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首页> 外文期刊>Infection and immunity >Binding and Internalization of Clostridium perfringens Iota-Toxin in Lipid Rafts
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Binding and Internalization of Clostridium perfringens Iota-Toxin in Lipid Rafts

机译：产气荚膜梭菌碘毒素在脂质筏中的结合和内在化

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Clostridium perfringens iota-toxin is a binary toxin composed of an enzymatic component (Ia) and a binding component (Ib). The oligomer of Ib formed in membranes induces endocytosis. We examined the binding and internalization of Ib by using Cy3-labeled Ib. Labeled Ib was retained at the membranes of MDCK cells for 60 min of incubation at 37°C, and later it was detected in cytoplasmic vesicles. To determine whether Ib associates with lipid rafts, we incubated MDCK cells with Ib at 4 or 37°C and fractionated the Triton-insoluble membranes. An Ib complex of 500 kDa was localized at 37°C to the insoluble fractions that fulfilled the criteria of lipid rafts, but it did not form at 4°C. The amount of complex in the raft fraction reached a maximum after 60 min of incubation at 37°C. When the cells that were preincubated with Ib at 4°C were incubated at 37°C, the complex was detected in the raft fraction. The treatment of MDCK cells with methyl-β-cyclodextrin reduced the localization of the Ib complex to the rafts and the rounding of the cells induced by Ia plus Ib. When 125I-labeled Ia was incubated with the cells in the presence of Ib at 37°C, it was localized in the raft fraction. Surface plasmon resonance analysis revealed that Ia binds to the oligomer of Ib. We conclude that Ib binds to a receptor in membranes and then moves to rafts and that Ia bound to the oligomer of Ib formed in the rafts is internalized.
机译：产气荚膜梭菌（Clostridium perfringens）碘毒素是一种由酶成分（Ia）和结合成分（Ib）组成的二元毒素。在膜中形成的Ib的低聚物诱导内吞作用。我们通过使用Cy3标记的Ib检查了Ib的结合和内在化。在37°C下孵育60分钟后，标记的Ib保留在MDCK细胞膜上，随后在细胞质囊泡中检测到。为了确定Ib是否与脂质筏相关，我们在4或37°C下将MDCK细胞与Ib孵育，并分馏Triton不溶膜。 500 kDa的Ib复合物在37°C定位于满足脂筏标准的不溶级分，但在4°C未形成。在37°C孵育60分钟后，筏级分中复合物的量达到最大值。将在4°C下用Ib预孵育的细胞在37°C下孵育时，在筏部分中检测到复合物。用甲基-β-环糊精处理MDCK细胞减少了Ib复合物在木筏上的定位以及由Ia加Ib诱导的细胞的圆形化。当 125 I标记的Ia在Ib存在下于37°C与细胞一起温育时，它定位在筏部分中。表面等离子体共振分析表明，Ia与Ib的低聚物结合。我们得出的结论是，Ib与膜中的受体结合，然后移至筏中，并且与筏中形成的Ib寡聚物结合的Ia被内化。

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