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Journal of Virology, January 2006, p. 941-950, Vol. 80, No. 2
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.2.941-950.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Monoclonal Antibodies Targeting the HR2 Domain and the Region Immediately Upstream of the HR2 of the S Protein Neutralize In Vitro Infection of Severe Acute Respiratory Syndrome Coronavirus

Kuo-Ming Lip,1,{dagger} Shuo Shen,1*,{dagger} Xiaoming Yang,2 Choong-Tat Keng,1 Aihua Zhang,2 Hsueh-Ling Janice Oh,1 Zhi-Hong Li,1 Le-Ann Hwang,1 Chih-Fong Chou,1 Burtram C. Fielding,1 Timothy H. P. Tan,1 Josef Mayrhofer,3 Falko G. Falkner,3 Jianlin Fu,1 Seng Gee Lim,1 Wanjin Hong,1 and Yee-Joo Tan1

Institute of Molecular and Cell Biology, Singapore 138673,1 Wuhan Institute of Biological Products, Wuhan, P. R. China 430060,2 Baxter Vaccines, Orth/Donau, Austria3

Received 7 June 2005/ Accepted 19 October 2005

We have previously shown that an Escherichia coli-expressed, denatured spike (S) protein fragment of the severe acute respiratory coronavirus, containing residues 1029 to 1192 which include the heptad repeat 2 (HR2) domain, was able to induce neutralizing polyclonal antibodies (C. T. Keng, A. Zhang, S. Shen, K. M. Lip, B. C. Fielding, T. H. Tan, C. F. Chou, C. B. Loh, S. Wang, J. Fu, X. Yang, S. G. Lim, W. Hong, and Y. J. Tan, J. Virol. 79:3289-3296, 2005). In this study, monoclonal antibodies (MAbs) were raised against this fragment to identify the linear neutralizing epitopes in the functional domain and to investigate the mechanisms involved in neutralization. Eighteen hybridomas secreting the S protein-specific MAbs were obtained. Binding sites of these MAbs were mapped to four linear epitopes. Two of them were located within the HR2 region and two immediately upstream of the HR2 domain. MAbs targeting these epitopes showed in vitro neutralizing activities and were able to inhibit cell-cell membrane fusion. These results provide evidence of novel neutralizing epitopes that are located in the HR2 domain and the spacer region immediately upstream of the HR2 of the S protein.

* Corresponding author. Mailing address: Institute of Molecular and Cell Biology, Proteos, 61 Biopolis Drive, Singapore 138673. Phone: 65 6586 9622. Fax: 65 67791117. E-mail: shenshuo{at}imcb.a-star.edu.sg.

{dagger} K.-M.L. and S.S. contributed equally to this article.

Journal of Virology, January 2006, p. 941-950, Vol. 80, No. 2
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.2.941-950.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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