{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,3]],"date-time":"2026-10-03T16:21:42Z","timestamp":1791044502195,"version":"4.1.0"},"reference-count":58,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2023,3,6]],"date-time":"2023-03-06T00:00:00Z","timestamp":1678060800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Biosafety Research Project","award":["20SWAQX23-002-002"],"award-info":[{"award-number":["20SWAQX23-002-002"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Toxins"],"abstract":"<jats:p>Botulinum neurotoxins (BoNTs) can cause nerve paralysis syndrome in mammals and other vertebrates. BoNTs are the most toxic biotoxins known and are classified as Class A biological warfare agents. BoNTs are mainly divided into seven serotypes A-G and new neurotoxins BoNT\/H and BoNT\/X, which have similar functions. BoNT proteins are 150 kDa polypeptide consisting of two chains and three domains: the light chain (L, catalytic domain, 50 kDa) and the heavy chain (H, 100 kDa), which can be divided into an N-terminal membrane translocation domain (HN, 50 kDa) and a C-terminal receptor binding domain (Hc, 50 kDa). In current study, we explored the immunoprotective efficacy of each functional molecule of BoNT\/F and the biological characteristics of the light chain-heavy N-terminal domain (FL-HN). The two structure forms of FL-HN (i.e., FL-HN-SC: single chain FL-HN and FL-HN-DC: di-chain FL-HN) were developed and identified. FL-HN-SC could cleave the vesicle associated membrane protein 2 (VAMP2) substrate protein in vitro as FL-HN-DC or FL. While only FL-HN-DC had neurotoxicity and could enter neuro-2a cells to cleave VAMP2. Our results showed that the FL-HN-SC had a better immune protection effect than the Hc of BoNT\/F (FHc), which indicated that L-HN-SC, as an antigen, provided the strongest protective effects against BoNT\/F among all the tested functional molecules. Further in-depth research on the different molecular forms of FL-HN suggested that there were some important antibody epitopes at the L-HN junction of BoNT\/F. Thus, FL-HN-SC could be used as a subunit vaccine to replace the FHc subunit vaccine and\/or toxoid vaccine, and to develop antibody immune molecules targeting L and HN domains rather than the FHc domain. FL-HN-DC could be used as a new functional molecule to evaluate and explore the structure and activity of toxin molecules. Further exploration of the biological activity and molecular mechanism of the functional FL-HN or BoNT\/F is warranted.<\/jats:p>","DOI":"10.3390\/toxins15030200","type":"journal-article","created":{"date-parts":[[2023,3,6]],"date-time":"2023-03-06T02:06:43Z","timestamp":1678068403000},"page":"200","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":10,"title":["Biological and Immunological Characterization of a Functional L-HN Derivative of Botulinum Neurotoxin Serotype F"],"prefix":"10.3390","volume":"15","author":[{"given":"Zhiying","family":"Li","sequence":"first","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"},{"name":"Pharmaceutical College, Henan University, Kaifeng 475001, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bolin","family":"Li","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiansheng","family":"Lu","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xuyang","family":"Liu","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"},{"name":"Pharmaceutical College, Henan University, Kaifeng 475001, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiao","family":"Tan","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"},{"name":"Institute of Life Science and Biotechnology, Beijing Jiaotong University, Beijing 100044, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rong","family":"Wang","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peng","family":"Du","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shuo","family":"Yu","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9680-7356","authenticated-orcid":false,"given":"Qing","family":"Xu","sequence":"additional","affiliation":[{"name":"Institute of Life Science and Biotechnology, Beijing Jiaotong University, Beijing 100044, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaobin","family":"Pang","sequence":"additional","affiliation":[{"name":"Pharmaceutical