Gschwedt M, Hamilton T H
Biochem J. 1972 Jul;128(3):611-6. doi: 10.1042/bj1280611.
Experiments performed with a cell-free system in tris-EDTA buffer, pH 7.4, indicate that the high-speed supernatant fraction of the rat uterus contains all the factors necessary to transform the 8S cytoplasmic oestradiol-receptor complex to the nuclear complex. The transformation is temperature-dependent. This nuclear complex was extracted in the form of a 5S particle with 0.4m-KCl from sediments of either uterine or heart nuclei that had been incubated together with the cytoplasmic soluble fraction of the uterus at 2 degrees C for 30min. This complex can also be obtained similarly from the soluble fraction of the uterus, incubated in the absence of nuclei. Previous warming of the soluble fraction to 37 degrees C for 7min was necessary for the successful extraction of the nuclear particle under these conditions of incubation. After an incubation of the transformed complex with the nuclear sediment at 37 degrees C for 7min, the 5S complex was extractable from the uterine nuclear sediment but not from the heart nuclear sediment, which may indicate the tissue specificity of the nuclear acceptor sites for the transformed complex. The extracted uterine nuclear complex sediments in the 5S region, but whether it is the native complex or a subunit or other part of the native complex resulting from the extraction with salt is unknown.
在pH 7.4的三羟甲基氨基甲烷-乙二胺四乙酸(tris-EDTA)缓冲液中使用无细胞系统进行的实验表明,大鼠子宫的高速上清液部分含有将8S细胞质雌二醇受体复合物转化为核复合物所需的所有因子。这种转化是温度依赖性的。该核复合物以5S颗粒的形式从子宫或心脏细胞核的沉淀物中用0.4m氯化钾提取,这些细胞核在2℃下与子宫细胞质可溶性部分一起孵育30分钟。这种复合物也可以类似地从子宫的可溶性部分中获得,在没有细胞核的情况下进行孵育。在这些孵育条件下,为了成功提取核颗粒,有必要将可溶性部分预先在37℃下加热7分钟。将转化后的复合物与核沉淀物在37℃下孵育7分钟后,5S复合物可从子宫核沉淀物中提取,但不能从心脏核沉淀物中提取,这可能表明转化复合物的核受体位点具有组织特异性。提取的子宫核复合物在5S区域沉淀,但它是天然复合物、亚基还是盐提取产生的天然复合物的其他部分尚不清楚。