A Rapid System to Study Virus-Antibody Interactions Using Native Mass Spectrometry

Topic Area Addressed: Emerging Viral Infections. Area of Encouragement: Rapid prediction of protective antigens/epitopes and testable correlates of protection on emerging or novel pathogens. Scientific Question: This proposal addresses two related yet independent scientific questions: (1) What is the observed glycosylation pattern on the SARS-CoV-2 Spike protein (vaccine target)? (2) Do different antibodies bind to SARS-CoV-2 with different affinities? Hypothesis: Our hypothesis is that native mass spectrometry can identify both glycosylation patterns in the Spike protein and antibody binding affinity to the spike protein. The energy employed during collision-induced dissociation will be correlated with antibody epitope binding. Rationale: SARS-CoV-2 presents an immediate problem for military mobilization - we currently lack a vaccine to immunize personnel, and we lack a test that functions as an indicator of protection status. This proposal will use native mass spectrometry to inform vaccine design (by characterizing Spike protein glycosylation patterns) and will provide an indication of antibody quality (by measuring the binding affinity of antibodies directly bound to the virus). Emerging viral infections require rapid surface epitope identification and characterization of neutralizing antibody responses. Native mass spectrometry has emerged as a tool to study macromolecular assemblies without requirements for crystallization or microscopy. As a tool, native mass spectrometry has already demonstrated the capability to identify viral particles and dissociate individual viral protein subunits. Work described in this application will identify protein epitopes available on the surface of SARS-CoV-2 for antibody binding. In addition, we will complex monoclonal antibodies directed against the SARS-CoV-2 Spike protein with virus particles and perform native mass spectrometry of the complex. Importantly, we will dissociate the antibody from virus using collision-induced dissociation (CID). The CID energy required to dissociate antibody directed against SARS-CoV-2 provides a framework for studying the relative strength of antibody binding to virus and a harbinger of antibody quality. Short and Long-Term Impact: Over the short term, we envision that defining glycosylation patterns (Aim 1) will impact the work of investigators that aim to develop a vaccine (including DoD research labs working on vaccine candidates) that accurately reflects the antigenic composition of circulating coronaviruses. Over the long term, our studies of antibody-virus interactions (Aim 2) will inform efforts to develop antibody tests that indicate antibody affinity as a proxy for antibody quality. Less