Yu Rui, Yi Shaoqiong, Yu Changming, Fang Ting, Liu Shuling, Yu Ting, Song Xiaohong, Fu Ling, Hou Lihua, Chen Wei
State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, 20 Dongdajie Street, Fengtai, Beijing 100071, China.
Clin Vaccine Immunol. 2011 Oct;18(10):1668-72. doi: 10.1128/CVI.05244-11. Epub 2011 Aug 3.
The C fragment of tetanus neurotoxin (TeNT-Hc) with different conformations was observed due to the four cysteine residues within it which could form different intramolecular disulfide bonds. In this study, we prepared and compared three types of monomeric TeNT-Hc with different conformational components: free sulfhydryls (50 kDa), bound sulfhydryls (44 kDa), and a mixture of the two conformational proteins (half 50 kDa and half 44 kDa). TeNT-Hc with bound sulfhydryls reduced its binding activity to ganglioside G(T1b) and neuronal PC-12 cells compared to what was seen for TeNT-Hc with free sulfhydryls. However, there was no significant difference among their immunogenicities in mice, including induction of antitetanus toxoid IgG titers, antibody types, and protective capacities against tetanus neurotoxin challenge. Our results showed that the conformational changes of TeNT-Hc resulting from disulfide bond formation reduced its ganglioside-binding activity but did not destroy its immunogenicity, and the protein still retained continuous B cell and T cell epitopes; that is, the presence of the ganglioside-binding site within TeNT-Hc may be not essential for the induction of a fully protective antitetanus response. TeNT-Hc with bound sulfhydryls may be developed into an ideal human vaccine with a lower potential for side effects.
破伤风神经毒素的C片段（TeNT-Hc）由于其内部的四个半胱氨酸残基可形成不同的分子内二硫键，从而呈现出不同的构象。在本研究中，我们制备并比较了三种具有不同构象成分的单体TeNT-Hc：游离巯基型（50 kDa）、结合巯基型（44 kDa）以及两种构象蛋白的混合物（50 kDa和44 kDa各占一半）。与游离巯基型TeNT-Hc相比，结合巯基型TeNT-Hc对神经节苷脂G(T1b)和神经元PC-12细胞的结合活性降低。然而，它们在小鼠体内的免疫原性并无显著差异，包括抗破伤风类毒素IgG滴度的诱导、抗体类型以及针对破伤风神经毒素攻击的保护能力。我们的结果表明，二硫键形成导致的TeNT-Hc构象变化降低了其神经节苷脂结合活性，但并未破坏其免疫原性，该蛋白仍保留连续的B细胞和T细胞表位；也就是说，TeNT-Hc中神经节苷脂结合位点的存在对于诱导完全保护性的抗破伤风反应可能并非必需。结合巯基型TeNT-Hc可能被开发成为一种副作用潜力较低的理想人用疫苗。
