Rees A M, Bell P A
Biochim Biophys Acta. 1975 Nov 10;411(1):121-32. doi: 10.1016/0304-4165(75)90291-3.
The glucocorticoid receptor protein present in the high-speed supernant fraction of rat thymus tissue is extremely unstable, having a half-life of about 2 h at 4 degrees C. It was found that the decline in steroid-binding capacity could be slowed, though not arrested completely, by the addition of sulphydryl-protecting agents such as 2-mercaptoethanol or dithiothreitol, and by EDTA. The inactivation was also partly reversed by these agents. 0.5 mM N-ethylmaleimideor p-chloromercuriphenylsulphonic acid inactivated the recptor at 4 degrees C, but the presence of bound steroid protected the receptor against this inactivation. Bound steroid did not protect the receptor against the action of higher concentrations of these reagents. Treatment of intact thymus cells with 2,4-dinitrophenol resulted in a reduction in the steroid-binding capacity of the supernatant fraction derived from these cells. This effect of 2,4-dinitrophenol could not be reversed by the presence of dithiothreitol in the extraction buffer. It is concluded that the inactivation of the receptor in vitro is at least partly due to the oxidation of one or more sulphydryl groups necessary for steroid binding; the process of oxidation does not account for the reduction in steroid binding observed in intact thymus cells under conditions of energy deprivation.
大鼠胸腺组织高速上清液组分中存在的糖皮质激素受体蛋白极不稳定，在4℃时半衰期约为2小时。研究发现，添加巯基保护剂如2-巯基乙醇或二硫苏糖醇以及EDTA后，类固醇结合能力的下降虽不能完全阻止，但可减缓。这些试剂也能部分逆转失活。0.5 mM N-乙基马来酰亚胺或对氯汞苯磺酸在4℃时可使受体失活，但结合的类固醇可保护受体免受这种失活作用。结合的类固醇不能保护受体免受更高浓度这些试剂的作用。用2,4-二硝基苯酚处理完整的胸腺细胞会导致这些细胞来源的上清液组分中类固醇结合能力降低。提取缓冲液中存在二硫苏糖醇不能逆转2,4-二硝基苯酚的这种作用。结论是，体外受体失活至少部分是由于类固醇结合所需的一个或多个巯基被氧化；氧化过程不能解释在能量剥夺条件下完整胸腺细胞中观察到的类固醇结合减少现象。
