O. A. Kondakova

Reduction of tobacco mosaic virus accumulation in transgenic plants producing non-functional viral transport proteins

Transgenic plants producing the 30K temperature-sensitive transport protein (TP) of tobacco mosaic virus (TMV) mutant Ni2519 (affecting cell-to-cell transport) were found to: (i) be susceptible to wild-type TMV U1 at 24 degrees C (a permissive temperature for Ni2519 TP), (ii) acquire a certain level of resistance to TMV U1 accumulation when maintained at 33 degrees C (a non-permissive temperature for Ni2519 TP) and (iii) lose the resistance to wild-type TMV after their transfer from 33 degrees C to 24 degrees C. It is suggested that reversible temperature-dependent conformational changes in Ni2519 TP are responsible for these phenomena and that production of a TP which is only partially functional in transgenic plants confers on these plants a resistance to the virus owing to reduction of the level of cell-to-cell transport. Transgenic tobacco plants producing the 32K TP of brome mosaic virus (BMV) acquired resistance to TMV U1 suggesting that BMV TP is partially functional in tobacco plants.

Plant virus transport function: complementation by helper viruses is non-specific.

Summary The possibility of complementation of the cell-to-cell spread (within the inoculated leaf) between different related and unrelated plant viruses has been studied. Various tobamoviruses (tobacco mosaic, sunn-hemp mosaic, cucumber green mottle mosaic viruses and ‘tobamovirus from orchids’) can facilitate each others replication in non-permissive hosts or at a temperature non-permissive for transport of one of the virus partners, probably by complementation of transport functions. Complementation of movement also occurred between some, but not all unrelated viruses tested. The complementation in transport function seems to be non-specific: it can occur between viruses even if their putative transport proteins significantly differ in structure. Consequently these viruses were classified tentatively into different ‘transport groups’.

New Synthetic Thrombin Inhibitors: Molecular Design and Experimental Verification

Background The development of new anticoagulants is an important goal for the improvement of thromboses treatments. Objectives The design, synthesis and experimental testing of new safe and effective small molecule direct thrombin inhibitors for intravenous administration. Methods Computer-aided molecular design of new thrombin inhibitors was performed using our original docking program SOL, which is based on the genetic algorithm of global energy minimization in the framework of a Merck Molecular Force Field. This program takes into account the effects of solvent. The designed molecules with the best scoring functions (calculated binding energies) were synthesized and their thrombin inhibitory activity evaluated experimentally in vitro using a chromogenic substrate in a buffer system and using a thrombin generation test in isolated plasma and in vivo using the newly developed model of hemodilution-induced hypercoagulation in rats. The acute toxicities of the most promising new thrombin inhibitors were evaluated in mice, and their stabilities in aqueous solutions were measured. Results New compounds that are both effective direct thrombin inhibitors (the best KI was <1 nM) and strong anticoagulants in plasma (an IC50 in the thrombin generation assay of approximately 100 nM) were discovered. These compounds contain one of the following new residues as the basic fragment: isothiuronium, 4-aminopyridinium, or 2-aminothiazolinium. LD50 values for the best new inhibitors ranged from 166.7 to >1111.1 mg/kg. A plasma-substituting solution supplemented with one of the new inhibitors prevented hypercoagulation in the rat model of hemodilution-induced hypercoagulation. Activities of the best new inhibitors in physiological saline (1 µM solutions) were stable after sterilization by autoclaving, and the inhibitors remained stable at long-term storage over more than 1.5 years at room temperature and at 4°C. Conclusions The high efficacy, stability and low acute toxicity reveal that the inhibitors that were developed may be promising for potential medical applications.

