Membrane fusion, mediated by viral spike glycoproteins, is a key process in the infection cycle of all enveloped human and animal viruses. The overall goal of this project is to understand the molecular mechanisms of Ebola and human immunodeficiency virus entry by membrane fusion, by taking combined state-of-the-art structural and cell biophysical imaging approaches. Although the crystal structures of the ectodomains of several enveloped virus spike glycoproteins have been determined to high resolution and although we and others have contributed NMR structures of the membrane- interactive parts of some of these proteins over the years, we are only beginning to understand the structural transformations that take place when these protein domains interact with membranes and each other, and how these structures drive membrane fusion. In the previous grant period we have made substantial progress towards elucidating the structures of the fusion loop, membrane proximal, and transmembrane domains of the Ebola virus envelope glycoprotein and their pH-dependencies in membrane environments. We have also discovered that HIV particles bind and fuse preferentially at lipid discontinuities in plasma membranes of infected cells and that this process depends critically on the level of membrane cholesterol. Building on these achievements, we now propose to (1) solve the structure of the Ebola virus fusion loop in interaction with its membrane-proximal and transmembrane domain, (2) determine the intracellular factors that trigger Ebola virus fusion in the late endosome, and (3) determine the role of membrane heterogeneity in forming the fusion pore for HIV entry.
Viral infections cause many devastating diseases in the US and abroad. Many viruses including influenza, Ebola, and human immunodeficiency viruses, are enveloped by a membrane that encloses their hereditary nucleic acids. To propagate these viruses infect cells by fusing their membrane envelopes with a target membrane of the cell either at the cell surface or in an internal organelle, i.e. the endosome. This project aims at discovering the fundamental mechanisms by which Ebola virus and HIV fuse with their respective target membranes. State--of--the--art structural, biophysical, and high- -resolution imaging approaches will be taken to delineate the cell entry mechanisms of these two viruses. Knowledge gained from this research will help to eventually develop new classes of viral entry inhibitors.
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