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SARS\u2010CoV entry is facilitated by the spike protein (S), which consists of an N\u2010terminal domain (S1) responsible for cellular attachment and a C\u2010terminal domain (S2) that mediates viral and host cell membrane fusion. The SARS\u2010CoV S2 is a potential drug target, as peptidomimetics against S2 act as potent fusion inhibitors. In this study, site\u2010directed mutagenesis and thermal stability experiments on electrostatic, hydrophobic, and polar residues to dissect their roles in stabilizing the S2 postfusion conformation was performed. It was shown that unlike the pH\u2010independent retroviral fusion proteins, SARS\u2010CoV S2 is stable over a wide pH range, supporting its ability to fuse at both the plasma membrane and endosome. A comprehensive SARS\u2010CoV S2 analysis showed that specific hydrophobic positions at the C\u2010terminal end of the HR2, rather than electrostatics are critical for fusion protein stabilization. Disruption of the conserved C\u2010terminal hydrophobic residues destabilized the fusion core and reduced the melting temperature by 30\u00b0C. The importance of the C\u2010terminal hydrophobic residues led us to identify a 42\u2010residue substructure on the central core that is structurally conserved in all existing CoV S2 fusion proteins (root mean squared deviation\u2009=\u20090.4 \u00c5). This is the first study to identify such a conserved substructure and likely represents a common foundation to facilitate viral fusion. We have discussed the role of key residues in the design of fusion inhibitors and the potential of the substructure as a general target for the development of novel therapeutics against CoV infections.<\/jats:p>","DOI":"10.1002\/pro.2442","type":"journal-article","created":{"date-parts":[[2014,2,12]],"date-time":"2014-02-12T12:44:42Z","timestamp":1392209082000},"page":"603-617","source":"Crossref","is-referenced-by-count":33,"title":["Influence of hydrophobic and electrostatic residues on SARS\u2010coronavirus S2 protein stability: Insights into mechanisms of general viral fusion and inhibitor design"],"prefix":"10.1002","volume":"23","author":[{"given":"Halil","family":"Aydin","sequence":"first","affiliation":[{"name":"Department of Laboratory Medicine and Pathobiology Faculty of Medicine, University of Toronto Toronto Ontario M5S 1A8 Canada"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Dina","family":"Al\u2010Khooly","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathobiology Faculty of Medicine, University of Toronto Toronto Ontario M5S 1A8 Canada"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jeffrey E.","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathobiology Faculty of Medicine, University of Toronto Toronto Ontario M5S 1A8 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