Red Clover Mottle Comovirus B-RNA Spreads between Cells in Tobamovirus-infected Tissues

Summary B component RNA (B-RNA) of red clover mottle comovirus (RCMV) was not transported between cells of inoculated leaf tissue unless it was co-inoculated with the M component. However when host plants were infected with sunn-hemp mosaic tobamovirus (SHMV) or mutant Ni118 of tobacco mosaic virus (TMV) before superinoculation, RCMV B-RNA was transported readily between cells. Plants were infected with SHMV or Ni118 and, 5 days after superinoculation with RCMV B component, protoplasts were isolated from the infected leaves and inoculated with RCMV M component. About 30% of such protoplasts multiplied RCMV (identified by immunofluorescence microscopy) whereas only 3 to 5% of protoplasts from similarly treated plants not initially infected with SHMV or Ni118 multiplied RCMV. Thus B-RNA spread from mixedly infected (SHMV + B-RNA or Ni118 + B-RNA) cells in the absence of M-RNA to the neighbouring cells. RCMV B-RNA spread in leaves infected with the temperature-sensitive coat protein TMV mutant Ni118 and grown at non-permissive temperature. Thus TMV coat protein is not involved in enabling RCMV RNA to be transported.

A quantum-chemical study of the mechanism of ionic conductivity in alkali borate glasses

The incorporation of alkali metal oxide molecules into a continuous random network of vitreous boron oxide is considered within the cluster approximation at a semiempirical MNDO level. It is found that the oxygen atom of the Li2O molecule easily forms an additional bond with the threefold-coordinated boron atom with the formation of the BO4 tetrahedron. Different types of metal cation motion in alkali borate glasses are treated. It is revealed that the M+ cation in neutral systems is predominantly located near the edges of a BO4- tetrahedron on the outside of the B-O-B corner fragments and can readily change its position. The M+ cation in the neighborhood of the pentaborate grouping makes a complete turn without considerable expenditure of energy. The vibrations with a change in the rotation angle from 180° to 270° are possible in tetraborates, and, only in the neighborhood of triborate, one position is substantially more stable than the other positions. The cation can migrate from center to center along chains of a continuous glass network on the outside of the B-O-B corner fragments. In the pentaborate and tetraborate centers, the cation can migrate into another chain and change its direction of motion.

Production of the tobacco mosaic virus (TMV) transport protein in transgenic plants is essential but insufficient for complementing foreign virus transport: a need for the full-length TMV genome or some other TMV-encoded product

We have reported previously that tobamoviruses enable the transport of red clover mottle comovirus (RCMV) in tobacco plants normally resistant to RCMV. Here we show that RCMV transport does not take place in transgenic tobacco plants (line To-4) producing the 30K transport protein of tobacco mosaic virus (TMV), whereas the transport of the TMV Ls1 mutant, the cell-to-cell movement of which is temperature sensitive, is complemented in these plants. However, RCMV transport is observed when these transgenic plants are infected with both RCMV and TMV Ls1 at the non-permissive temperature (33 degrees C). It is suggested that (i) the hypothetical modification of transgenic plant plasmodesmata by the TMV 30K transport protein can specifically mediate the cell-to-cell movement of the homologous virus (TMV), but is insufficient to mediate RCMV transport; (ii) the presence of the full-length TMV genome or a certain TMV-encoded product(s) besides the 30K protein is essential for complementation of the RCMV transport function. The possibility that line To-4 might provide enough 30K protein to complement TMV Ls1 but not RCMV cannot be ruled out. During double infection the mutant 30K protein may, in concert with the wild-type 30K protein, provide the transport function for RCMV.

Computation of hydration free energies of organic solutes with an implicit water model.

