial releases from "Pechenganickel" enterprise.

High-resolution structure of a membrane protein transferred from amphipol to a lipidic mesophase.

Amphipols (APols) have become important tools for the stabilization, folding, and in vitro structural and functional studies of membrane proteins (MPs). Direct crystallization of MPs solubilized in APols would be of high importance for structural biology. However, despite considerable efforts, it is still not clear whether MP/APol complexes can form well-ordered crystals suitable for X-ray crystallography. In the present work, we show that an APol-trapped MP can be crystallized in meso. Bacteriorhodopsin (BR) trapped by APol A8-35 was mixed with a lipidic mesophase, and crystallization was induced by adding a precipitant. The crystals diffract beyond 2 Å. The structure of BR was solved to 2 Å and found to be indistinguishable from previous structures obtained after transfer from detergent solutions. We suggest the proposed protocol of in meso crystallization to be generally applicable to APol-trapped MPs.

[Evolutionary admonition: allergy].

The paper concerns phylogeny development of allergic reactivity, the most probable antecedents of allergic IgE antibodies, high affinity IgE receptor (Fc(epsilon)RI), presents comparative character of IgE - Fc(epsilon)RI and IgY- CHIR-AB1 interactions. The paper has given an insight of allergy as evolutionary selected reactivity for highly organized animals. This reactivity is directed to organization of allergen-specific inflammation and serves as biologically expedient, high-specific and high-sensitive reaction in response to allergen entering into the organism because of barrier tissue dysfunction. Such insight has raised a question on consequences of thoroughgoing allergy reactivity elimination for highly organized animals and their posterity.

Ground state structure of D75N mutant of sensory rhodopsin II in complex with its cognate transducer.

The complex of sensory rhodopsin II (NpSRII) with its cognate transducer (NpHtrII) mediates negative phototaxis in halobacteria Natronomonas pharaonis. Upon light activation NpSRII triggers, by means of NpHtrII, a signal transduction chain homologous to the two component system in eubacterial chemotaxis. Here we report on the crystal structure of the ground state of the mutant NpSRII-D75N/NpHtrII complex in the space group I212121. Mutations of this aspartic acid in light-driven proton pumps dramatically modify or/and inhibit protein functions. However, in vivo studies show that the similar D75N mutation retains functionality of the NpSRII/NpHtrII complex. The structure provides the molecular basis for the explanation of the unexpected observation that the wild and the mutant complexes display identical physiological response on light excitation.

[Molecular basis of allergy control].

The article describes the picture of the interaction between IgE and its high-affinity receptor (Fc(epsilon)RI) leading to cellular triggering and clinical manifestations of IgE-dependent allergy. New understanding of the key event of an allergic response has enabled new pharmacological interventions at this stage.

[Comparative antihistamine and anti-allergic effects of various antihistamine preparations]. / Sravnitel'noe izuchenie antigistaminnoi i antiallergicheskoi aktivnosti nekotorykh protivogistaminnykh preparatov.

AIM: To compare antihistaminic and antiallergic activity of antihistaminic drugs of the latest generation (ebastin, cetirisine, fexofenadine, loratadine) and antihistaminic drugs of the first generation (clemastin) in the same patients with pollenosis. MATERIAL AND METHODS: Skin prick-titration with 10-dilution histamine and specific allergen, provocative nasal titration with 2-dilution histamine and allergen before and after a single intake of H1-antagonists were made in 30 patients in stable clinical remission of pollenosis during maximal antihistamine activity of the above drugs. RESULTS: Systemic administration of the known H1-antagonists suppresses histamine sensitivity of both skin and nasal mucosa in the same degree. Drugs with more potent antihistaminic activity (fexofenadin and cetirisin) inhibited allergen-induced reactions more effectively. The order of the tested drugs by suppression of allergen-provoked skin and nasal reactions (by lowering antiallergic activity) is the following: fexofenadin and cetirisin > ebastin and loratadin > clemastin. CONCLUSION: The above drugs of the latest generation seem to posses antiallergic activity not only due to antihistaminic effect but also due to other mechanisms. Different suppressive action of H1-antagonists reflects also individual sensitivity to different drugs. The factor of individual sensitivity of the patients to a pharmacological action of the drug may be crucial in the selection of the most effective medicine for each patient. This is confirmed by the data of individual sensitivity of the patient to antihistaminic and antiallergic action of H1-antagonists. The illustrated method may be helpful for individual selection of H1-antagonists for treatment of patients with allergic diseases.

[Therapeutic effectiveness of histaglobin preparations in patients with allergic rhinitis and chronic urticaria]. / Terapevticheskaia éffektivnost' preparatov gistaglobina u bol'nykh allergicheskim rinitom i khronicheskoi krapivnitsei.

AIM: To evaluate therapeutic effects of histaglobin drugs (histaglobin and histaglobin-triplex) in patients with continuous allergic rhinitis (CAR) and chronic recurrent (idiopathic) urticaria (CRU). MATERIALS AND METHODS: The drugs were given to 45 patients with CAR and high sensitivity to household allergens confirmed by cutaneous diagnostic tests or high IgE level, and 40 patients with CRU with typical cutaneous lesions. The drugs were injected subcutaneously (a total of 6 injections, 2 ml of solution each) with the interval 2-3 or 3-4 days. Histaglobin was given as 12 mg of normal human immunoglobulin + 0.00015 mg of histamine dihydrochloride. Histamine-triplex as 36 mg of immunoglobulin + 0.00045 mg of histamine dihydrochloride. The treatment was repeated in a months (3 injections). Clinical response was assessed in scores by the scale of clinical symptoms. RESULTS: Histaglobin relieved symptoms of CAR (from 7.34 +/- 0.095 to 1.7 +/- 0.04 scores on the treatment day 28, p < 0.01). After the repeated course CAR symptoms attenuated to 1.6 +/- 0.057 scores). Extranasal symptoms significantly reduced too. Excellent and good results were achieved in 80% of the patients. Positive results were also obtained in 82.5% of CRU patients. Tolerance of histaglobin and histaglobin-triplex was good. CONCLUSION: In the tested regimen, both histaglobin and histaglobin-triplex proved effective in CAR and CRU when routine treatment failed.

[Chemical modification of allergen leading to changes in its epitopic activity]. / Khimicheskaia modifikatsiia allergena, vedushchaia k izmeneniiu ego épitopnoi aktivnosti.

Modification of a model allergen ovalbumin (OA) with succinylation led to a decrease of its allergenicity measured by passive cutaneous anaphylaxis reaction, RAST inhibition assay and basophil histamine release. Modified OA stimulated OA-specific T-cell hybrid 3DO-548 to produce IL-2 at the same level as in case of non-modified OA. Modified OA did not induce anti-OA IgE, but did induce anti-OA IgG antibodies. This approach to chemical modification of allergen-selective blockade of B-cell epitopes while not affecting T-cell epitopes suggests new opportunities in creation of safe and effective allergovaccines.