College, Henan University, Kaifeng 475001, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5124-9688","authenticated-orcid":false,"given":"Yunzhou","family":"Yu","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhixin","family":"Yang","sequence":"additional","affiliation":[{"name":"Beijing Institute of Biotechnology, Beijing 100071, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,3,6]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"200","DOI":"10.1124\/pr.116.012658","article-title":"Botulinum neurotoxins: Biology, pharmacology, and toxicology","volume":"69","author":"Pirazzini","year":"2017","journal-title":"Pharmacol. Rev."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Choudhury, S., Baker, M.R., Chatterjee, S., and Kumar, H. (2021). Botulinum toxin: An update on pharmacology and newer products in development. Toxins, 13.","DOI":"10.3390\/toxins13010058"},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Rossetto, O., and Montecucco, C. (2019). Tables of toxicity of botulinum and tetanus neurotoxins. Toxins, 11.","DOI":"10.3390\/toxins11120686"},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Rasetti-Escargueil, C., and Popoff, M.R. (2019). Antibodies and vaccines against botulinum toxins: Available measures and novel approaches. Toxins, 11.","DOI":"10.3390\/toxins11090528"},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Ben David, A., Barnea, A., Diamant, E., Dor, E., Schwartz, A., Torgeman, A., and Zichel, R. (2021). Small molecule receptor binding inhibitors with in vivo efficacy against botulinum neurotoxin serotypes a and e. Int. J. Mol. Sci., 22.","DOI":"10.3390\/ijms22168577"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Hobbs, R.J., Thomas, C.A., Halliwell, J., and Gwenin, C.D. (2019). Rapid detection of botulinum neurotoxins\u2014A review. Toxins, 11.","DOI":"10.3390\/toxins11070418"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"290","DOI":"10.1016\/j.resmic.2014.09.007","article-title":"Historical and current perspectives on clostridium botulinum diversity","volume":"166","author":"Smith","year":"2015","journal-title":"Res. Microbiol."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Peck, M.W., Smith, T.J., Anniballi, F., Austin, J.W., Bano, L., Bradshaw, M., Cuervo, P., Cheng, L.W., Derman, Y., and Dorner, B.G. (2017). Historical perspectives and guidelines for botulinum neurotoxin subtype nomenclature. Toxins, 9.","DOI":"10.3390\/toxins9010038"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Tehran, D.A., and Pirazzini, M. (2018). Novel botulinum neurotoxins: Exploring underneath the iceberg tip. Toxins, 10.","DOI":"10.3390\/toxins10050190"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1007\/164_2019_342","article-title":"The structure and classification of botulinum toxins","volume":"Volume 263","author":"Dong","year":"2021","journal-title":"Botulinum Toxin Therapy"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"811","DOI":"10.1146\/annurev-biochem-013118-111654","article-title":"Botulinum and tetanus neurotoxins","volume":"88","author":"Dong","year":"2019","journal-title":"Annu. Rev. Biochem."},{"key":"ref_12","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. Rev. Microbiol."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Luvisetto, S. (2021). Botulinum neurotoxins in central nervous system: An overview from animal models to human therapy. Toxins, 13.","DOI":"10.3390\/toxins13110751"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1016\/j.toxicon.2020.02.006","article-title":"Neuronal selectivity of botulinum neurotoxins","volume":"178","author":"Poulain","year":"2020","journal-title":"Toxicon"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Cai, S., Kumar, R., and Singh, B.R. (2021). Clostridial neurotoxins: Structure, function and implications to other bacterial toxins. Microorganisms, 9.","DOI":"10.3390\/microorganisms9112206"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Fonfria, E., Elliott, M., Beard, M., Chaddock, J.A., and Krupp, J. (2018). Engineering botulinum toxins to improve and expand targeting and snare cleavage activity. Toxins, 10.","DOI":"10.3390\/toxins10070278"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"789","DOI":"10.1038\/nsmb.1626","article-title":"Mode of vamp substrate recognition and inhibition of clostridium botulinum neurotoxin f","volume":"16","author":"Agarwal","year":"2009","journal-title":"Nat. Struct. Mol. Biol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"277","DOI":"10.1042\/BJ20101310","article-title":"Molecular mechanisms of substrate recognition and specificity of botulinum neurotoxin serotype f","volume":"433","author":"Chen","year":"2011","journal-title":"Biochem. J."