A new approach for computing hydration free energies ΔGsolv of organic solutes is formulated and parameterized. The method combines a conventional PCM (polarizable continuum model) computation for the electrostatic component ΔGel of ΔGsolv and a specially detailed algorithm for treating the complementary nonelectrostatic contributions (ΔGnel). The novel features include the following: (a) two different cavities are used for treating ΔGel and ΔGnel. For the latter case the cavity is larger and based on thermal atomic radii (i.e., slightly reduced van der Waals radii). (b) The cavitation component of ΔGnel is taken to be proportional to the volume of the large cavity. (c) In the treatment of van der Waals interactions, all solute atoms are counted explicitly. The corresponding interaction energies are computed as integrals over the surface of the larger cavity; they are based on Lennard Jones (LJ) type potentials for individual solute atoms. The weighting coefficients of these LJ terms are considered as fitting parameters. Testing this method on a collection of 278 uncharged organic solutes gave satisfactory results. The average error (RMSD) between calculated and experimental free energy values varies between 0.15 and 0.5 kcal/mol for different classes of solutes. The larger deviations found for the case of oxygen compounds are probably due to a poor approximation of H‐bonding in terms of LJ potentials. For the seven compounds with poorest fit to experiment, the error exceeds 1.5 kcal/mol; these outlier points were not included in the parameterization procedure. Several possible origins of these errors are discussed.

Quantum chemical study of the network modification in vitreous B2O3

Abstract The possible ways of rearrangement of a random B2O3 network were investigated in a cluster approach on semiempirical MNDO level. Interlayer interaction was found to be able to form metastable hypervalent structures with fourfold-coordinated B and threefold-coordinated O atoms. The B–O bonds in such configurations were weaker than ordinary ones and are similar for intra- and interlayer interactions. As a result, in such structures the moderate-energy switching of B–O bonds and linking of neighbor layers are possible. Destruction of the network is shown to start at puckered regions of the network and at the boroxol rings. The formation of fragments with -BO defects is the most energetically favorable, and we expect to find a band in the region of 2000 cm−1 (vibration of BO double bond) in the IR-spectrum of molten B2O3. Implantation of a -BO fragment into the B–O bond of the neighbor layer with creation of interlayer linkage is possible. As a result the B2O3 network will have the properties of a three-dimensional structure.

High-performance atomistic modeling of optical thin films deposited by energetic processes

In this paper we present a computationally effective approach to classical molecular dynamic simulation of thin film growth with orientation on cluster supercomputing facilities. The goal of the developed approach is to investigate structural heterogeneities of thin films deposited on substrates at a nanoscale level. These heterogeneities depend on the experimental conditions of a deposition process being used. They have essential influence on practical properties of thin films and their modeling is important for achieving further progress in thin film optical technology. The presented research is focused on silicon dioxide thin films growth. A special force field, oriented on the atomistic description of the silicon dioxide deposition on fused silica substrate, has been developed and applied to the molecular dynamic simulation with the GROMACS package. The validity of the developed simulation approach is verified using atomic clusters consisting of up to 106 atoms and having characteristic dimensions of up to 30 nm. Its computational efficiency is tested using up to 2048 cores. The dependence of achievable efficiency on model parameters is discussed.

Study of rubella candidate vaccine based on a structurally modified plant virus

Abstract A novel rubella candidate vaccine based on a structurally modified plant virus – spherical particles (SPs) – was developed. SPs generated by the thermal remodelling of the tobacco mosaic virus are promising platforms for the development of vaccines. SPs combine unique properties: biosafety, stability, high immunogenicity and the effective adsorption of antigens. We assembled in vitro and characterised complexes (candidate vaccine) based on SPs and the rubella virus recombinant antigen. The candidate vaccine induced a strong humoral immune response against rubella. The IgG isotypes ratio indicated the predominance of IgG1 which plays a key role in immunity to natural rubella infection. The immune response was generally directed against the rubella antigen within the complexes. We suggest that SPs can act as a platform (depot) for the rubella antigen, enhancing specific immune response. Our results demonstrate that SPs‐antigen complexes can be an effective and safe candidate vaccine against rubella. Graphical abstract Figure. No caption available. HighlightsIn vitro assembly and characterisation of a rubella candidate vaccine based on modified plant virus (spherical particles).Candidate vaccine induces efficient humoral immune response against rubella.Acute and chronic toxicity studies demonstrate biosafety of candidate vaccine.Candidate vaccine demonstrated good results and can be considered as a promising safe tool against the rubella virus.