Zakula Z, Moudgil V K
Department of Biological Sciences, Oakland University, Rochester, MI 48309-4401.
Biochim Biophys Acta. 1991 Apr 17;1092(2):188-95. doi: 10.1016/0167-4889(91)90156-r.
Steroid receptor antagonists are important biochemical probes for understanding the mode of steroid hormone action. We have studied the interaction between rat liver glucocorticoid receptor and a newly synthesized antisteroid ZK98299 (13-antigestagen; [11-beta-(4-dimethylaminophenyl)-17a-hydroxy-17 beta-(3- hydroxypropyl)-13 alpha-methyl-4,9-gonadien-3-one]). Glucocorticoid receptor from freshly prepared hepatic cytosol bound [3H]ZK98299 with affinity approximately equal to that of [3H]triamcinolone acetonide. The binding of both steroids reached a maximum at 4 h at 0 degrees C. Both ligands were able to compete for the steroid binding site but progesterone, estradiol and dihydrotestosterone (DHT) failed to compete for the [3H]ZK98299 and [3H]triamcinolone acetonide binding. While [3H]ZK98299 binding to glucocorticoid receptor could occur in the presence of iodoacetamide and N-ethylmaleimide (NEM), [3H]triamcinolone acetonide binding capacity was completely abolished following such treatments. The [3H]ZK98299-receptor complexes sedimented as 9 S and 4 S molecules under control (4 degrees C) and receptor transforming (23 degrees C) conditions, and exhibited a faster rate of dissociation at 23 degrees C when compared with [3H]triamcinolone acetonide-receptor complexes. These results indicate that ZK98299 interacts with hepatic glucocorticoid receptor. The differential effects of iodoacetamide and NEM on the interaction of glucocorticoid receptor with ZK98299 and triamcinolone acetonide, and the faster rate of dissociation of [3H]ZK98299-receptor complexes suggest that treatment with these agents (NEM and iodoacetamide) results in distinct conformational changes in glucocorticoid receptor structure with respect to triamcinolone acetonide and ZK98299 binding. Alternatively, ZK98299 may be interacting with a site which is distinct from one which accepts triamcinolone acetonide.
类固醇受体拮抗剂是理解类固醇激素作用模式的重要生化探针。我们研究了大鼠肝脏糖皮质激素受体与新合成的抗类固醇ZK98299（13-抗孕激素；[11-β-(4-二甲基氨基苯基)-17α-羟基-17β-(3-羟丙基)-13α-甲基-4,9-孕二烯-3-酮]）之间的相互作用。新鲜制备的肝细胞溶胶中的糖皮质激素受体与[3H]ZK98299结合的亲和力约等于与[3H]曲安奈德的亲和力。两种类固醇的结合在0℃下4小时达到最大值。两种配体都能够竞争类固醇结合位点，但孕酮、雌二醇和二氢睾酮（DHT）未能竞争[3H]ZK98299和[3H]曲安奈德的结合。虽然[3H]ZK98299与糖皮质激素受体的结合可以在碘乙酰胺和N-乙基马来酰亚胺（NEM）存在的情况下发生，但经过此类处理后，[3H]曲安奈德的结合能力完全丧失。在对照（4℃）和受体转化（23℃）条件下，[3H]ZK98299-受体复合物以9S和4S分子形式沉降，与[3H]曲安奈德-受体复合物相比，在23℃时表现出更快的解离速率。这些结果表明ZK98299与肝脏糖皮质激素受体相互作用。碘乙酰胺和NEM对糖皮质激素受体与ZK98299和曲安奈德相互作用的不同影响，以及[3H]ZK98299-受体复合物更快的解离速率表明，用这些试剂（NEM和碘乙酰胺）处理会导致糖皮质激素受体结构相对于曲安奈德和ZK98299结合发生明显的构象变化。或者，ZK98299可能与一个不同于接受曲安奈德的位点相互作用。
