Plisov S Iu, Merkulova T I, Baranova L V, Kumarev V P, Merkulov V M, Sokolenko A A, Kaĭkina I I, Salganik R I
Mol Biol (Mosk). 1990 Jul-Aug;24(4):1109-16.
Interaction of highly purified glucocorticoid receptor complex (GIRC) with synthetic DNA-fragment of mouse metallotionein 1 gene promoter from -209 to -252 b.p. (MTwt) was investigated. By means of nitrocellulose filter binding assay this fragment was shown to contain specific GIRC-binding site. In order to analyse the fine structure of the site, two variants of this DNA-fragment were synthesized and used in gel retardation assay. GIRC specific binding was shown to retain throughout interaction with the fragment in which all base pairs in the surroundings of generally accepted GIRC-binding site consensus G--ACA---TGTTCT C--TGT---ACAAGA were substituted by means of transitions, but it was weaker than the GIRC-binding with MTwt, where the mentioned consensus was situated in the natural surroundings. Complete loss of the GIRC-binding ability was observed when five CG pairs were substituted by AT ones. Two of the CG pairs belonged to the mentioned consensus. Comparison of the data obtained with results of computer analysis allows to consider the consensus as a "core" of GIRC-binding site, flanked with additional elements, interacting with GIRC.
研究了高度纯化的糖皮质激素受体复合物（GIRC）与小鼠金属硫蛋白1基因启动子-209至-252碱基对（MTwt）的合成DNA片段之间的相互作用。通过硝酸纤维素滤膜结合试验表明，该片段含有特异性GIRC结合位点。为了分析该位点的精细结构，合成了该DNA片段的两个变体并用于凝胶阻滞试验。结果显示，在与片段的整个相互作用过程中，GIRC特异性结合得以保留，其中在普遍认可的GIRC结合位点共有序列G--ACA---TGTTCT C--TGT---ACAAGA周围的所有碱基对通过转换进行了替换，但它比与MTwt的GIRC结合弱，在MTwt中上述共有序列位于自然环境中。当五个CG对被AT对替换时，观察到GIRC结合能力完全丧失。其中两个CG对属于上述共有序列。将所得数据与计算机分析结果进行比较，可以认为该共有序列是GIRC结合位点的“核心”，两侧是与GIRC相互作用的其他元件。
