Differential effects of chelating agents on cytosolic glucocorticoid receptor stability and nuclear binding in vitro

The effect of several metal chelators (EDTA, EGTA, and 1,10 phenanthroline) on rat liver glucocorticoid receptor properties in vitro was investigated. At 4 degrees C 10 mM EDTA (unlike 10 mM EGTA and 10 mM 1,10 phenanthroline) had a significant stabilizing effect on unbound hepatic glucocorticoid receptors. At higher temperature (25 degrees C) 10 mM EGTA appeared to act as a chemical stabilizer of unbound receptors. 1,10 Phenanthroline had no stabilizing effect at either temperature. Scatchard analysis indicated that the alteration in receptor binding after incubation at 4 and 25 degrees C in the presence and absence of chelating agents was due to a change in the number of steroid binding sites rather than perturbation of receptor affinity. Unlike results obtained with unbound receptors, all three chelating agents appeared to enhance prebound glucocorticoid-receptor complex inactivation. Interestingly these chelating reagents also significantly altered glucocorticoid-receptor complex binding to isolated nuclei.