Enveloped viruses access their host cells by binding to receptors on the plasma membrane and then undergoing fusion with the host membrane. Both binding and fusion are mediated by a specific viral “spike” protein that is typically primed for fusion activation by proteolytic cleavage to expose the fusion peptide. Coronavirus fusion spike protein (CoV S) is a complex biomolecular machine that has a novel fusion peptide with has a great deal of inherent flexibility in its fusion reaction. This is exploited by these viruses in their diverse entry pathways and is a primary determinant of viral tropism. We have pioneered the concept that that the proteolytic cleavage events in S that lead to membrane fusion occur both at the interface of the receptor binding (S1) and fusion (S2) domains (called S1/S2), as well as adjacent to a structurally and functionally novel fusion peptide within S2 (called S2’). Thus, there are notable differences between CoV S and most other class I fusion proteins including: 1) that the proteolytic events liberating the fusion peptide are diverse, and 2) that the fusion peptide itself is atypical in sequence compared to other fusion peptides, containing a mixture of important hydrophobic and negativelycharged residues, and may represent a larger than normal fusion “platform” instead of a defined “peptide”. Thus fusion peptide activity is likely controlled by reorganization of the fusion platform, based on both hydrophobic (i.e. lipid-binding) and ionic (i.e. Ca2+ and pH) interactions. Despite the recent availability of two S structures, there remains a very limited mechanistic understanding of membrane fusion for the CoV family, or any structural information to correlate structural biology aspects of S to its function in membrane fusion. This information is critical to understanding viral pathogenesis and CoV emergence into the human population. We propose to develop a panel of monoclonal antibodies to the SARS-CoV-2/COVID-19 fusion peptide that will be used as tools to understand the fusion mechanism of coronaviruses, and which will be integrated into our biophysical, biochemical, and in vivo approach to study the unique cleavage-activated regulation of CoV S protein. These antibodies will also provide a platform for development of novel broadly-acting therapeutic antibodies for SARSCoV- 2 and other coronaviruses. Moving the field forward with these innovative studies will provide critical knowledge about CoV entry and tropism needed to safeguard human health from an emerging pathogen likely to cause severe outbreaks, and for which few or no medical countermeasures exist.