The HC fragment of tetanus toxin forms stable, concentration-dependent dimers via an intermolecular disulphide bond
The HC fragment of tetanus toxin forms stable, concentration-dependent dimers via an intermolecular disulphide bond
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DOI：
10.1016/j.jmb.2006.09.050
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发表时间：
2007-01-05
影响因子：
5.6
通讯作者：
Fairweather, Neil
中科院分区：
文献类型：
--
作者：
Qazi, Omar;Bolgiano, Barbara;Fairweather, Neil
蛋白质寡聚化是许多细菌毒素毒性的先决条件。例子包括成孔细胞毒素链球菌溶血素0，其寡聚化以在膜中形成大孔，以及炭疽毒素的保护性抗原，其中七聚体复合物对于将致死因子和水肿因子递送至细胞胞质溶胶是必需的。梭菌神经毒素与神经元细胞上的受体的结合被很好地表征，但是关于这些毒素的四级结构和在中毒过程中寡聚化的作用知之甚少。我们已经研究了破伤风毒素的受体结合结构域（H-C）的寡聚化，其保留了全长毒素的结合和运输特性。电泳，尺寸排阻色谱法和质谱法被用来证明，H-C在溶液中经历浓度依赖性的寡聚化。还原剂被发现影响HC寡聚化，并使用诱变，Cys 869被证明是必不可少的这一过程。此外，低聚状态和四级结构的HC在溶液中进行了评估，使用同步辐射小角X射线散射。从头算形状分析和刚体建模加上诱变数据允许在溶液中的二聚体H-C的一个明确的模型的建设。我们提出了一个可能的机制H-C寡聚化，并讨论这可能与毒性。
Protein oligomerisation is a prerequisite for the toxicity of a number of bacterial toxins. Examples include the pore-forming cytotoxin streptolysin 0, which oligomerises to form large pores in the membrane and the protective antigen of anthrax toxin, where a heptameric complex is essential for the delivery of lethal factor and edema factor to the cell cytosol. Binding of the clostridial neurotoxins to receptors on neuronal cells is well characterised, but little is known regarding the quaternary structure of these toxins and the role of oligomerisation in the intoxication process. We have investigated the oligomerisation of the receptor binding domain (H-C) of tetanus toxin, which retains the binding and trafficking properties of the full-length toxin. Electrophoresis, size exclusion chromatography and mass spectrometry were used to demonstrate that H-C undergoes concentration-dependent oligomerisation in solution. Reducing agents were found to affect HC oligomerisation and, using mutagenesis, Cys869 was shown to be essential for this process. Furthermore, the oligomeric state and quaternary structure of HC in solution was assessed using synchrotron small-angle Xray scattering. Ab initio shape analysis and rigid body modelling coupled with mutagenesis data allowed the construction of an unequivocal model of dimeric H-C in solution. We propose a possible mechanism for H-C oligomerisation and discuss how this may relate to toxicity.