Format

Send to

Choose Destination
J Biol Chem. 2017 Jan 20;292(3):872-883. doi: 10.1074/jbc.M116.758102. Epub 2016 Nov 30.

Structural Analysis of Single Domain Antibodies Bound to a Second Neutralizing Hot Spot on Ricin Toxin's Enzymatic Subunit.

Author information

1
From the New York Structural Biology Center, New York, New York 10027, mrudolph@nysbc.org.
2
the Division of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, New York 12208, and.
3
From the New York Structural Biology Center, New York, New York 10027.
4
the Division of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, New York 12208, and nicholas.mantis@health.ny.gov.
5
the Department of Biomedical Sciences, University at Albany, Albany, New York 12201.

Abstract

Ricin toxin is a heterodimer consisting of RTA, a ribosome-inactivating protein, and RTB, a lectin that facilitates receptor-mediated uptake into mammalian cells. In previous studies, we demonstrated that toxin-neutralizing antibodies target four spatially distinct hot spots on RTA, which we refer to as epitope clusters I-IV. In this report, we identified and characterized three single domain camelid antibodies (VHH) against cluster II. One of these VHHs, V5E1, ranks as one of the most potent ricin-neutralizing antibodies described to date. We solved the X-ray crystal structures of each of the three VHHs (E1, V1C7, and V5E1) in complex with RTA. V5E1 buries a total of 1,133 Å2 of surface area on RTA and makes primary contacts with α-helix A (residues 18-32), α-helix F (182-194), as well as the F-G loop. V5E1, by virtue of complementarity determining region 3 (CDR3), may also engage with RTB and potentially interfere with the high affinity galactose-recognition element that plays a critical role in toxin attachment to cell surfaces and intracellular trafficking. The two other VHHs, E1 and V1C7, bind epitopes adjacent to V5E1 but display only weak toxin neutralizing activity, thereby providing structural insights into specific residues within cluster II that may be critical contact points for toxin inactivation.

KEYWORDS:

antibody; antigen; crystal structure; epitope mapping; neutralizing; ricin; toxin

PMID:
27903650
PMCID:
PMC5247660
DOI:
10.1074/jbc.M116.758102
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for HighWire Icon for PubMed Central
Loading ...
Support Center