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Journal of Virology, May 2004, p. 4552-4560, Vol. 78, No. 9
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.9.4552-4560.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Amino Acids 270 to 510 of the Severe Acute Respiratory Syndrome Coronavirus Spike Protein Are Required for Interaction with Receptor

Gregory J. Babcock,* Diana J. Esshaki, William D. Thomas Jr., and Donna M. Ambrosino

Massachusetts Biologic Laboratories, University of Massachusetts Medical School, Jamaica Plain, Massachusetts 02130

Received 13 October 2003/ Accepted 2 January 2004

A novel coronavirus, severe acute respiratory syndrome coronavirus (SARS-CoV), has recently been identified as the causative agent of severe acute respiratory syndrome (SARS). SARS-CoV appears similar to other coronaviruses in both virion structure and genome organization. It is known for other coronaviruses that the spike (S) glycoprotein is required for both viral attachment to permissive cells and for fusion of the viral envelope with the host cell membrane. Here we describe the construction and expression of a soluble codon-optimized SARS-CoV S glycoprotein comprising the first 1,190 amino acids of the native S glycoprotein (S1190). The codon-optimized and native S glycoproteins exhibit similar molecular weight as determined by Western blot analysis, indicating that synthetic S glycoprotein is modified correctly in a mammalian expression system. S1190 binds to the surface of Vero E6 cells, a cell permissive to infection, as demonstrated by fluorescence-activated cell sorter analysis, suggesting that S1190 maintains the biologic activity present in native S glycoprotein. This interaction is blocked with serum obtained from recovering SARS patients, indicating that the binding is specific. In an effort to map the ligand-binding domain of the SARS-CoV S glycoprotein, carboxy- and amino-terminal truncations of the S1190 glycoprotein were constructed. Amino acids 270 to 510 were the minimal receptor-binding region of the SARS-CoV S glycoprotein as determined by flow cytometry. We speculate that amino acids 1 to 510 of the SARS-CoV S glycoprotein represent a unique domain containing the receptor-binding site (amino acids 270 to 510), analogous to the S1 subunit of other coronavirus S glycoproteins.

* Corresponding author. Mailing address: Massachusetts Biologic Laboratories, University of Massachusetts Medical School, 305 South St., Jamaica Plain, MA 02130. Phone: (617) 983-6415. Fax: (617) 983-6477. E-mail: greg.babcock{at}umassmed.edu.

Journal of Virology, May 2004, p. 4552-4560, Vol. 78, No. 9
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.9.4552-4560.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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