Abstract The sudden emergence of novel viruses such as SARS-CoV may cause severe disease and economic losses that call for rapid development of novel anti-virals. To develop new approaches for the treatment of SARS, I will study the structure and function of the unique long interhelical (IH) domain of the spike glycoprotein (S) of SARS-CoV. Receptor-induced conformational changes in S on virions lead to fusion of the viral envelope with host cell membranes and virus entry. I will identify and optimize peptides from the IH domain that block these conformational changes in S and prevent infection. Proposed research and summarized methodology. In Aim 1 I will test the functional conservation between the IH domains of the S proteins of SARS-CoV and murine coronavirus MHV by interchanging their IH domains. The gene encoding the chimeric MHV S protein with the SARS-CoV IH domain will be introduced into the MHV genome by targeted RNA recombination, and the resulting viruses tested for infectivity. Retrovirus pseudotypes expressing the chimeric SARS-CoV S with the MHV IH domain will be assayed for infection of receptor-expressing cells. Chimeric proteins with smaller swaps in the IH domains, and truncations and site-directed mutations will be engineered in the IH domains to identify regions of functional importance in virus entry that can serve as targets for drug development. In Aim 2 I will characterize receptor-induced conformational changes in the IH domains of SARS-CoV and MHV spike proteins using structural and biochemical approaches (e.g. circular dichroism, analytical ultracentrifugation and X-ray crystallography). Soluble ectodomains of the wild type, chimeric and mutant S proteins will be purified, and their structural and biochemical properties will be compared. This will provide further insight into the important roles of the IH domain in the receptor-induced conformation changes leading to virus entry. The resulting structural and biochemical knowledge will aid in the development of novel anti-virals that target the IH domain. In Aim 3 I will identify and optimize peptides from the IH domain of SARS-CoV and MHV S that block infection. I will determine the location in S where the inhibitory peptides bind using chemical cross-linking followed by mass spectrometry. These data, together with the structural information from Aim 2 will enable me to modify the peptides in order to optimize inhibition of virus entry. The optimized peptides will be lead compounds for development of anti-virals to prevent entry of SARS-CoV and other coronaviruses. This strategy for rapid development of anti-viral peptides can be applied to any emerging virus that is found by sequencing of the genome to have a type 1 viral fusion glycoprotein.