{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T07:07:00Z","timestamp":1782284820985,"version":"3.54.5"},"reference-count":68,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2020,9,27]],"date-time":"2020-09-27T00:00:00Z","timestamp":1601164800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01AI139087; R21AI139690; R21AI123920; R01AI125704"],"award-info":[{"award-number":["R01AI139087; R21AI139690; R21AI123920; R01AI125704"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000057","name":"National Institute of General Medical Sciences","doi-asserted-by":"publisher","award":["P30 GM124165"],"award-info":[{"award-number":["P30 GM124165"]}],"id":[{"id":"10.13039\/100000057","id-type":"DOI","asserted-by":"publisher"}]},{"name":"NIH-ORIP HEI grant","award":["S10OD021527"],"award-info":[{"award-number":["S10OD021527"]}]},{"name":"U.S. DOE under Contract","award":["DE-AC02-06CH11357"],"award-info":[{"award-number":["DE-AC02-06CH11357"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Toxins"],"abstract":"<jats:p>Botulinum neurotoxin serotype E (BoNT\/E) is one of the major causes of human botulism, which is a life-threatening disease caused by flaccid paralysis of muscles. After receptor-mediated toxin internalization into motor neurons, the translocation domain (HN) of BoNT\/E transforms into a protein channel upon vesicle acidification in endosomes and delivers its protease domain (LC) across membrane to enter the neuronal cytosol. It is believed that the rapid onset of BoNT\/E intoxication compared to other BoNT serotypes is related to its swift internalization and translocation. We recently identified two neutralizing single-domain camelid antibodies (VHHs) against BoNT\/E1 termed JLE-E5 and JLE-E9. Here, we report the crystal structures of these two VHHs bound to the LCHN domain of BoNT\/E1. The structures reveal that these VHHs recognize two distinct epitopes that are partially overlapping with the putative transmembrane regions on HN, and therefore could physically block membrane association of BoNT\/E1. This is confirmed by our in vitro studies, which show that these VHHs inhibit the structural change of BoNT\/E1 at acidic pH and interfere with BoNT\/E1 association with lipid vesicles. Therefore, these two VHHs neutralize BoNT\/E1 by preventing the transmembrane delivery of LC. Furthermore, structure-based sequence analyses show that the 3-dimensional epitopes of these two VHHs are largely conserved across many BoNT\/E subtypes, suggesting a broad-spectrum protection against the BoNT\/E family. In summary, this work improves our understanding of the membrane translocation mechanism of BoNT\/E and paves the way for developing VHHs as diagnostics or therapeutics for the treatment of BoNT\/E intoxication.<\/jats:p>","DOI":"10.3390\/toxins12100616","type":"journal-article","created":{"date-parts":[[2020,9,28]],"date-time":"2020-09-28T10:39:58Z","timestamp":1601289598000},"page":"616","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":17,"title":["Two VHH Antibodies Neutralize Botulinum Neurotoxin E1 by Blocking Its Membrane Translocation in Host Cells"],"prefix":"10.3390","volume":"12","author":[{"given":"Kwok-Ho","family":"Lam","sequence":"first","affiliation":[{"name":"Department of Physiology &amp; Biophysics, University of California, Irvine, CA 92617, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4046-1704","authenticated-orcid":false,"given":"Kay","family":"Perry","sequence":"additional","affiliation":[{"name":"NE-CAT and Department of Chemistry and Chemical Biology, Cornell University, Argonne National Laboratory, Argonne, IL 60439, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9738-1361","authenticated-orcid":false,"given":"Charles B.","family":"Shoemaker","sequence":"additional","affiliation":[{"name":"Department of Infectious Disease and Global Health, Cummings School of Veterinary Medicine, Tufts University, North Grafton, MA 01536, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0348-7363","authenticated-orcid":false,"given":"Rongsheng","family":"Jin","sequence":"additional","affiliation":[{"name":"Department of Physiology &amp; Biophysics, University of California, Irvine, CA 92617, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,9,27]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"535","DOI":"10.1038\/nrmicro3295","article-title":"Botulinum neurotoxins: Genetic, structural and mechanistic insights","volume":"12","author":"Rossetto","year":"2014","journal-title":"Nat. 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