@article{57c0d47da1e04fa4adfc86ae5b1c1062,
title = "Camelid single domain antibodies (VHHs) as neuronal cell intrabody binding agents and inhibitors of Clostridium botulinum neurotoxin (BoNT) proteases",
abstract = "Botulinum neurotoxins (BoNTs) function by delivering a protease to neuronal cells that cleave SNARE proteins and inactivate neurotransmitter exocytosis. Small (14 kDa) binding domains specific for the protease of BoNT serotypes A or B were selected from libraries of heavy chain only antibody domains (VHHs or nanobodies) cloned from immunized alpacas. Several VHHs bind the BoNT proteases with high affinity (KD near 1 nM) and include potent inhibitors of BoNT/A protease activity (Ki near 1 nM). The VHHs retain their binding specificity and inhibitory functions when expressed within mammalian neuronal cells as intrabodies. A VHH inhibitor of BoNT/A protease was able to protect neuronal cell SNAP25 protein from cleavage following intoxication with BoNT/A holotoxin. These results demonstrate that VHH domains have potential as components of therapeutic agents for reversal of botulism intoxication.",
keywords = "BoNT, Botulinum, Intrabody, Metalloproteinase, Nanobody, Neurotoxin, VHH",
author = "Tremblay, {Jacqueline M.} and Kuo, {Chueh Ling} and Claudia Abeijon and Jorge Sepulveda and George Oyler and Xuebo Hu and Jin, {Moonsoo M.} and Shoemaker, {Charles B.}",
note = "Funding Information: We are grateful to Dr. David Wilson for preparing some of the A-Lc and to Dr. Randall Kincaid for providing the GST fusion proteins to A-Lc and B-Lc. We greatly appreciate the assistance of Tony Pernthaner and Sally Cole for immunizing the alpacas, titering alpaca sera and preparing alpaca cDNA, and Dr. David Maass for preparing the A-Lc VHH-display library. We thank Nick Salzameda for testing the VHHs for inhibition of B-Lc activity. Finally we thank Dr. Patrick Skelly and Dr. Saul Tzipori for helpful discussions. This project was funded in part with Federal funds from the NIAID , NIH , DHHS , under Contract No. N01-AI-30050 and Award Number U54 AI057159. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Allergy and Infectious Diseases or the National Institutes of Health. ",
year = "2010",
month = nov,
doi = "10.1016/j.toxicon.2010.07.003",
language = "English (US)",
volume = "56",
pages = "990--998",
journal = "Toxicon",
issn = "0041-0101",
publisher = "Elsevier Ltd",
number = "6",
}