A Collection of Single-Domain Antibodies that Crowd Ricin Toxin's Active Site

Siva Krishna Angalakurthi; David J Vance; Yinghui Rong; Chi My Thi Nguyen; Michael J Rudolph; David Volkin; C Russell Middaugh; David D Weis; Nicholas J Mantis

doi:10.3390/antib7040045

A Collection of Single-Domain Antibodies that Crowd Ricin Toxin's Active Site

Antibodies (Basel). 2018 Dec;7(4):45. doi: 10.3390/antib7040045. Epub 2018 Dec 17.

Authors

Siva Krishna Angalakurthi¹, David J Vance², Yinghui Rong², Chi My Thi Nguyen³, Michael J Rudolph³, David Volkin¹, C Russell Middaugh¹, David D Weis⁴, Nicholas J Mantis²

Affiliations

¹ Department of Pharmaceutical Chemistry and Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, KS 660451, USA; s214a774@ku.edu (S.K.A.); volkin@ku.edu (D.V.); middaugh@ku.edu (C.R.M.).
² Division of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA; david.vance@health.ny.gov (D.J.V.); yinghui.rong@health.ny.gov (Y.R.).
³ New York Structural Biology Center (NYSBC), New York, NY 10027, USA; ntchimy@gmail.com (C.M.T.N.); mrudolph@nysbc.org (M.J.R.).
⁴ Department of Chemistry and Ralph Adams Institute for Bioanalytical Chemistry, University of Kansas, Lawrence, KS 660451, USA; dweis@ku.edu.

Abstract

In this report, we used hydrogen exchange-mass spectrometry (HX-MS) to identify the epitopes recognized by 21 single-domain camelid antibodies (V_HHs) directed against the ribosome-inactivating subunit (RTA) of ricin toxin, a biothreat agent of concern to military and public health authorities. The V_HHs, which derive from 11 different B-cell lineages, were binned together based on competition ELISAs with IB2, a monoclonal antibody that defines a toxin-neutralizing hotspot ("cluster 3") located in close proximity to RTA's active site. HX-MS analysis revealed that the 21 V_HHs recognized four distinct epitope subclusters (3.1-3.4). Sixteen of the 21 V_HHs grouped within subcluster 3.1 and engage RTA α-helices C and G. Three V_HHs grouped within subcluster 3.2, encompassing a-helices C and G, plus α-helix B. The single V_HH in subcluster 3.3 engaged RTA α-helices B and G, while the epitope of the sole V_HH defining subcluster 3.4 encompassed α-helices C and E, and β-strand h. Modeling these epitopes on the surface of RTA predicts that the 20 V_HHs within subclusters 3.1-3.3 physically occlude RTA's active site cleft, while the single antibody in subcluster 3.4 associates on the active site's upper rim.

Keywords: antibody; biodefense; camelid; hydrogen exchange-mass spectrometry; toxin; vaccine.

Grants and funding

HHSN272201400021C/AI/NIAID NIH HHS/United States