Moskaug J O, Sandvig K, Olsnes S
J Biol Chem. 1987 Jul 25;262(21):10339-45.
When 125I-labeled nicked diphtheria toxin bound to Vero cells was exposed to pH less than 5.0, a small fraction was reduced to yield A- and B-fragments. The pH required for reduction correlates well with that required to induce intoxication, and the amount of A-fragment released was of the same order as that required to intoxicate the cells. Conditions that protect cells against intoxication, such as acidification of the cytosol, treatment with anion transport inhibitors, or treatment with anti-diphtheria toxin antibodies, prevented the reduction of the interfragment disulfide in cell-bound toxin. In vitro, thioredoxin reduced nicked diphtheria toxin only at pH 5.0 and lower, and the reduction was inhibited by anti-toxin antibodies. This indicates that a conformational change in the toxin, necessary for reduction by the thioredoxin system, is prevented by the antibodies. Reduction by glutathione and cysteine was most efficient at neutral pH and was not inhibited by anti-toxin. The results are consistent with the possibility that cell-mediated reduction of the interfragment disulfide is a measure of the entry of fragment A into the cytosol.
当与Vero细胞结合的125I标记的缺口白喉毒素暴露于pH小于5.0的环境时，一小部分会被还原产生A片段和B片段。还原所需的pH与诱导中毒所需的pH密切相关，释放的A片段量与使细胞中毒所需的量处于同一数量级。保护细胞免受中毒的条件，如细胞质酸化、用阴离子转运抑制剂处理或用抗白喉毒素抗体处理，可防止细胞结合毒素中片段间二硫键的还原。在体外，硫氧还蛋白仅在pH 5.0及更低时还原缺口白喉毒素，且这种还原被抗毒素抗体抑制。这表明抗体阻止了硫氧还蛋白系统还原毒素所必需的毒素构象变化。谷胱甘肽和半胱氨酸在中性pH下的还原效率最高，且不受抗毒素抑制。这些结果与细胞介导的片段间二硫键还原是A片段进入细胞质的一种衡量指标这一可能性一致。
