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首页> 外文期刊>Infection and immunity >Purification and characterization of a Chinese hamster ovary cell elongation factor of Vibrio hollisae.
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Purification and characterization of a Chinese hamster ovary cell elongation factor of Vibrio hollisae.

机译：霍乱弧菌中国仓鼠卵巢细胞延长因子的纯化和鉴定。

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The halophilic bacterium Vibrio hollisae, isolated from patients with diarrhea, produces an extracellular toxin which elongates Chinese hamster ovary (CHO) cells. We purified this toxin to homogeneity by sequential ammonium sulfate precipitation, gel filtration with Sephacryl S-200, hydrophobic interaction chromatography with phenyl-Sepharose CL-4B, ion-exchange chromatography with DEAE-Sephadex A-50, and affinity chromatography. The toxin is heat labile and sensitive to proteases, with an isoelectric point of about 6.5 and molecular weights of about 83,000 and 80,000, as estimated by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, respectively. The toxin did not react with immunoaffinity-purified antibodies to cholera toxin in a plate enzyme-linked immunosorbent assay and in a Western blot, and its activity could not be neutralized by anti-cholrea toxin serum. Mixed gangliosides and gangliosides GM1, GD1a, GD1b, Gq1b, GT1b, GD2, GD3, GM2, and GM3 failed to block its activity. Elongation of CHO cells induced by the toxin was not accompanied by an increase in the levels of cyclic AMP. The toxin induced intestinal fluid accumulation in suckling mice. These results and the lack of homology between V. hollisae DNA and DNA coding for cholera toxin or the heat-labile toxin of Escherichia coli suggest that the V. hollisae toxin is structurally and functionally different from other CHO cell-elongating toxins.
机译：从腹泻患者中分离出来的嗜盐菌霍乱弧菌会产生一种细胞外毒素，这种毒素会拉长中国仓鼠卵巢（CHO）细胞。我们通过依次进行硫酸铵沉淀，Sephacryl S-200凝胶过滤，苯基-Sepharose CL-4B疏水相互作用色谱，DEAE-Sephadex A-50离子交换色谱和亲和色谱将毒素纯化至均质。该毒素是热不稳定的并且对蛋白酶敏感，通过凝胶过滤和十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分别估计，其等电点约为6.5，分子量约为83,000和80,000。在平板酶联免疫吸附测定和Western印迹中，该毒素未与免疫亲和纯化的霍乱毒素抗体反应，并且其抗胆汁毒素血清无法抵消其活性。混合的神经节苷脂和神经节苷脂GM1，GD1a，GD1b，Gq1b，GT1b，GD2，GD3，GM2和GM3无法阻止其活性。毒素诱导的CHO细胞伸长并没有伴随环AMP含量的增加。该毒素在乳鼠中诱导肠液积聚。这些结果以及霍乱弧菌DNA与编码霍乱毒素或大肠杆菌的热不稳定毒素的DNA之间缺乏同源性，这表明霍乱弧菌毒素在结构和功能上与其他CHO细胞伸长毒素不同。

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