Oku Y, Yutsudo T, Hirayama T, O'Brien A D, Takeda Y
Department of Bacterial Infection, Institute of Medical Science, University of Tokyo, Japan.
Microb Pathog. 1989 Feb;6(2):113-22. doi: 10.1016/0882-4010(89)90014-4.
A cytotoxin to Vero cells (Vero toxin), which was immunologically related to Shiga-like toxin II (SLT-II) (or VT2), was purified from a stain of Escherichia coli isolated from a patient with hemolytic uremic syndrome. The toxin was active on Vero cells but much less active on HeLa cells, a property similar to that of the recently identified SLT-II variant from E. coli strains that caused edema disease of swine. Thus the toxin purified in this report was tentatively named Shiga-like toxin II variant (Vero toxin 2 variant). The purification procedures consisted of ammonium sulfate fractionation, DEAE-Sepharose CL-6B column chromatography, chromatofocusing column chromatography, and repeated high performance liquid chromatography (HPLC) on TSK-gel G-2000SW column and on TSK-gel DEAE-5PW columns. About 90 micrograms of purified toxin was obtained from 451 of the culture supernatant with a yield of about 16%. The purified toxin consisted of A and B subunits of molecular sizes similar to those of SLT-II (VT2). The isoelectric point of the purified toxin was 6.1, which was different from that of SLT-II (VT2) (pI = 4.1). In an Ouchterlony double gel diffusion test, purified toxin and SLT-II (VT2) formed precipitin lines with spur formation against anti-purified toxin and anti-SLT-II (anti-VT2), respectively. The purified toxin was cytotoxic to Vero cells, about 6 pg of the toxin killing 50% of the Vero cells, and showed lethal toxicity to mice when injected intraperitoneally, the LD50 being about 2.7 ng per mouse.
从一名溶血尿毒综合征患者分离出的大肠杆菌菌株中纯化出一种对Vero细胞有细胞毒性的毒素（Vero毒素），它与志贺样毒素II（SLT-II，或VT2）在免疫学上相关。该毒素对Vero细胞有活性，但对HeLa细胞的活性要低得多，这一特性与最近从引起猪水肿病的大肠杆菌菌株中鉴定出的SLT-II变体相似。因此，本报告中纯化的毒素被暂命名为志贺样毒素II变体（Vero毒素2变体）。纯化步骤包括硫酸铵分级分离、DEAE-琼脂糖CL-6B柱色谱、色谱聚焦柱色谱，以及在TSK-gel G-2000SW柱和TSK-gel DEAE-5PW柱上重复进行高效液相色谱（HPLC）。从451升培养上清液中获得了约90微克的纯化毒素，产率约为16%。纯化的毒素由分子大小与SLT-II（VT2）相似的A和B亚基组成。纯化毒素的等电点为6.1，与SLT-II（VT2）的等电点（pI = 4.1）不同。在双向琼脂扩散试验中，纯化毒素和SLT-II（VT2）分别与抗纯化毒素和抗SLT-II（抗VT2）形成了带有刺突的沉淀线。纯化毒素对Vero细胞具有细胞毒性，约6皮克毒素可杀死50%的Vero细胞，腹腔注射时对小鼠显示出致死毒性，半数致死量约为每只小鼠2.7纳克。
