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Journal of Virology, July 2002, p. 6780-6790, Vol. 76, No. 13
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.13.6780-6790.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Dissection of Human Immunodeficiency Virus Type 1 Entry with Neutralizing Antibodies to gp41 Fusion Intermediates

Hana Golding,1* Marina Zaitseva,1 Eve de Rosny,1,{dagger} Lisa R. King,1 Jody Manischewitz,1 Igor Sidorov,2 Miroslaw K. Gorny,3 Susan Zolla-Pazner,3,4 Dimiter S. Dimitrov,2 and Carol D. Weiss1

Division of Viral Products, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland 20892,1 Center for Cancer Research, NCI-Frederick, National Institutes of Health, Frederick, Maryland 21702,2 Department of Pathology, New York University School of Medicine, New York, New York 10016,3 Research Center for AIDS and HIV Infection, Veterans Affairs Medical Center, New York, New York 100104

Received 5 February 2002/ Accepted 4 April 2002

Human immunodeficiency virus type 1 (HIV-1) entry requires conformational changes in the transmembrane subunit (gp41) of the envelope glycoprotein (Env) involving transient fusion intermediates that contain exposed coiled-coil (prehairpin) and six-helix bundle structures. We investigated the HIV-1 entry mechanism and the potential of antibodies targeting fusion intermediates to block Env-mediated membrane fusion. Suboptimal temperature (31.5°C) was used to prolong fusion intermediates as monitored by confocal microscopy. After transfer to 37°C, these fusion intermediates progressed to syncytium formation with enhanced kinetics compared with effector-target (E/T) cell mixtures that were incubated only at 37°C. gp41 peptides DP-178, DP-107, and IQN17 blocked fusion more efficiently (5- to 10-fold-lower 50% inhibitory dose values) when added to E/T cells at the suboptimal temperature prior to transfer to 37°C. Rabbit antibodies against peptides modeling the N-heptad repeat or the six-helix bundle of gp41 blocked fusion and viral infection at 37°C only if preincubated with E/T cells at the suboptimal temperature. Similar fusion inhibition was observed with human six-helix bundle-specific monoclonal antibodies. Our data demonstrate that antibodies targeting gp41 fusion intermediates are able to bind to gp41 and arrest fusion. They also indicate that six-helix bundles can form prior to fusion and that the lag time before fusion occurs may include the time needed to accumulate preformed six-helix bundles at the fusion site.

* Corresponding author. Mailing address: Division of Viral Products, Center for Biologics Evaluation and Research, Food and Drug Administration, Bldg. 29A, Rm. 1A21, HFM-454, 8800 Rockville Pike, Bethesda, MD 20892. Phone: (301) 827-0784. Fax: (301) 496-1810. E-mail: goldingh{at}cber.fda.gov.

{dagger} Present address: Institut de Biologie Structurale, F-38027 Grenoble Cedex 1, France.

Journal of Virology, July 2002, p. 6780-6790, Vol. 76, No. 13
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.13.6780-6790.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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