Functional EL-HN Fragment as a Potent Candidate Vaccine for the Prevention of Botulinum Neurotoxin Serotype E

Li, Zhen; Lu, Jian-Sheng; Tan, Xiao; Wang, Rong; Chen, Nansheng; Yu, Yun-Zhou; Yang, Zhengming

doi:10.3390/toxins14020135

Cited by 6 publications

(12 citation statements)

References 49 publications

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“…In recent years, the L and/or HN domains of BoNTs have been proved to generate protective antibodies [ 35 , 36 , 42 , 43 ]. In our studies, we also found that the L-HN molecules of BoNT serotypes A, B, and E have strong immunoprotective effects [ 30 , 37 , 38 ] as per the reports of Shone et al on the translocation and effector domains BoNT/A and B [ 44 ]. The L-HN molecule of BoNT/E has been shown to have better immunoprotective efficacy than its Hc domain [ 32 ], which indicated that the L-HN molecule is superior to toxoid vaccines in terms of production and immune protection efficacy.…”

Section: Discussionsupporting

confidence: 87%

“…To further study the biological properties and functions of FL-HN at the cellular level, neuro-2a cells were selected as the cell model for related experiments. The molecules derived from BoNT/F were incubated with neuro-2a cells, and the activity of FL-HN was assessed by Western blotting, which could detect the content of cleaved rVAMP2 protein at the cellular level [ 36 , 37 , 38 , 39 ]. After co-incubating FL-HN with neuro-2a cells for 12 h, proteins in the cells were extracted for Western blotting detection.…”

Section: Resultsmentioning

confidence: 99%

“…However, some studies have found that the translocation domains in BoNT/A and BoNT/B are also involved in the binding process [ 45 , 46 ]. In our study, we found that the EL-HN molecule, lacking the Hc domain, can also exert a little neurotoxicity in animals, indicating HN might play a receptor binding role in neurotoxicity, similar to Hc domain [ 38 ]. To explore the biological activity of FL-HN, we mutated HELIH (amino acids 227–231) of FL-HN into AALIA to construct and prepare FmL-HN.…”

Section: Discussionmentioning

confidence: 99%

“…Currently, the recombinant Hc subunit vaccines of BoNT/A, B, E, and F all have good protection against the biologically active BoNTs of their own serotype after immunization [ 30 , 31 , 32 , 33 , 34 ]. The L and HN domains of BoNTs have also been developed as recombinant subunit vaccines with strong potency to prevent animal botulism [ 30 , 35 , 36 , 37 , 38 ].…”

Section: Introductionmentioning

confidence: 99%

“…To define the function of the L-HN molecules of all BoNTs, we explored their biological and immunological characteristics [ 30 , 37 , 38 ]. Subsequently, in the present study, BoNT/F was selected to further explore the biological and immunological characterization of its functional molecules.…”

Section: Introductionmentioning

confidence: 99%

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Biological and Immunological Characterization of a Functional L-HN Derivative of Botulinum Neurotoxin Serotype F

Lu²,

Liu

et al. 2023

Toxins

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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.

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Section: Discussionsupporting

confidence: 87%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Biological and Immunological Characterization of a Functional L-HN Derivative of Botulinum Neurotoxin Serotype F

Lu²,

Liu

et al. 2023

Toxins

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Characterization of a novel tetravalent botulism antitoxin based on receptor-binding domain of BoNTs

