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Abstract
The Coronavirus Disease 2019 (COVID-19) global pandemic caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) virus emphasized the vital role of SARS-CoV-2 Spike (S) protein binding its angiotensin converting enzyme 2 (ACE2) host cell receptor. Understanding the molecular features that influence the S-ACE2 interaction is essential for the development of effective anti-S therapeutics. While the receptor binding domain (RBD) of ACE2 is known to directly interact with ACE2, the precise influence of N-glycans on S and ACE2, in addition to specific RBD mutations. The S protein is a large viral surface protein that contains the Receptor Binding Domain (RBD) that directly binds to the ACE2 receptor, which allows conformational changes for host protease cleavage and viral infection. Thus, the strength of S-ACE2 binding affinity is a major factor of viral infection and transmission.This thesis aims to elucidate how variations in glycosylation and predominant RBD mutations of the SARS-CoV-2 S glycoprotein influence interactions with receptor and antibody binding. Through a combination of structural analyses and in vitro binding assays via biolayer interferometry, this research provides insight into the binding affinities Spike-ACE2 complexes in quantitative terms. Key results revealed that specific N-glycans on ACE2 and S mediated binding. Additionally, specific RBD mutations lead to altered binding affinities observed in SARS-CoV-2 variants of concern (VOC) for ACE2 and monoclonal antibodies (mAbs). These insights will provide understanding of the factors that mediate therapeutic mAb binding to their target antigen, and the design parameters that should be considered as demonstrated in anti-S therapeutic mAb interactions.