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

In this report, we used hydrogen exchange-mass spectrometry (HX-MS) to identify the epitopes recognized by 21 single-domain camelid antibodies (V<sub>H</sub>Hs) directed against the ribosome-inactivating subunit (RTA) of ricin toxin, a biothreat agent of concern to military and public he...

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Main Authors:	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
Format:	Article
Language:	English
Published:	MDPI AG 2018-12-01
Series:	Antibodies
Subjects:	toxin antibody camelid vaccine biodefense hydrogen exchange-mass spectrometry
Online Access:	https://www.mdpi.com/2073-4468/7/4/45

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spelling	doaj-08b529d18940415d831f6223da371ea82020-11-24T23:31:41ZengMDPI AGAntibodies2073-44682018-12-01744510.3390/antib7040045antib7040045A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active SiteSiva Krishna Angalakurthi0David J. Vance1Yinghui Rong2Chi My Thi Nguyen3Michael J. Rudolph4David Volkin5C. Russell Middaugh6David D. Weis7Nicholas J. Mantis8Department of Pharmaceutical Chemistry and Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, KS 660451, USADivision of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, NY 12208, USADivision of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, NY 12208, USANew York Structural Biology Center (NYSBC), New York, NY 10027, USANew York Structural Biology Center (NYSBC), New York, NY 10027, USADepartment of Pharmaceutical Chemistry and Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, KS 660451, USADepartment of Pharmaceutical Chemistry and Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, KS 660451, USADepartment of Chemistry and Ralph Adams Institute for Bioanalytical Chemistry, University of Kansas, Lawrence, KS 660451, USADivision of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, NY 12208, USAIn this report, we used hydrogen exchange-mass spectrometry (HX-MS) to identify the epitopes recognized by 21 single-domain camelid antibodies (V<sub>H</sub>Hs) directed against the ribosome-inactivating subunit (RTA) of ricin toxin, a biothreat agent of concern to military and public health authorities. The V<sub>H</sub>Hs, 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<sub>H</sub>Hs recognized four distinct epitope subclusters (3.1–3.4). Sixteen of the 21 V<sub>H</sub>Hs grouped within subcluster 3.1 and engage RTA α-helices C and G. Three V<sub>H</sub>Hs grouped within subcluster 3.2, encompassing α-helices C and G, plus α-helix B. The single V<sub>H</sub>H in subcluster 3.3 engaged RTA α-helices B and G, while the epitope of the sole V<sub>H</sub>H 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<sub>H</sub>Hs 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.https://www.mdpi.com/2073-4468/7/4/45toxinantibodycamelidvaccinebiodefensehydrogen exchange-mass spectrometry
collection	DOAJ
language	English
format	Article
sources	DOAJ
author	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
spellingShingle	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 A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site Antibodies toxin antibody camelid vaccine biodefense hydrogen exchange-mass spectrometry
author_facet	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
author_sort	Siva Krishna Angalakurthi
title	A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site
title_short	A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site
title_full	A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site
title_fullStr	A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site
title_full_unstemmed	A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site
title_sort	collection of single-domain antibodies that crowd ricin toxin’s active site
publisher	MDPI AG
series	Antibodies
issn	2073-4468
publishDate	2018-12-01
description	In this report, we used hydrogen exchange-mass spectrometry (HX-MS) to identify the epitopes recognized by 21 single-domain camelid antibodies (V<sub>H</sub>Hs) directed against the ribosome-inactivating subunit (RTA) of ricin toxin, a biothreat agent of concern to military and public health authorities. The V<sub>H</sub>Hs, 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<sub>H</sub>Hs recognized four distinct epitope subclusters (3.1–3.4). Sixteen of the 21 V<sub>H</sub>Hs grouped within subcluster 3.1 and engage RTA α-helices C and G. Three V<sub>H</sub>Hs grouped within subcluster 3.2, encompassing α-helices C and G, plus α-helix B. The single V<sub>H</sub>H in subcluster 3.3 engaged RTA α-helices B and G, while the epitope of the sole V<sub>H</sub>H 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<sub>H</sub>Hs 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.
topic	toxin antibody camelid vaccine biodefense hydrogen exchange-mass spectrometry
url	https://www.mdpi.com/2073-4468/7/4/45
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A Collection of Single-Domain Antibodies that Crowd Ricin Toxin’s Active Site

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