The severe acute respiratory syndrome-associated coronavirus spike protein: Identification and characterization of the fusion-inducing domains and design of fusion inhibitory peptides
Description
The severe acute respiratory syndrome (SARS) epidemic of 2002--2003 resulted in 774 deaths in 29 countries worldwide. The etiological agent of SARS (SARS-CoV) was quickly identified as belonging to the family Coronaviridae. Like other CoV, SARS-CoV enters cell through fusion of the viral membrane with the target cell membrane. This process is facilitated by the viral spike (S) glycoprotein. It is believed that the S1 subunit of the SARS-CoV S protein mediates binding to the cellular receptor(s) ACE2 and/or DC209L, while the S2 subunit, a predicted class I viral fusion protein, drives the fusion reaction The studies presented herein further characterize the SARS-CoV S protein by identifying and characterizing the putative fusion peptide and determining the functional role of the conserved aromatic domain. The hydrophobic stretch of 19 amino acids, corresponding to residues 770--788, was shown to be the putative fusion peptide of the SARS-CoV S2 subunit. A peptide analogous to this region strongly partitioned into the membranes of lipid vesicles, adopting a beta-sheet structure, and was capable of inducing fusion and membrane permeabilization In addition, biophysical studies of the aromatic domains of the SARS-CoV, mouse hepatitis virus (MHV) and the human CoV OC43 S2 subunits showed that these regions also strongly partitioned into lipid membranes and induced lipid vesicle permeabilization. Based on these biophysical studies, we propose that during membrane apposition these domains function to destabilize the leaflets of the apposed viral and target cell membranes by forming a continuous hydrophobic surface track resulting in membrane fusion and subsequent viral nucleocapsid entry Lastly, peptides analogous to regions of the S2 subunit were tested as inhibitors of CoV infectivity. Peptides analogous to the N-terminus or to the pretransmembrane domain of the S2 subunit inhibited CoV plaque formation by 40--50%. In addition, peptides analogous to the SARS-CoV or MHV loop region inhibited viral plaque formation by >80%. This effect was dose-dependent and not attributable to the propensity of the peptides to adopt a defined secondary structure. The antiviral activity of the CoV peptides tested provides an attractive basis for the development of new peptide inhibitors for fusion proteins of other viruses