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"10","DOI":"10.1055\/s-0035-1571215","article-title":"The botulinum toxin as a therapeutic agent: Molecular structure and mechanism of action in motor and sensory systems","volume":"36","author":"Kumar","year":"2016","journal-title":"Semin. Neurol."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Gardner, A.P., and Barbieri, J.T. (2018). Light chain diversity among the botulinum neurotoxins. Toxins, 10.","DOI":"10.3390\/toxins10070268"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"67","DOI":"10.1016\/j.toxicon.2020.02.001","article-title":"Potency and stability of a trivalent, catalytically inactive vaccine against botulinum neurotoxin serotypes c, e and f (tricef)","volume":"176","author":"Webb","year":"2020","journal-title":"Toxicon"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"481","DOI":"10.1586\/erv.13.37","article-title":"What next for botulism vaccine development?","volume":"12","author":"Webb","year":"2013","journal-title":"Expert Rev. Vaccines"},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Poulain, B., and Popoff, M.R. (2019). Why are botulinum neurotoxin-producing bacteria so diverse and botulinum neurotoxins so toxic?. Toxins, 11.","DOI":"10.3390\/toxins11010034"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"233","DOI":"10.1089\/hyb.2012.0017","article-title":"High sensitivity elisa for detection of botulinum neurotoxin serotype f","volume":"31","author":"Liu","year":"2012","journal-title":"Hybridoma"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"2041","DOI":"10.1016\/j.vaccine.2018.02.042","article-title":"Safety and immunogenicity of investigational recombinant botulinum vaccine, rbv a\/b, in volunteers with pre-existing botulinum toxoid immunity","volume":"36","author":"Khouri","year":"2018","journal-title":"Vaccine"},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Sundeen, G., and Barbieri, J.T. (2017). Vaccines against botulism. Toxins, 9.","DOI":"10.3390\/toxins9090268"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Lou, J., and Marks, J.D. (2018). Botulinum neurotoxins (bonts)\u2014Antibody and vaccine. Toxins, 10.","DOI":"10.3390\/toxins10120495"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"6998","DOI":"10.1128\/IAI.73.10.6998-7005.2005","article-title":"Characterization of the antibody response to the receptor binding domain of botulinum neurotoxin serotypes a and e","volume":"73","author":"Baldwin","year":"2005","journal-title":"Infect. Immun."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1314","DOI":"10.1128\/IAI.01025-07","article-title":"Subunit vaccine against the seven serotypes of botulism","volume":"76","author":"Baldwin","year":"2008","journal-title":"Infect. Immun."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2978","DOI":"10.1016\/j.vaccine.2020.02.060","article-title":"Immunological characterisation and immunoprotective efficacy of functional domain antigens of botulinum neurotoxin serotype A","volume":"38","author":"Liu","year":"2020","journal-title":"Vaccine"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"755","DOI":"10.1080\/21645515.2018.1547613","article-title":"Development and evaluation of candidate subunit vaccine against botulinum neurotoxin serotype b","volume":"15","author":"Shi","year":"2019","journal-title":"Hum. Vaccin. Immunother."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"100","DOI":"10.1080\/21645515.2019.1633878","article-title":"Development and evaluation of candidate subunit vaccine and novel antitoxin against botulinum neurotoxin serotype e","volume":"16","author":"Shi","year":"2020","journal-title":"Hum. Vaccin. Immunother."