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The injectable hydrated calcium phosphate bone-like paste (hCPP) was developed with suitable rheological characteristics, enabling unhindered injection through standard 23G needles. In vitro assays showed the cytocompatibility of hCPP with mesenchymal embryonic C3H10T1/2 cell cultures. The hCPP was composed of aggregated micro-sized particles with sphere-like shapes and low crystallinity. The ability of hCPP particles to adsorb serum proteins (FBS) was investigated. The hCPP demonstrated high protein adsorption capacity, indicating its potential in various biomedical applications. The results of the in vivo assay upon subcutaneous injection in Wistar rats indicated nontoxicity and biocompatibility of experimental hCPP, as well as gradual resorption of hCPP, comparable to the period of bone regeneration. The data obtained are of great interest for the development of commercial highly effective osteoplastic materials for bone tissue regeneration and augmentation.

X-ray imaging of virus particles at European XFEL could eventually allow solving their complete structure, potentially approaching resolution of other structural virology methods. To achieve this ambitious goal with today’s technologies, several mL of purified virus suspension containing at least 1012particles per mL are required. Such large amounts of concentrated suspension have never before been obtained for enveloped viruses. Tick-borne encephalitis virus (TBEV) represents an attractive model system for the development of enveloped virus purification and concentration protocols, given the availability of large amounts of inactivated virus material provided by vaccine manufacturing facilities. Here we present the development of a TBEV vaccine purification and concentration scheme combined with a quality control protocol allowing substantial amounts of highly concentrated non-aggregated suspension to be obtained. Preliminary single particle imaging experiments were performed for this sample at European XFEL, showing distinct diffraction patterns.

A Lyapunov-Krasovskii functional with prescribed derivative whose construction does not require the stability of the system is introduced. It leads to the presentation of stability/instability theorems. By evaluating the functional at initial conditions depending on the fundamental matrix we are able to present necessary and sufficient stability conditions expressed exclusively in terms of the delay Lyapunov matrix for integral delay systems. Some examples illustrate and validate the stability conditions., This paper has been submitted to International Journal of Robust and Nonlinear Control

The severe COVID‐19 pandemic drives the research toward the SARS‐CoV‐2 virion structure and the possible therapies against it. Here, we characterized the β‐propiolactone inactivated SARS‐CoV‐2 virions using transmission electron microscopy (TEM) and atomic force microscopy (AFM). We compared the SARS‐CoV‐2 samples purified by two consecutive chromatographic procedures (size exclusion chromatography [SEC], followed by ion‐exchange chromatography [IEC]) with samples purified by ultracentrifugation. The samples prepared using SEC and IEC retained more spikes on the surface than the ones prepared using ultracentrifugation, as confirmed by TEM and AFM. TEM showed that the spike (S) proteins were in the pre‐fusion conformation. Notably, the S proteins could be recognized by specific monoclonal antibodies. Analytical TEM showed that the inactivated virions retained nucleic acid. Altogether, we demonstrated that the inactivated SARS‐CoV‐2 virions retain the structural features of native viruses and provide a prospective vaccine candidate., SARS‐CoV‐2 virions inactivated with β‐propiolactone were analyzed using transmission electron microscopy (TEM) and atomic force microscopy.The S proteins could be recognized by specific monoclonal antibodies.Analytical TEM showed that the inactivated virions retained nucleic acid.

Tick-borne encephalitis virus (TBEV) is a RNA-containing enveloped virus, a member of the Flaviviridae family. Here we describe a detailed analysis of the size and structure of inactivated TBEV (the Sofyin-Chumakov TBEV strain, used in vaccines). Four analytical methods were used to analyze individual TBEV particles—negative staining TEM, cryo-EM, AFM, and NTA. All methods confirmed that the particles were monodisperse, and their mean size was ~50 nm. Cryo-EM data were used to obtain a 3D electron density model of the virus with clearly distinguishable E-proteins. STEM-EELS analysis detected phosphorous in the particles, which was interpreted as the RNA signal. Altogether, the described analytical procedures can be valuable for the further analysis of vaccine inactivated virus samples.

Tick-borne encephalitis virus (TBEV) is an enveloped RNA virus, a member of the genus Flavivirus (family Flaviviridae). Here, we provide a detailed analysis of the size and structure of the inactivated TBEV vaccine strain Sofjin-Chumakov. Four analytical methods were used to analyze individual TBEV particles—negative staining TEM, cryo-EM, atomic force microscopy (AFM), and nanoparticle tracking analysis (NTA). All methods confirmed that the particles were monodisperse and that their mean size was ~50 nm. Cryo-EM data allowed us to obtain a 3D electron density model of the virus with clearly distinguishable E protein molecules. STEM-EELS analysis detected phosphorus in the particles, which was interpreted as an indicator of RNA presence. Altogether, the described analytical procedures can be valuable for the characterization of inactivated vaccine virus samples.

