Abstract OR-8: Cryo-EM Structure of Mature Yellow Fever Virus

Background: Yellow fever virus (YFV) is the prototype virus of the genus Flavivirus. It is endemic to sub-Saharan Africa and tropical South America. YF disease ranges from asymptomatic to severe jaundice and hemorrhagic fever. The flavivirus virion core is enveloped by a lipid membrane with integrated membrane (M) proteins and envelope (E) proteins that form the outer surface of the virion. The Е protein provides stability to the viral particle and is responsible for early infection stages. Flaviviruses are heterogeneous in nature, which is related to their maturation process. Samples always contain mature, immature, half-mature, and damaged particles. Thus, cryo-EM is a method of choice for their structure determination. During the early stages of cryo-EM development, structures of flaviviruses were studied at 10–20 Å resolution. However, due to the progress of recent years, it became possible to determine flavivirus structures at a resolution of 5.6-2.6 Å. The cryo-EM method was used to obtain structural data of virions of dengue fever virus, Zika virus, TBE virus, etc. For YFV, only the cryo-EM structure of the immature virions at a low resolution of 25 Å was determined (Y. Zhang et al. 2003 doi:10.1093/emboj/cdg270). However, the structure of mature (most infectious) YFV particles is still unknown. Methods: Virus sample was produced in Vero cell culture. YFV-17D was inactivated and purified using ultracentrifugation. The concentration of viral particles in the target inactivated YFV-17D (iYFV-17D) was evaluated by spectrophotometry and by estimating the concentration of E protein determined by PAGE electrophoresis. Preliminary quality control of iYFV-17D sample was performed using negative staining TEM. Cryo-EM data were collected using cryo-TEM Krios (Thermo-Fisher, USA) at 300kV using DED Falcon II. Dataset was preprocessed using Warp. Further processing was performed in Relion 3.1 and CisTEM. Model building was carried out using Isolde, Phenix and Coot software. Results: A protocol of production and purification of highly concentrated (~2x1012) monodisperse inactivated iYFV-17D sample was developed. Cryo-EM structure of mature iYFV was solved at 4.1 Å resolution. The structure of YFV is similar to other known flavivirus structures, with 180 copies of protein E arranged in a herringbone pattern that makes up the icosahedral shell. Conclusion: The high-resolution structure of mature iYFV-17D allowed to elucidate special features of this flavivirus and may be useful for vaccine improvement and drug development

Functionally specialized human CD4+ T-cell subsets express physicochemically distinct TCRs

The organizational integrity of the adaptive immune system is determined by functionally discrete subsets of CD4+ T cells, but it has remained unclear to what extent lineage choice is influenced by clonotypically expressed T-cell receptors (TCRs). To address this issue, we used a high-throughput approach to profile the αβ TCR repertoires of human naive and effector/memory CD4+ T-cell subsets, irrespective of antigen specificity. Highly conserved physicochemical and recombinatorial features were encoded on a subset-specific basis in the effector/memory compartment. Clonal tracking further identified forbidden and permitted transition pathways, mapping effector/memory subsets related by interconversion or ontogeny. Public sequences were largely confined to particular effector/memory subsets, including regulatory T cells (Tregs), which also displayed hardwired repertoire features in the naive compartment. Accordingly, these cumulative repertoire portraits establish a link between clonotype fate decisions in the complex world of CD4+ T cells and the intrinsic properties of somatically rearranged TCRs

Comprehensive surface treatment of high-speed steel tool

One of the promising directions of hardening of high-speed steel tool is the creation on their surface of the layered structures with the gradient of physic-chemical properties between the wear-resistant coatings to the base material. Among the methods of such surface modification, a special process takes place based on the use of pulsed high-intensity charged particle beams. The high speed of heating and cooling allows structural-phase transformations in the surface layer, which cannot be realized in a stationary mode. The treatment was conducted in a RITM-SP unit, which constitutes a combination of a source of low-energy high-current electron beams “RITM” and two magnetron spraying systems on a single vacuum chamber. The unit enables deposition of films on the surface of the desired product and subsequent liquid-phase mixing of materials of the film and the substrate by an intense pulse electron beam. The article discusses features of the structure of the subsurface layer of high-speed steel M2, modified by surface alloying of a low-energy high-current electron beam, and its effect on the wear resistance of the tool when dry cutting hard to machine Nickel alloy. A significant decrease of intensity of wear of high-speed steel with combined treatment happens due to the displacement of the zone of wear and decrease the radius of rounding of the cutting edge because of changes in conditions of interaction with the material being treated

Increase of Productivity and Neutralization of Pathological Processes in Plants of Grain and Fruit Crops with the Help of Aqueous Solutions Activated by Plasma of High-Frequency Glow Discharge