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1819","DOI":"10.1128\/CVI.00239-08","article-title":"Evaluation of a recombinant hc of clostridium botulinum neurotoxin serotype f as an effective subunit vaccine","volume":"15","author":"Yu","year":"2008","journal-title":"Clin. Vaccine. Immunol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"2048621","DOI":"10.1080\/21645515.2022.2048621","article-title":"Development and evaluation of a tetravalent botulinum vaccine","volume":"18","author":"Shi","year":"2022","journal-title":"Hum. Vaccin. Immunother."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"15776","DOI":"10.1038\/srep15776","article-title":"Antigenic sites on the hn domain of botulinum neurotoxin a stimulate protective antibody responses against active toxin","volume":"5","author":"Ayyar","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"51","DOI":"10.1016\/j.pep.2018.01.014","article-title":"High level expression and immunochemical characterization of botulinum neurotoxin type f light chain","volume":"146","author":"Chauhan","year":"2018","journal-title":"Protein Expr. Purif."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1044","DOI":"10.1007\/s12640-021-00337-x","article-title":"Recombinant l-hn fusion antigen derived from the l and hn domains of botulinum neurotoxin b stimulates a protective antibody response against active neurotoxin","volume":"39","author":"Li","year":"2021","journal-title":"Neurotox. Res."},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Li, Z., Lu, J., Tan, X., Wang, R., Xu, Q., Yu, Y., and Yang, Z. (2022). Functional el-hn fragment as a potent candidate vaccine for the prevention of botulinum neurotoxin serotype e. Toxins, 14.","DOI":"10.3390\/toxins14020135"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Parrera, G.S., Astacio, H., Tunga, P., Anderson, D.M., Hall, C.L., and Richardson, J.S. (2021). Use of botulism antitoxin heptavalent (a, b, c, d, e, f, g)-(equine) (bat((r))) in clinical study subjects and patients: A 15-year systematic safety review. Toxins, 14.","DOI":"10.3390\/toxins14010019"},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Lonati, D., Schicchi, A., Crevani, M., Buscaglia, E., Scaravaggi, G., Maida, F., Cirronis, M., Petrolini, V.M., and Locatelli, C.A. (2020). Foodborne botulism: Clinical diagnosis and medical treatment. Toxins, 12.","DOI":"10.3390\/toxins12080509"},{"key":"ref_41","unstructured":"Centers for Disease Control and Prevention (CDC) (2011). Notice of cdc\u2019s discontinuation of investigational pentavalent (abcde) botulinum toxoid vaccine for workers at risk for occupational exposure to botulinum toxins. MMWR Morb. Mortal. Wkly. Rep., 60, 1454\u20131455."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1615\/CritRevImmunol.v30.i2.50","article-title":"Mode of action of botulinum neurotoxins: Current vaccination strategies and molecular immune recognition","volume":"30","author":"Aoki","year":"2010","journal-title":"Crit. Rev. Immunol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"827","DOI":"10.1016\/j.vaccine.2017.12.064","article-title":"Enhancing toxin-based vaccines against botulism","volume":"36","author":"Przedpelski","year":"2018","journal-title":"Vaccine"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"2795","DOI":"10.1128\/IAI.01252-08","article-title":"Bivalent recombinant vaccine for botulinum neurotoxin types a and b based on a polypeptide comprising their effector and translocation domains that is protective against the predominant a and b subtypes","volume":"77","author":"Shone","year":"2009","journal-title":"Infect. Immun."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"52","DOI":"10.1016\/j.jsb.2010.11.010","article-title":"Structure and activity of a functional derivative of clostridium botulinum neurotoxin b","volume":"174","author":"Masuyer","year":"2011","journal-title":"J. Struct. Biol."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"1465","DOI":"10.1128\/IAI.00063-15","article-title":"The c-terminal heavy-chain domain of botulinum neurotoxin a is not the only site that binds neurons, as the n-terminal heavy-chain domain also plays a very active role in toxin-cell binding and interactions","volume":"83","author":"Ayyar","year":"2015","journal-title":"Infect. Immun."