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Two Synthetic Antibodies that Recognize and Neutralize Distinct Proteolytic Forms of the Ebola Virus Envelope Glycoprotein

Authors

  • Jayne F. Koellhoffer,

    1. Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (USA)
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    • These authors contributed equally to this work.

  • Dr. Gang Chen,

    1. Banting and Best Department of Medical Research, Terrence Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, 160 College Street, Toronto, Ontario M5S 3E1 (Canada)
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    • These authors contributed equally to this work.

  • Rohini G. Sandesara,

    1. Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (USA)
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    • These authors contributed equally to this work.

  • Dr. Shridhar Bale,

    1. Department of Immunology and Microbial Science and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037 (USA)
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  • Prof. Dr. Erica Ollmann Saphire,

    1. Department of Immunology and Microbial Science and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037 (USA)
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  • Prof. Dr. Kartik Chandran,

    Corresponding author
    1. Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (USA)
    • Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (USA)
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  • Prof. Dr. Sachdev S. Sidhu,

    Corresponding author
    1. Banting and Best Department of Medical Research, Terrence Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, 160 College Street, Toronto, Ontario M5S 3E1 (Canada)
    • Banting and Best Department of Medical Research, Terrence Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, 160 College Street, Toronto, Ontario M5S 3E1 (Canada)
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  • Prof. Dr. Jonathan R. Lai

    Corresponding author
    1. Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (USA)
    • Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461 (USA)
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Abstract

Ebola virus (EBOV) is a highly pathogenic member of the Filoviridae family of viruses that causes severe hemorrhagic fever. Infection proceeds through fusion of the host cell and viral membranes, a process that is mediated by the viral envelope glycoprotein (GP). Following endosomal uptake, a key step in viral entry is the proteolytic cleavage of GP by host endosomal cysteine proteases. Cleavage exposes a binding site for the host cell receptor Niemann-Pick C1 (NPC1) and may induce conformational changes in GP leading to membrane fusion. However, the precise details of the structural changes in GP associated with proteolysis and the role of these changes in viral entry have not been established. Here, we have employed synthetic antibody technology to identify antibodies targeting EBOV GP prior to and following proteolysis (i.e. in the “uncleaved” [GPUNCL] and “cleaved” [GPCL] forms). We identified antibodies with distinct recognition profiles: FabCL bound preferentially to GPCL (EC50=1.7 nM), whereas FabUNCL bound specifically to GPUNCL (EC50=75 nM). Neutralization assays with GP-containing pseudotyped viruses indicated that these antibodies inhibited GPCL- or GPUNCL-mediated viral entry with specificity matching their recognition profiles (IC50: 87 nM for IgGCL; 1 μM for FabUNCL). Competition ELISAs indicate that FabCL binds an epitope distinct from that of KZ52, a well-characterized EBOV GP antibody, and from that of the luminal domain of NPC1. The binding epitope of FabUNCL was also distinct from that of KZ52, suggesting that FabUNCL binds a novel neutralization epitope on GPUNCL. Furthermore, the neutralizing ability of FabCL suggests that there are targets on GPCL available for neutralization. This work showcases the applicability of synthetic antibody technology to the study of viral membrane fusion, and provides new tools for dissecting intermediates of EBOV entry.

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