Analysis of a fuctor involving trnslocation of LT to the cell.

涉及 LT 转移至细胞的功能因子的分析。

基本信息

批准号：
09670302
负责人：
TSUJI Takao
金额：
$ 1.92万
依托单位：
Fujita Health University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09670302/
关键词：
ETEC Entocytosis GM1 ganglioside Enterotoxin (LT)LT

项目摘要

We have been analyzing whether there might be a factor involving the endocytosis of ETEC enterotoxin (LT) to target cell except GMI ganglioside receptor. After LT was added to the CHO cell, Immunoprecipitates with anti-LT antibody to cell lysate was analyzed with Western blotting with anti-LT antibody. There was a band, of which molecular weight was 22,000 except the monomer of LT-B and might shift the monomer upper side of gel. Then we examined this factor. However, some other examiner reported that this shift of the monomer was caused by the formation of dimer of LT-B in the cell. Therefore, we examined whether other factor might be involved in the endocytosis of LT to make an antibody blocking the endocytosis of LT-B.Some rabbits were immunized by cell lysate, which was prepared from NCTC 2071 defecting GM1 ganglioside and we can prepared the anti-serum partially blocking the internization of LT to CHO cells. Then, we are now examining a factor reacting to this antiserum.Morecover, while we received this grant, we reported data as followed;1. LT is also coded in the chromosome, except the plasmid.2. The mutant defecting nicking site of A subunit near Arg 192 have strong adjuvant action to VZV, KKLH, Ovalbumin, BGG and measle virus.3. The nicking site near Arg 192 of the A subnit is not involved in some biological activity of LT.

我们一直在分析除GMI神经节苷脂受体外是否还有其他因素参与ETEC肠毒素（LT）对靶细胞的内吞作用。将 LT 添加至 CHO 细胞后，使用抗 LT 抗体通过蛋白质印迹分析细胞裂解液中抗 LT 抗体的免疫沉淀。除LT-B单体外，还有一条分子量为22,000的条带，可能会移动凝胶的单体上侧。然后我们检查了这个因素。然而，其他一些检查者报告说，单体的这种转变是由细胞中 LT-B 二聚体的形成引起的。因此，我们检测是否有其他因素参与了LT的内吞作用，从而制备了阻断LT-B内吞作用的抗体。用细胞裂解物免疫一些兔子，该细胞裂解物是由缺陷GM1神经节苷脂的NCTC 2071制备的，我们可以制备抗LT-B的抗体。 -血清部分阻断LT对CHO细胞的内化。然后，我们现在正在研究对该抗血清产生反应的因素。此外，当我们收到这笔资助时，我们报告了以下数据：1。除质粒外，LT也编码在染色体中。2． A亚基Arg 192附近缺损突变位点对VZV、KKLH、卵清蛋白、BGG、麻疹病毒有较强的佐剂作用。 3． A 亚基 Arg 192 附近的切口位点不参与 LT 的某些生物活性。