We present a new necessary and sufficient condition of exponential stability of neutral-type linear time invariant (LTI) systems with one delay. The new stability criterion is given in terms of a block matrix, which exclusively depends on the delay Lyapunov matrix and is of finite dimension.

Theoretical, methodological, and empirical substantiation of the problem of values’ development of a serviceman’s family after the implementation of the civil-military relations reform in Russia in 2012 is presented. On the basis of the study from the sociological survey “Family Culture of the Russian Servicemen”, the trends in updating family values in the post-reform period are revealed. The identification of the changes in the values of a serviceman’s family made it possible to clarify and reveal the structure of military families and the ways transforming not only the vital interests and priorities of this family, but also the changes in labor motives, needs and social needs of their members, as a whole. It is proved that the value priorities of military personnel in modern conditions should be characterized as one of the generalized indicators of their professional cultural potential. With regards to the priorities of life values, military families are conventionally divided into several groups: 1) 20% “money savers” (the families whose members have predominant values related to material well-being, the desire for high earnings); 2) 17% “health-oriented” (the families that have the most important priority in maintaining health, developing the physical potential of their members); 3) 10% “strong family members” (the families whose members have predominant values related to their own family, striving to maintain good relations in the family); 4) 10% “career professionals” (the families whose life activities are aimed at ensuring the conditions for career growth of military personnel, and increasing their military educational potential); 5) 6% “self-sacrificing patriots” (the families that focus on finding things to their liking, realizing themselves in interesting military activities, meeting the need for military intellectual work, subordinating their own interests, sacrificing them for the good of society, the country and the state).

In this article, necessary stability conditions for integral delay systems with distributed single delay are presented. Based on new properties that connect the Lyapunov delay matrix and the fundamental matrix of this class of equations, a complete type Lyapunov–Krasovskii functional is introduced. It allows us to present necessary stability conditions in terms of the delay Lyapunov matrix. The result is illustrated by some examples.

For the experimental study of the role of the R65 residue in the stabilization of β-lactamases with the substitution of M182T, predicted by the analysis of the residue interaction networks (RINs), homogeneous preparations of the recombinant TEM-type β-lactamases with substitutions of R65L, M182A, and a combination of R65L and M182T mutations were obtained. The kinetic parameters of these enzymes were determined for penicillin, ceftazidime, cephalothin and CENTA. None of the investigated substitutions changed the substrate specificity of enzymes against β-lactam antibiotics. The substitution R65L leads to a decrease in thermal stability; the substitution of M182A and the combination of substitutions of R65L and M182T improve the thermal stability of β-lactamase in comparison with the wild-type enzyme TEM-1. Using differential scanning calorimetry, it was determined the enthalpy (ΔH, kJ/mol) and the denaturation temperature (Tmp, °C) for β-lactamases TEM-1, TEM(M182A), TEM-135(M182T), and TEM (R65L + M182T), which are, respectively, 554.0 and 50.8, 573.7 and 51.7, 654.4 and 55.9, and 647.4 and 51.9. The hypothesis of molecular mechanism, explaining the stabilizing role of M182T substitution in TEM type β‑lactamases, was supplemented by the effect of changing the conformation of the R65 residue and the appearance of its new contacts with the residues of the Ω-loop of β-lactamase.

Bacterial beta-lactamases and carbapenemases confer resistance to beta-lactam antibiotics, including penicillins, cephalosporins, carbapenems, and monobactams. Their wide distribution among the bacteria that cause infectious diseases in humans and animals represent a global threat. We have developed a biochip with colorimetric detection based on horseradish peroxidase for the simultaneous identification of genes for all clinically relevant class A beta-lactamases and carbapenemases of classes A, B, and D, including 25 single substitutions in the nucleotide sequence encoding the key amino acid substitutions in class A beta-lactamases. The conditions for allele-specific hybridization of biotin-labeled target DNA with oligonucleotide probes immobilized on the surface of the biochip have been optimized. A method of multiplex amplification of all of the studied genes in one reaction with the simultaneous incorporation of biotin was developed to obtain the target DNA. The biochip was validated with mixtures of the beta-lactamase and carbapenemase genes, as well as 68 DNA samples isolated from clinical strains of gram-negative bacteria. The total DNA sample analysis time was ~4 h. A high specificity of the identification of genes in mixtures was demonstrated, which can be used in the study of multidrug-resistant bacteria.

