Construction and Characterization of HIV-1 env-Pseudoviruses of the Recombinant Form CRF63_02A and Subtype A6.

HIV-1 env-pseudoviruses are a useful tool in the search for antiviral drugs (entry inhibitors) and evaluation of the efficacy of HIV-1 vaccines. Given the high genetic variability of HIV-1, it is necessary to regularly update the panels of pseudoviruses in accordance with the emergence of new strains. Based on genetic variants of HIV-1 circulating in the regions of the Siberian Federal District, 13 HIV-1 env-pseudoviruses of recombinant form CRF63_02A and subtype A6 were obtained. Most pseudoviruses have been shown to be sensitive to neutralization by bnAbs VRC01, PGT126, and 10E8, moderately sensitive to bnAbs PG9 and 4E10, and resistant to bnAbs 2G12, PG16, and 2F5. All obtained variants of pseudoviruses are CCR5-tropic.

The Structural and Immunological Properties of Chimeric Proteins Containing HIV-1 MPER Sites.

The human immunodeficiency virus (HIV-1) poses a serious risk to global public health. The development of a safe and effective vaccine could stop the HIV/AIDS pandemic. Much of the research focused on HIV-1 prevention through vaccination is aimed at developing immunogens and immunization strategies to induce the formation of antibodies with neutralizing activity against a broad range of HIV-1 isolates (bNAbs). The objective of this study was to develop immunogens capable of targeting an immune response to MPER, one of the regions of bNAb binding in Env. Two immunogens carrying MPER fragments on their scaffolds (protein YkuJ Bacillus subtilis and artificial polypeptide TBI) were constructed. Circular dichroism spectroscopy was used to show that the secondary structure of the immunogens was consistent with their theoretical models. The antigenic structure of the MPER-TBI and YkuJ-MPER proteins was characterized using bNAbs that recognize HIV-1 MPER (2F5, 4E10, and 10E8). The rabbit model made it possible to show the immunogenicity of the constructed recombinant proteins. The resulting serum was found to be cross-reactive with immunogens carrying MPER. The constructs designed and characterized in this study can be used for targeting the humoral immune response to MPER, which is known to be one of the sites of HIV-1 vulnerability.

Characteristics of Artificial Immunogens Containing Peptide Mimotopes of HIV-1 Epitopes Recognized by Monoclonal Antibodies 2F5 and 2G12.

The paper describes construction of TBI-based recombinant proteins TBI-2F5 and TBI-2G12 that contain peptide mimotopes of HIV-1 epitopes recognized by broadly neutralizing antibodies 2F5 and 2G12, respectively. The capacity of the immunogens to induce neutralizing antibodies was evaluated. The sera of BALB/c mice immunized with recombinant proteins TBI, TBI-2F5, and TBI-2G12 neutralized HIV-1 env-pseudoviruses. Moreover, pooled serum from mice immunized with TBI-2F5 and TBI-2G12 neutralized env-pseudoviruses of HIV-1 subtype B more effectively than individual sera.

[The impact of the antibody 2F5 biotinylation on the selection of the peptides from combinatorial phage library].

The impact of monoclonal antibodies (mAb) biotinylation on the output and the repertoire of selected peptides in the biopanning procedure were tested. A comparative analysis of the peptides selected from phage library using the biotinylated and non-biotinylated mAb 2F5 was performed. It was shown that the output of peptides homologous to the native epitope was 1.7-fold higher for biotinylated antibodies, whereas the binding capacity of the selected phages with mAb 2F5 in ELISA was higher in the case of using non-biotinylated antibodies. It should be noted that the phages exposing peptides, which have 4-5 amino acid sequence similarity with the native epitope, demonstrate the highest binding affinity. The phages that expose peptides with 3 amino acid sequence similarity demonstrate different binding affinity: from the smallest to the largest. Based on the obtained data, it is safe to suggest that the rational biopanning may proceed in accordance with the task.

[Antiviral activity of aqueous extracts and polysaccharide fractions from mycelium and fruit bodies of higher fungi].

Sixty preparations of basidiomycetes (Ganoderma, Lentinus, Pleurotus, Laetiporus, Polyporus, Inonotus, Flammulina, Grifola, Trametes) were investigated with respect to their toxicity for Vero cells and antiviral activity. The antiviral activity was estimated with the use of the West Nile virus and type 2 Herpes simplex. It was shown that 11 preparations of Ganoderma, Lentinus and Pleurotus completely inhibited the infective activity in doses not lower than 1000 TCD50 (the West Nile virus) and 100 PPU (type 2 Herpes simplex). The antiviral activity of the preparations was likely due to the content of polysaccharides or their derivatives in the composition. It increased with increasing of the quantity of the total polysaccharide fraction or its concentration.

[Ca2+ transport by the membrane-bound monomeric form of Ca-ATPase of sarcoplasmic reticulum]. / Transport Ca2+ membranosviazannoi monomernoi formoi Ca-ATF-azy sarkoplazmaticheskogo retikuluma.

Chromatography of a mixture of phospholipid and Ca-dependent ATPase of sarcoplasmic reticulum solubilized with choleate in a column with anionite results in proteolyposome formation. It was shown that dilution of ATPase with phospholipids makes it possible to reduce the concentration of ATPase in the membrane. Using the bifunctional reagent difluorodinitrobenzene it was demonstrated that reduction of the ATPase concentration in the membrane leads to the dissociation of the enzyme to monomers. It was disclosed that the monomeric form of ATPase is capable of active Ca2+ transport.

[Dependence of thermal stability of Ca-ATPase from sarcoplasmic reticulum on its concentration in the membrane]. / Zavisimost' termostabil'nosti Ca-ATPazy sarkoplazmaticheskogo Retikuluma ot ee kontsentratsii v membrane.

Chromatography of a mixture of phospholipids and Ca-dependent ATPase from sarcoplasmic reticulum solubilized by cholate on a column with anionite results in a formation of proteoliposomes. Using the freeze-fracture technique, it was shown that the density of intramembrane particles in fractured proteoliposomes formed is reversely proportional to the lipid--protein ratio. This is indicative of possible changes in ATPase concentration in the membrane, when the above-mentioned method was used. It was shown that dilution of ATPase with phospholipids increases its thermal stability. This is probably due to aggregation of the ATPase molecules and increase in the length of the borderline of lipids around each enzyme molecule.