Takuhiro Matsumura, Mayu Kitamura, Sho Amatsu, Aki Yamaguchi, Nobuhide Kobayashi, Masahiro Yutani, Yukako Fujinaga
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引用次数: 0
摘要
肉毒中毒是一种致命的神经麻痹病症,由肉毒梭状芽孢杆菌及相关菌种产生的肉毒神经毒素(BoNT)引起。毒素中和抗体是治疗 BoNT 中毒最有效的方法。我们生成了中和 B 型肉毒杆菌神经毒素(BoNT/B)的人类单克隆抗体,命名为 M2 和 M4。这些抗体的组合对 BoNT/B 的毒性有很强的中和作用。在这项研究中,我们分析了这些抗体在体外的作用机制。M4 与重链的 C 端(受体结合域)结合,抑制 BoNT/B 与神经元 PC12 细胞的结合。虽然 M2 能识别轻(L)链(金属蛋白酶结构域),但在裂解试验中并不能抑制底物(VAMP2)的裂解。在较晚的时间点,M2 增加了 BoNT/B 在 PC12 细胞中的表面定位,这表明 M2 抑制了 L 链从突触小泡向细胞膜的转运。这些结果表明,M2和M4可分别抑制BoNT/B的不同过程,而多步骤抑制对单克隆抗体组合的协同效应非常重要。我们的研究结果可能有助于开发针对BoNTs的有效治疗抗体。
Neutralization mechanism of human monoclonal antibodies against type B botulinum neurotoxin
Botulism is a deadly neuroparalytic condition caused by the botulinum neurotoxin (BoNT) produced by Clostridium botulinum and related species. Toxin-neutralizing antibodies are the most effective treatments for BoNT intoxication. We generated human monoclonal antibodies neutralizing type B botulinum neurotoxin (BoNT/B), designated M2 and M4. The combination of these antibodies exhibited a strong neutralizing effect against BoNT/B toxicity. In this study, we analyzed the mechanisms of action of these antibodies in vitro. M4 binds to the C-terminus of the heavy chain (the receptor-binding domain) and inhibits BoNT/B binding to neuronal PC12 cells. Although M2 recognized the light (L) chain (the metalloprotease domain), it did not inhibit substrate (VAMP2) cleavage in the cleavage assay. M2 increased the surface localization of BoNT/B in PC12 cells at a later time point, suggesting that M2 inhibits the translocation of the L chain from synaptic vesicles to the cytosol. These results indicate that M2 and M4 inhibit the different processes of BoNT/B individually and that multistep inhibition is important for the synergistic effect of the combination of monoclonal antibodies. Our findings may facilitate the development of effective therapeutic antibodies against BoNTs.
期刊介绍:
Microbiology and Immunology is published in association with Japanese Society for Bacteriology, Japanese Society for Virology, and Japanese Society for Host Defense Research. It is peer-reviewed publication that provides insight into the study of microbes and the host immune, biological and physiological responses.
Fields covered by Microbiology and Immunology include:Bacteriology|Virology|Immunology|pathogenic infections in human, animals and plants|pathogenicity and virulence factors such as microbial toxins and cell-surface components|factors involved in host defense, inflammation, development of vaccines|antimicrobial agents and drug resistance of microbes|genomics and proteomics.