The resistance of bacteria to β-lactam antibiotics is primarily caused by the production of β-lactamases. Here, novel crystal structures of the native β-lactamase TEM-171 and two complexes with the widely used inhibitor tazobactam are presented, alongside complementary data from UV spectroscopy and fluorescence quenching. The six chemically identical β-lactamase molecules in the crystallographic asymmetric unit displayed different degrees of disorder. The tazobactam intermediate was covalently bound to the catalytic Ser70 in the trans-enamine configuration. While the conformation of tazobactam in the first complex resembled that in published β-lactamase–tazobactam structures, in the second complex, which was obtained after longer soaking of the native crystals in the inhibitor solution, a new and previously unreported tazobactam conformation was observed. It is proposed that the two complexes correspond to different stages along the deacylation path of the acyl-enzyme intermediate. The results provide a novel structural basis for the rational design of new β-lactamase inhibitors.

The problem of autism spectrum disorders (ASD) and early childhood autism, in particular, has become one of the most discussed problems of child psychiatry in recent years. For example, a search for publications with the keywords "Autism Spectrum Disorder" in PubMed for 2019 offers 4523 results, and for 2010 1795 links. This is due to a number of reasons.

The design of effective target-specific drugs for COVID-19 treatment has become an intriguing challenge for modern science. The SARS-CoV-2 main protease, Mpro, responsible for the processing of SARS-CoV-2 polyproteins and production of individual components of viral replication machinery, is an attractive candidate target for drug discovery. Specific Mpro inhibitors have turned out to be promising anticoronaviral agents. Thus, an effective platform for quantitative screening of Mpro-targeting molecules is urgently needed. Here, we propose a pre–steady-state kinetic analysis of the interaction of Mpro with inhibitors as a basis for such a platform. We examined the kinetic mechanism of peptide substrate binding and cleavage by wild-type Mpro and by its catalytically inactive mutant C145A. The enzyme induces conformational changes of the peptide during the reaction. The inhibition of Mpro by boceprevir, telaprevir, GC-376, PF-00835231, or thimerosal was investigated. Detailed pre–steady-state kinetics of the interaction of the wild-type enzyme with the most potent inhibitor, PF-00835231, revealed a two-step binding mechanism, followed by covalent complex formation. The C145A Mpro mutant interacts with PF-00835231 approximately 100-fold less effectively. Nevertheless, the binding constant of PF-00835231 toward C145A Mpro is still good enough to inhibit the enzyme. Therefore, our results suggest that even noncovalent inhibitor binding due to a fine conformational fit into the active site is sufficient for efficient inhibition. A structure-based virtual screening and a subsequent detailed assessment of inhibition efficacy allowed us to select two compounds as promising noncovalent inhibitor leads of SARS-CoV-2 Mpro.

The increasing antibiotic resistance is a clinical problem worldwide. Numerous Gram-negative bacteria have already become resistant to the most widely used class of antibacterial drugs, β-lactams. One of the main mechanisms is inactivation of β-lactam antibiotics by bacterial β-lactamases. Appearance and spread of these enzymes represent a continuous challenge for the clinical treatment of infections and for the design of new antibiotics and inhibitors. Drug repurposing is a prospective approach for finding new targets for drugs already approved for use. We describe here the inhibitory potency of known detoxifying antidote 2,3-dimercaptopropane-1-sulfonate (unithiol) against metallo-β-lactamases. Unithiol acts as a competitive inhibitor of meropenem hydrolysis by recombinant metallo-β-lactamase NDM-1 with the KI of 16.7 µM. It is an order of magnitude lower than the KI for l-captopril, the inhibitor of angiotensin-converting enzyme approved as a drug for the treatment of hypertension. Phenotypic methods demonstrate that the unithiol inhibits natural metallo-β-lactamases NDM-1 and VIM-2 produced by carbapenem-resistant K. pneumoniae and P. aeruginosa bacterial strains. The 3D full atom structures of unithiol complexes with NDM-1 and VIM-2 are obtained using QM/MM modeling. The thiol group is located between zinc cations of the active site occupying the same place as the catalytic hydroxide anion in the enzyme–substrate complex. The sulfate group forms both a coordination bond with a zinc cation and hydrogen bonds with the positively charged residue, lysine or arginine, responsible for proper orientation of antibiotics upon binding to the active site prior to hydrolysis. Thus, we demonstrate both experimentally and theoretically that the unithiol is a prospective competitive inhibitor of metallo-β-lactamases and it can be utilized in complex therapy together with the known β-lactam antibiotics.