SARS-CoV-2 Spike protein peptides displayed in the Pyrococcus furiosus RAD system preserve epitopes antigenicity, immunogenicity, and virus-neutralizing activity of antibodies

Abstract

Amongst the potential contribution of protein or peptide-display systems to study epitopes with relevant immunological features, the RAD display system stands out as a highly stable scaffold protein that allows the presentation of constrained target peptides. Here, we employed the RAD display system to present peptides derived from the SARS-CoV-2 Spike (S) protein as a tool to detect specific serum antibodies and to generate polyclonal antibodies capable of inhibiting SARS-CoV-2 infectivity in vitro. 44 linear S-derived peptides were genetically fused with the RAD scaffold (RAD-SCoV-epitopes) and screened for antigenicity with sera collected from COVID-19-infected patients. In a second step, selected RAD-SCoV-epitopes were used to immunize mice and generate antibodies. Phenotypic screening showed that some of these antibodies were able to recognize replicating viral particles in VERO CCL-81 and most notably seven of the RAD-SCoV-epitopes were able to induce antibodies that inhibited viral infection. Our findings highlight the RAD display system as an useful platform for the immunological characterization of peptides and a potentially valuable strategy for the design of antigens for peptide-based vaccines, for epitope-specific antibody mapping, and for the development of antibodies for diagnostic and therapeutic purposes.

Neutralizing antibody response after immunization with a COVID‐19 bivalent vaccine: Insights to the future

“…In addition, instead of using plaque or focus reduction neutralization test (PRNT or FRNT, respectively) to access the antibody neutralization titers, we used CPE‐VNT, a technique that has been successfully applied to measure serum NAbs both in humans and in other mammalian species. Although PRNT, and FRNT are considered golden standard, the CPE‐VNT has shown to be particularly useful for studying larger sets of samples 18,26,27 . The same technique has been used in our previous studies 13,14 and we preferred not to change the methodology in the present study.…”

“…are considered golden standard, the CPE-VNT has shown to be particularly useful for studying larger sets of samples. 18,26,27 The same technique has been used in our previous studies 13,14 and we preferred not to change the methodology in the present study.…”

Successful production of antibodies against extra cytoplasmic loops of the Mycobacterium tuberculosis ABC transporter Rv1819c

“…In previous studies, we showed that this system was suitable for production of antibodies against extracellular loops of permeases from ABC transporters of Escherichia coli and Staphylococcus aureus without the need for expression and purification of complete membrane proteins [6]. The system also was applied to evaluate the immunogenicity of sequences from SARS-CoV-2 Spike protein, production of antibodies and selection of antibodies with ability to neutralize the virus infection in human cells [7].…”