Poor, Taylor A.Jones, Lisa M.Sood, AmikaLeser, George P.Plasencia, Manolo D.Rempel, Don L.Jardetzky, Theodore S.Woods, Robert J.Gross, Michael L.Lamb, Robert A.2016-02-292016-02-292014-06-24Poor, T. A., Jones, L. M., Sood, A., Leser, G. P., Plasencia, M. D., Rempel, D. L., … Lamb, R. A. (2014). Probing the paramyxovirus fusion (F) protein-refolding event from pre- to postfusion by oxidative footprinting. Proceedings of the National Academy of Sciences of the United States of America, 111(25), E2596–E2605. http://doi.org/10.1073/pnas.1408983111https://hdl.handle.net/1805/8553To infect a cell, the Paramyxoviridae family of enveloped viruses relies on the coordinated action of a receptor-binding protein (variably HN, H, or G) and a more conserved metastable fusion protein (F) to effect membrane fusion and allow genomic transfer. Upon receptor binding, HN (H or G) triggers F to undergo an extensive refolding event to form a stable postfusion state. Little is known about the intermediate states of the F refolding process. Here, a soluble form of parainfluenza virus 5 F was triggered to refold using temperature and was footprinted along the refolding pathway using fast photochemical oxidation of proteins (FPOP). Localization of the oxidative label to solvent-exposed side chains was determined by high-resolution MS/MS. Globally, metastable prefusion F is oxidized more extensively than postfusion F, indicating that the prefusion state is more exposed to solvent and is more flexible. Among the first peptides to be oxidatively labeled after temperature-induced triggering is the hydrophobic fusion peptide. A comparison of peptide oxidation levels with the values of solvent-accessible surface area calculated from molecular dynamics simulations of available structural data reveals regions of the F protein that lie at the heart of its prefusion metastability. The strong correlation between the regions of F that experience greater-than-expected oxidative labeling and epitopes for neutralizing antibodies suggests that FPOP has a role in guiding the development of targeted therapeutics. Analysis of the residue levels of labeled F intermediates provides detailed insights into the mechanics of this critical refolding event.en-USPublisher PolicyMass spectroscopyProtein refoldingViral fusion proteinArticle