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Journal of Virology, January 2003, p. 830-840, Vol. 77, No. 2
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.2.830-840.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Conformational Changes in the Spike Glycoprotein of Murine Coronavirus Are Induced at 37°C either by Soluble Murine CEACAM1 Receptors or by pH 8

Bruce D. Zelus,1 Jeanne H. Schickli,1 Dianna M. Blau,1 Susan R. Weiss,2 and Kathryn V. Holmes1*

Department of Microbiology, University of Colorado Health Sciences Center, Denver, Colorado 80262,1 Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 191042

Received 25 June 2002/ Accepted 15 October 2002

The spike glycoprotein (S) of the murine coronavirus mouse hepatitis virus (MHV) binds to viral murine CEACAM receptor glycoproteins and causes membrane fusion. On virions, the 180-kDa S glycoprotein of the MHV-A59 strain can be cleaved by trypsin to form the 90-kDa N-terminal receptor-binding subunit (S1) and the 90-kDa membrane-anchored fusion subunit (S2). Incubation of virions with purified, soluble CEACAM1a receptor proteins at 37°C and pH 6.5 neutralizes virus infectivity (B. D. Zelus, D. R. Wessner, R. K. Williams, M. N. Pensiero, F. T. Phibbs, M. deSouza, G. S. Dveksler, and K. V. Holmes, J. Virol. 72:7237-7244, 1998). We used liposome flotation and protease sensitivity assays to investigate the mechanism of receptor-induced, temperature-dependent virus neutralization. After incubation with soluble receptor at 37°C and pH 6.5, virions became hydrophobic and bound to liposomes. Receptor binding induced a profound, apparently irreversible conformational change in S on the viral envelope that allowed S2, but not S1, to be degraded by trypsin at 4°C. Various murine CEACAM proteins triggered conformational changes in S on recombinant MHV strains expressing S glycoproteins of MHV-A59 or MHV-4 (MHV-JHM) with the same specificities as seen for virus neutralization and virus-receptor activities. Increased hydrophobicity of virions and conformational change in S2 of MHV-A59 could also be induced by incubating virions at pH 8 and 37°C, without soluble receptor. Surprisingly, the S protein of recombinant MHV-A59 virions with a mutation, H716D, that precluded cleavage between S1 and S2 could also be triggered to undergo a conformational change at 37°C by soluble receptor at neutral pH or by pH 8 alone. A novel 120-kDa subunit was formed following incubation of the receptor-triggered SA59H716D virions with trypsin at 4°C. The data show that unlike class 1 fusion glycoproteins of other enveloped viruses, the murine coronavirus S protein can be triggered to a membrane-binding conformation at 37°C either by soluble receptor at neutral pH or by alkaline pH alone, without requiring previous activation by cleavage between S1 and S2.

* Corresponding author. Mailing address: Department of Microbiology, Campus Box B-175, University of Colorado Health Sciences Center, 4200 East 9th Ave., Denver, CO 80262. Phone: (303) 315-7329. Fax: (303) 315-6785. E-mail: kathryn.holmes{at}uchsc.edu.

Journal of Virology, January 2003, p. 830-840, Vol. 77, No. 2
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.2.830-840.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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