In this work, we, for the first time, manufactured a plasma-chemical reactor operating at a frequency of 0.11 MHz. The reactor allows for the activation of large volumes of liquids in a short time. The physicochemical properties of activated liquids (concentration of hydrogen peroxide, nitrate anions, redox potential, electrical conductivity, pH, concentration of dissolved gases) are characterized in detail. Antifungal activity of aqueous solutions activated by a glow discharge has been investigated. It was shown that aqueous solutions activated by a glow discharge significantly reduce the degree of presence of phytopathogens and their effect on the germination of such seeds. Seeds of cereals (sorghum and barley) and fruit (strawberries) crops were studied. The greatest positive effect was found in the treatment of sorghum seeds. Moreover, laboratory tests have shown a significant increase in sorghum drought tolerance. The effectiveness of the use of glow-discharge-activated aqueous solutions was shown during a field experiment, which was set up in the saline semi-desert of the Northern Caspian region. Thus, the technology developed by us makes it possible to carry out the activation of aqueous solutions on an industrial scale. Water activated by a glow discharge exhibits antifungicidal activity and significantly accelerates the development of the grain and fruit crops we studied. In the case of sorghum culture, glow-discharge-activated water significantly increases drought resistance

Activation of Tissue Reparative Processes by Glow-Type Plasma Discharges as an Integral Part of the Therapy of Decubital Ulcers

The results of a clinical study of the complex treatment of pressure ulcers using the method of activation of reparative processes in tissues by cold plasma discharges initiated by high-frequency current are presented. Activation was carried out with a specialized device generating cold plasma discharges at frequencies of 0.11, 2.64, and 6.78 MHz. It was shown that the process of activation in the skin and muscle tissues of the bedsore zone proceeds most efficiently when using a current with a frequency of 6.78 MHz as compared to currents with a frequency of 2.64 and 0.11 MHz. For a needle electrode with a diameter of 0.3 mm, the optimal exposure parameters were power—(5.0 ± 1.5) W and time—(2.0–3.0) s. The results of the analysis of histological samples, histochemical, and bacteriological analysis confirmed the effect and showed the dynamics of the process of activation of reparative processes in the tissues of the bedsore wound under the influence of cold plasma discharges and a decrease in microbial contamination. The most pronounced effect of activation was formed during the period from 14 to 21 days. The effectiveness of therapy by the method of activation of reparative processes with cold plasma discharges, according to the criterion of the rate of wound healing, ranged from 14 to 16%, depending on the etiology of the decubitus wound. It is concluded that the activation of tissue reparative processes by glow-type plasma discharges as an integral part of the treatment of decubital ulcers is an effective link in the complex treatment of pressure sores

Activation of Tissue Reparative Processes by Glow-Type Plasma Discharges as an Integral Part of the Therapy of Decubital Ulcers

The results of a clinical study of the complex treatment of pressure ulcers using the method of activation of reparative processes in tissues by cold plasma discharges initiated by high-frequency current are presented. Activation was carried out with a specialized device generating cold plasma discharges at frequencies of 0.11, 2.64, and 6.78 MHz. It was shown that the process of activation in the skin and muscle tissues of the bedsore zone proceeds most efficiently when using a current with a frequency of 6.78 MHz as compared to currents with a frequency of 2.64 and 0.11 MHz. For a needle electrode with a diameter of 0.3 mm, the optimal exposure parameters were power—(5.0 ± 1.5) W and time—(2.0–3.0) s. The results of the analysis of histological samples, histochemical, and bacteriological analysis confirmed the effect and showed the dynamics of the process of activation of reparative processes in the tissues of the bedsore wound under the influence of cold plasma discharges and a decrease in microbial contamination. The most pronounced effect of activation was formed during the period from 14 to 21 days. The effectiveness of therapy by the method of activation of reparative processes with cold plasma discharges, according to the criterion of the rate of wound healing, ranged from 14 to 16%, depending on the etiology of the decubitus wound. It is concluded that the activation of tissue reparative processes by glow-type plasma discharges as an integral part of the treatment of decubital ulcers is an effective link in the complex treatment of pressure sores

Effect of alpha-lactalbumin and lactoferrin oleic acid complexes on chromatin structural organization

This work focuses on the study of multimeric alpha-lactalbumin oleic acid and lactoferrin oleic acid complexes. The purpose of the research is to study possible mechanisms involved in their pro-apoptotic activities, as seen in some tumor cell cultures. Complexes featuring oleic acid (OA) with human alpha-lactalbumin (hAl) or with bovine alpha-lactalbumin (bAl), and human lactoferrin (hLf) were investigated using small-angle neutron scattering (SANS). It was shown that while alpha-lactalbumin protein complexes were formed on the surface of polydisperse OA micelles, the lactoferrin complexes comprised a monodisperse system of nanoscale particles. Both hAl and hLf complexes appeared to interact with the chromatin of isolated nuclei affecting chromatin structural organization. The possible roles of these processes in the specific anti-tumor activity of these complexes are discussed