Mutational analysis of fusion peptide-like regions in the mouse hepatitis virus strain A59 spike protein.
The coronavirus peplomer protein S is responsible for attachment and fusion during viral entry as well as for the induction of cell to cell fusion. While several regions within S have been shown to influence the ability to induce fusion, the region of the protein actually responsible for fusion, the fusion peptide, has not yet been identified. We identified two hydrophobic peptides (peptides #1 and #2) within MHV-A59 S2 as possible fusion domains. This was based on hydrophobicity, conservation among coronavirus S proteins and the prediction of a sided helix conformation. Using site directed mutagenesis and an in vitro cell to cell fusion assay we showed that substitution of hydrophobic amino acids with charged amino acids, within the predicted hydrophobic face of either of these two peptides eliminated fusion. Within peptide #1 substitution of the same hydrophobic amino acids with other hydrophobic amino acids or substitution of polar amino acids with charged or polar amino acids had little effect on fusion. Thus peptides #1 and #2 remain likely candidates for the MHV fusion peptide. A third previously identified peptide within S2 (Chambers et al., 1990) is unlikely as a fusion peptide as it is not well conserved among coronaviruses and substitution within the hydrophobic face with charged amino acids does not effect fusion.
Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
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D000595:Amino Acid Sequence; D000818:Animals; D008561:Membrane Fusion; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D006517:Murine hepatitis virus / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000235:genetics / Q000378:metabolism*; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*
D008562:Membrane Glycoproteins; D010455:Peptides; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578553:spike protein, mouse hepatitis virus