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Journal of Virology, September 2008, p. 9134-9142, Vol. 82, No. 18
0022-538X/08/$08.00+0     doi:10.1128/JVI.00394-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Mapping of the CXCR4 Binding Site within Variable Region 3 of the Feline Immunodeficiency Virus Surface Glycoprotein{triangledown}

Magnus Sundstrom,1 Rebecca L. White,1 Aymeric de Parseval,2 K. Jagannadha Sastry,3 Garrett Morris,1 Chris K. Grant,4 and John H. Elder1*

The Scripps Research Institute, Department of Molecular Biology, La Jolla, California,1 Public Health Research Institute, Newark, New Jersey,2 The University of Texas M. D. Anderson Cancer Center, Department of Immunology, Houston, Texas,3 Custom Monoclonals International, Inc., West Sacramento, California4

Received 23 February 2008/ Accepted 25 June 2008

Feline immunodeficiency virus (FIV) shares with T-cell tropic strains of human immunodeficiency virus type 1 (HIV-1) the use of the chemokine receptor CXCR4 for cellular entry. In order to map the interaction of the FIV envelope surface unit (SU) with CXCR4, full-length FIV SU-Fc as well as constructs with deletions of extended loop L2, V3, V4, or V5 were produced in stable CHO cell lines. Binding studies were performed using these proteins on 3201 cells (CXCR4hi CD134–), with or without the CXCR4 inhibitor AMD3100. The findings established that SU binding to CXCR4 specifically requires the V3 region of SU. Synthetic peptides spanning the V3 region as well as a panel of monoclonal antibodies (MAbs) to SU were used to further map the site of CXCR4 interaction. Both the SU V3-specific antibodies and the full-length V3 peptide potently blocked binding of SU to CXCR4 and virus entry. By using a set of nested peptides overlapping a region of SU specifically recognized by CD134-dependent neutralizing V3 MAbs, we showed that the neutralizing epitope and the region required for CXCR4 binding are within the same contiguous nine-amino-acid sequence of V3. Site-directed mutagenesis was used to reveal that serine 393 and tryptophan 394 at the predicted tip of V3 are required to facilitate entry into the target cell via CXCR4. Although the amino acid sequences are not identical between FIV and HIV, the ability of FIV to bind and utilize both feline and human CXCR4 makes the feline model an attractive venue for development of broad-based entry antagonists.

* Corresponding author. Mailing address: Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, MB-14, La Jolla, CA 92037. Phone: (858) 784-8270. Fax: (858) 784-2750. E-mail: jelder{at}scripps.edu

{triangledown} Published ahead of print on 2 July 2008.

Journal of Virology, September 2008, p. 9134-9142, Vol. 82, No. 18
0022-538X/08/$08.00+0     doi:10.1128/JVI.00394-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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