Shi¹,

Lu²,

Mao³

et al. 2023

Appl Microbiol Biotechnol

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Botulinum neurotoxin (BoNTs; serotypes A, B, E, and F) cause botulism disease in humans, which could be effectively treated using antitoxins. Herein, we established a novel receptor-binding domain (RBD)-based antitoxin using recombinant C terminal heavy chain (Hc) domains of BoNTs as immunogens. Immunization of horses with these recombinant Hc domains allowed the purification and digestion of IgGs from hyper-immune sera to produce high-quality and high-efficiency monovalent botulism antitoxin F(ab′) 2 against each BoNT (M-BATs). However, these M-BATs could not bind or neutralize other serotypes of BoNTs, and that there were no cross-protective effects among these M-BATs. This suggested the need to prepare tetravalent antitoxins to neutralize the four BoNTs simultaneously. Thus, these M-BATs were formulated into a novel tetravalent botulism antitoxin (T-BAT), in which a 10-ml volume contained 10000 IU of BoNT/A and 5000 IU of BoNT/B, BoNT/E, and BoNT/F antitoxins. The novel antitoxin preparation could prevent and treat the four mixed botulinum neurotoxins simultaneously in vivo, representing strong efficacy in an animal poisoning model. Moreover, these antibodies in T-BAT could bind the RBD, whereas conventional antitoxins based on inactivated toxins mainly bind the light chain or heavy chain translocation domain (HN) and weakly bind the important RBD in current experimental conditions. The high levels of RBD-specific novel antitoxins can efficiently bind the RBD and neutralize natural or recombinant toxins containing this RBD. The findings of the present study experimentally support the use of RBD-specific antitoxins to treat BoNT serotype A, B, E, and F-mediated botulism. This study demonstrated the concept of developing potent novel multivalent antitoxins against all BoNTs or other toxins, using the RBD of these toxins as an alternative antigen to inactivated toxins. Key points • Antitoxins based on the receptor-binding domains of botulinum neurotoxins were made. • Novel antitoxin binds RBD; traditional antitoxin mainly binds light chain or HN domain. • A tetravalent antitoxin could prevent and treat the four mixed neurotoxins in vivo. Supplementary Information The online version contains supplementary material available at 10.1007/s00253-023-12515-2.

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A human bispecific antibody neutralizes botulinum neurotoxin serotype A

Lu,

Jiang,

Guo

et al. 2023

Sci Rep

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Botulinum neurotoxin (BoNT) shows high lethality and toxicity, marking it as an important biological threat. The only effective post-exposure therapy is botulinum antitoxin; however, such products have great potential for improvement. To prevent or treat BoNT, monoclonal antibodies (mAbs) are promising agents. Herein, we aimed to construct a bispecific antibody (termed LUZ-A1-A3) based on the anti-BoNT/A human monoclonal antibodies (HMAb) A1 and A3. LUZ-A1-A3 binds to the Hc and L-HN domains of BoNT/A, displaying potent neutralization activity against BoNT/A (124 × higher than that of HMAb A1 or HMAb A3 alone and 15 × higher than that of the A1 + A3 combination). LUZ-A1-A3 provided effective protection against BoNT/A in an in vivo mouse model. Mice were protected from infection with 500 × LD50 of BoNT/A by LUZ-A1-A3 from up to 7 days before intraperitoneal administration of BoNT/A. We also demonstrated the effective therapeutic capacity of LUZ-A1-A3 against BoNT/A in a mouse model. LUZ-A1-A3 (5 μg/mouse) neutralized 20 × LD50 of BoNT/A at 3 h after intraperitoneal BoNT/A administration and complete neutralized 20 × LD50 of BoNT/A at 0.5 h after intraperitoneal BoNT/A administration. Thus, LUZ-A1-A3 is a promising agent for the pre-exposure prophylaxis and post-exposure treatment of BoNT/A.

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Functional EL-HN Fragment as a Potent Candidate Vaccine for the Prevention of Botulinum Neurotoxin Serotype E

Cited by 6 publications

References 49 publications

Biological and Immunological Characterization of a Functional L-HN Derivative of Botulinum Neurotoxin Serotype F

Biological and Immunological Characterization of a Functional L-HN Derivative of Botulinum Neurotoxin Serotype F

Characterization of a novel tetravalent botulism antitoxin based on receptor-binding domain of BoNTs

A human bispecific antibody neutralizes botulinum neurotoxin serotype A

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