Howard K J, Distelhorst C W
Department of Medicine, Case Western Reserve University School of Medicine, Cleveland, Ohio.
Biochem Biophys Res Commun. 1988 Mar 30;151(3):1226-32. doi: 10.1016/s0006-291x(88)80497-2.
Glucocorticoid receptors in the IM-9 human lymphoblastoid cell line were affinity labeled with [3H]dexamethasone 21-mesylate and activated to a DNA-binding form by filtration through a Bio-Gel A-1.5m column. The 90 kDa heat shock protein, HSP90, was identified by labeling IM-9 cells with 35S-methionine at both 37 degrees C and 42 degrees C and purified to near homogeneity by sequential chromatography through DE52 and hydroxyapatite. Addition of purified HSP90 to activated, affinity labeled glucocorticoid receptors in a molecular ratio of 16 to 1 inhibited the binding of the receptors to DNA-cellulose. HSP90 did not affect the binding of other proteins to DNA-cellulose, indicating that the inhibitory effect of HSP90 was specific for the glucocorticoid receptor. These results suggest that HSP90 may associate with the glucocorticoid receptor, masking its DNA-binding site and thereby inhibiting receptor interaction with DNA.
用[3H]地塞米松21-甲磺酸盐对人IM-9淋巴母细胞系中的糖皮质激素受体进行亲和标记，并通过Bio-Gel A-1.5m柱过滤将其激活为DNA结合形式。通过在37℃和42℃下用35S-甲硫氨酸标记IM-9细胞来鉴定90 kDa热休克蛋白HSP90，并通过DE52和羟基磷灰石顺序色谱法将其纯化至接近均一。以16比1的分子比将纯化的HSP90添加到活化的、亲和标记的糖皮质激素受体中，可抑制受体与DNA纤维素的结合。HSP90不影响其他蛋白质与DNA纤维素的结合，表明HSP90的抑制作用对糖皮质激素受体具有特异性。这些结果表明，HSP90可能与糖皮质激素受体结合，掩盖其DNA结合位点，从而抑制受体与DNA的相互作用。
