Wuhan Sequence-Based Recombinant Antigens Expressed in E. coli Elicit Antibodies Capable of Binding with Omicron S-Protein.

The development of cross-reactive vaccines is one of the central aims of modern vaccinology. Continuous mutation and the emergence of new SARS-CoV-2 variants and subvariants create the problem of universal coronavirus vaccine design. Previously, the authors devised three recombinant coronavirus antigens, which were based on the sequence collected in 2019 (the Wuhan variant) and produced in an E. coli bacterial expression system. The present work has shown, for the first time, that these recombinant antigens induce the production of antibodies that clearly interact with produced in CHO full-length S-protein of the Omicron variant. The immunogenicity of these recombinant antigens was studied in formulations with different adjuvants: Freund's adjuvant, Al(OH)3 and an adjuvant based on spherical particles (SPs), which are structurally modified plant virus. All adjuvanted formulations effectively stimulated Omicron-specific IgG production in mice. These universal coronavirus antigens could be considered the main component for the further development of broad-spectrum coronavirus vaccines for the prevention of SARS-CoV-2 infection. The present work also provides evidence that the synthetic biology approach is a promising strategy for the development of highly cross-reactive vaccines. Moreover, it is important to note that the bacterial expression system might be appropriate for the production of antigenically active universal antigens.

Recombinant Protein Vaccines against Human Betacoronaviruses: Strategies, Approaches and Progress.

Betacoronaviruses have already troubled humanity more than once. In 2002-2003 and 2012, the SARS-CoV and MERS-CoV, respectively, caused outbreaks of respiratory syndromes with a fatal outcome. The spread of the SARS-CoV-2 coronavirus has become a pandemic. These three coronaviruses belong to the genus Betacoronavirus and have a zoonotic origin. The emergence of new coronavirus infections in the future cannot be ruled out, and vaccination is the main way to prevent the spread of the infection. Previous experience in the development of vaccines against SARS and MERS has helped to develop a number of vaccines against SARS-CoV-2 in a fairly short time. Among them, there are quite a few recombinant protein vaccines, which seem to be very promising in terms of safety, minimization of side effects, storage and transportation conditions. The problem of developing a universal betacoronavirus vaccine is also still relevant. Here, we summarize the information on the designing of vaccines based on recombinant proteins against highly pathogenic human betacoronaviruses SARS-CoV, MERS-CoV and SARS-CoV-2.

Plant Viruses as Adjuvants for Next-Generation Vaccines and Immunotherapy.

Vaccines are the cornerstone of infectious disease control and prevention. The outbreak of SARS-CoV-2 has confirmed the urgent need for a new approach to the design of novel vaccines. Plant viruses and their derivatives are being used increasingly for the development of new medical and biotechnological applications, and this is reflected in a number of preclinical and clinical studies. Plant viruses have a unique combination of features (biosafety, low reactogenicity, inexpensiveness and ease of production, etc.), which determine their potential. This review presents the latest data on the use of plant viruses with different types of symmetry as vaccine components and adjuvants in cancer immunotherapy. The discussion concludes that the most promising approaches might be those that use structurally modified plant viruses (spherical particles) obtained from the Tobacco mosaic virus. These particles combine high adsorption properties (as a carrier) with strong immunogenicity, as has been confirmed using various antigens in animal models. According to current research, it is evident that plant viruses have great potential for application in the development of vaccines and in cancer immunotherapy.

Vaccine Candidate Against COVID-19 Based on Structurally Modified Plant Virus as an Adjuvant.

A recombinant vaccine candidate has been developed based on the major coronaviruses' antigen (S protein) fragments and a novel adjuvant-spherical particles (SPs) formed during tobacco mosaic virus thermal remodeling. The receptor-binding domain and the highly conserved antigenic fragments of the S2 protein subunit were chosen for the design of recombinant coronavirus antigens. The set of three antigens (Co1, CoF, and PE) was developed and used to create a vaccine candidate composed of antigens and SPs (SPs + 3AG). Recognition of SPs + 3AG compositions by commercially available antibodies against spike proteins of SARS-CoV and SARS-CoV-2 was confirmed. The immunogenicity testing of these compositions in a mouse model showed that SPs improved immune response to the CoF and PE antigens. Total IgG titers against both proteins were 9-16 times higher than those to SPs. Neutralizing activity against SARS-CoV-2 in serum samples collected from hamsters immunized with the SPs + 3AG was demonstrated.

Thermal remodelling of Alternanthera mosaic virus virions and virus-like particles into protein spherical particles.

The present work addresses the thermal remodelling of flexible plant viruses with a helical structure and virus-like particles (VLPs). Here, for the first time, the possibility of filamentous Alternanthera mosaic virus (AltMV) virions' thermal transition into structurally modified spherical particles (SP) has been demonstrated. The work has established differences in formation conditions of SP from virions (SPV) and VLPs (SPVLP) that are in accordance with structural data (on AltMV virions and VLPs). SP originate from AltMV virions through an intermediate stage. However, the same intermediate stage was not detected during AltMV VLPs' structural remodelling. According to the biochemical analysis, AltMV SPV consist of protein and do not include RNA. The structural characterisation of AltMV SPV/VLP by circular dichroism, intrinsic fluorescence spectroscopy and thioflavin T fluorescence assay has been performed. AltMV SPV/VLP adsorption properties and the availability of chemically reactive surface amino acids have been analysed. The revealed characteristics of AltMV SPV/VLP indicate that they could be applied as protein platforms for target molecules presentation and for the design of functionally active complexes.

Investigation of the calcium-induced activation of the bacteriophage T5 peptidoglycan hydrolase promoting host cell lysis.

Peptidoglycan hydrolase of bacteriophage T5 (EndoT5) is a Ca2+-dependent l-alanyl-d-glutamate peptidase, although the mode of Ca2+ binding and its physiological significance remain obscure. Site-directed mutagenesis was used to elucidate the role of the polar amino acids of the mobile loop of EndoT5 (111-130) in Ca2+ binding. The mutant proteins were purified to electrophoretic homogeneity, the overall structures were characterized by circular dichroism, and the calcium dissociation constants were determined via NMR spectroscopy. The data suggest that polar amino acids D113, N115, and S117 of EndoT5 are involved in the coordination of calcium ions by forming the core of the EF-like Ca2+-binding loop while the charged residues D122 and E123 of EndoT5 contribute to maintaining the loop net charge density. The results suggest that Ca2+ binding to the EndoT5 molecule could be essential for the stabilization of the long mobile loop in the catalytically active "open" conformation. The possible mechanism of Ca2+ regulation of EndoT5 activity during bacteriophage T5's life cycle through the Ca2+ concentration difference between the cytoplasm and the periplasm of the host bacteria cell has been discussed. The study reveals valuable insight into the role of calcium in the regulation of phage-induced bacterial lysis.