},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Burgin, D., Perier, C., Hackett, G., Elliott, M., Kwan, D., Hornby, F., Mir, I., Maignel, J., Liu, S.M., and Beard, M. (2021). New modified recombinant botulinum neurotoxin type f with enhanced potency. Toxins, 13.","DOI":"10.3390\/toxins13120834"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1678","DOI":"10.1016\/j.bbapap.2016.09.001","article-title":"Effects of membrane properties on the binding activities of the hn and hc heavy-chain domains of botulinum neurotoxin a","volume":"1864","author":"Ayyar","year":"2016","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"502","DOI":"10.1111\/1348-0421.12796","article-title":"Translocation domain of botulinum neurotoxin a subtype 2 potently induces entry into neuronal cells","volume":"64","author":"Kohda","year":"2020","journal-title":"Microbiol. Immunol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"319","DOI":"10.1615\/CritRevImmunol.v27.i4.30","article-title":"Molecular bases of protective immune responses against botulinum neurotoxin a--how antitoxin antibodies block its action","volume":"27","author":"Atassi","year":"2007","journal-title":"Crit. Rev. Immunol."},{"key":"ref_51","doi-asserted-by":"crossref","unstructured":"Fan, Y., Garcia-Rodriguez, C., Lou, J., Wen, W., Conrad, F., Zhai, W., Smith, T.J., Smith, L.A., and Marks, J.D. (2017). A three monoclonal antibody combination potently neutralizes multiple botulinum neurotoxin serotype f subtypes. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0174187"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"321","DOI":"10.1093\/protein\/gzq111","article-title":"Neutralizing human monoclonal antibodies binding multiple serotypes of botulinum neurotoxin","volume":"24","author":"Geren","year":"2011","journal-title":"Protein Eng. Des. Sel."},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Garcia-Rodriguez, C., Razai, A., Geren, I.N., Lou, J., Conrad, F., Wen, W.H., Farr-Jones, S., Smith, T.J., Brown, J.L., and Skerry, J.C. (2018). A three monoclonal antibody combination potently neutralizes multiple botulinum neurotoxin serotype e subtypes. Toxins, 10.","DOI":"10.3390\/toxins10030105"},{"key":"ref_54","doi-asserted-by":"crossref","unstructured":"Garcia-Rodriguez, C., Yan, S., Geren, I.N., Knopp, K.A., Dong, J., Sun, Z., Lou, J., Conrad, F., Wen, W.H., and Farr-Jones, S. (2021). A four-monoclonal antibody combination potently neutralizes multiple botulinum neurotoxin serotypes c and d. Toxins, 13.","DOI":"10.3390\/toxins13090641"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"1606","DOI":"10.1093\/infdis\/jiv770","article-title":"Immunological characterization and neutralizing ability of monoclonal antibodies directed against botulinum neurotoxin type h","volume":"213","author":"Fan","year":"2016","journal-title":"J. Infect. Dis."},{"key":"ref_56","doi-asserted-by":"crossref","unstructured":"Tremblay, J.M., Vazquez-Cintron, E., Lam, K.H., Mukherjee, J., Bedenice, D., Ondeck, C.A., Conroy, M.T., Bodt, S.M.L., Winner, B.M., and Webb, R.P. (2020). Camelid vhh antibodies that neutralize botulinum neurotoxin serotype e intoxication or protease function. Toxins, 12.","DOI":"10.3390\/toxins12100611"},{"key":"ref_57","doi-asserted-by":"crossref","unstructured":"Lam, K.H., Tremblay, J.M., Perry, K., Ichtchenko, K., Shoemaker, C.B., and Jin, R. (2022). Probing the structure and function of the protease domain of botulinum neurotoxins using single-domain antibodies. PLoS Pathog., 18.","DOI":"10.1371\/journal.ppat.1010169"},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"271","DOI":"10.1016\/j.clim.2010.07.005","article-title":"Development and evaluation of candidate vaccine and antitoxin against botulinum neurotoxin serotype f","volume":"137","author":"Yu","year":"2010","journal-title":"Clin. Immunol."}],"container-title":["Toxins"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6651\/15\/3\/200\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T18:48:40Z","timestamp":1760122120000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6651\/15\/3\/200"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,3,6]]},"references-count":58,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2023,3]]}},"alternative-id":["toxins15030200"],"URL":"https:\/\/doi.org\/10.3390\/toxins15030200","relation":{},"ISSN":["2072-6651"],"issn-type":[{"value":"2072-6651","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,3,6]]}}}