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    High-Resolution Epitope Positioning of a Large Collection of Neutralizing and Nonneutralizing Single-Domain Antibodies on the Enzymatic and Binding Subunits of Ricin Toxin

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    Issue Date
    2017-12-05
    Author
    Vance, David J.
    Tremblay, Jacqueline M.
    Rong, Yinghui
    Krishna Angalakurthi, Siva
    Volkin, David B.
    Middaugh, C. Russell
    Weis, David D.
    Shoemaker, Charles B.
    Mantis, Nicholas J.
    Publisher
    American Society for Microbiology
    Type
    Article
    Article Version
    Scholarly/refereed, publisher version
    Rights
    © 2017 American Society for Microbiology. All Rights Reserved.
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    Abstract
    We previously produced a heavy-chain-only antibody (Ab) VH domain (VHH)-displayed phage library from two alpacas that had been immunized with ricin toxoid and nontoxic mixtures of the enzymatic ricin toxin A subunit (RTA) and binding ricin toxin B subunit (RTB) (D. J. Vance, J. M. Tremblay, N. J. Mantis, and C. B. Shoemaker, J Biol Chem 288:36538–36547, 2013, https://doi.org/10.1074/jbc.M113.519207). Initial and subsequent screens of that library by direct enzyme-linked immunosorbent assay (ELISA) yielded more than two dozen unique RTA- and RTB-specific VHHs, including 10 whose structures were subsequently solved in complex with RTA. To generate a more complete antigenic map of ricin toxin and to define the epitopes associated with toxin-neutralizing activity, we subjected the VHH-displayed phage library to additional “pannings” on both receptor-bound ricin and antibody-captured ricin. We now report the full-length DNA sequences, binding affinities, and neutralizing activities of 68 unique VHHs: 31 against RTA, 33 against RTB, and 4 against ricin holotoxin. Epitope positioning was achieved through cross-competition ELISAs performed with a panel of monoclonal antibodies (MAbs) and verified, in some instances, with hydrogen-deuterium exchange mass spectrometry. The 68 VHHs grouped into more than 20 different competition bins. The RTA-specific VHHs with strong toxin-neutralizing activities were confined to bins that overlapped two previously identified neutralizing hot spots, termed clusters I and II. The four RTB-specific VHHs with potent toxin-neutralizing activity grouped within three adjacent bins situated at the RTA-RTB interface near cluster II. These results provide important insights into epitope interrelationships on the surface of ricin and delineate regions of vulnerability that can be exploited for the purpose of vaccine and therapeutic development.
    URI
    http://hdl.handle.net/1808/27394
    DOI
    https://doi.org/10.1128/CVI.00236-17
    Collections
    • Chemistry Scholarly Works [587]
    Citation
    Vance DJ, Tremblay JM, Rong Y, Angalakurthi SK, Volkin DB, Middaugh CR, Weis DD, Shoemaker CB, Mantis NJ. 2017. Highresolution epitope positioning of a large collection of neutralizing and nonneutralizing single-domain antibodies on the enzymatic and binding subunits of ricin toxin. Clin Vaccine Immunol 24:e00236-17. https://doi.org/10.1128/CVI.00236-17.

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    Contact KU ScholarWorks
    785-864-8983
    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    785-864-8983

    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    Image Credits
     

     

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