Interaction of rat liver glucocorticoid receptor with adenosine 5'-triphosphate. Characterization of interaction by use of ATP-sepharose affinity chromatography

Abstract

An interaction between rat liver glucocorticoid—receptor complex and immobilized ATP was identified. Rat liver cytosol preparations were incubated with [3H]triamcinolone acetonide for 4 h at 4 degrees C and partially purified by precipitation with (NH4)2SO4 before use. The resulting glucocorticoid—receptor complex could be selectively adsorbed on to columns of ATP—Sepharose. The freshly prepared cytosol [3H]triamcinolone acetonide—receptor complex had very little affinity for binding to the ATP—Sepharose column, but acquired this ability on temperature- or salt-activation. The presence of 10 mM-sodium molybdate during this salt- or temperature-dependent activation blocked the binding of the receptor complex to ATP—Sepharose. The interaction is reversible, since it can be disrupted by high-salt conditions. A competitive binding assay, using free nucleotides in samples to be chromatographed, revealed a preferential interaction between ATP and the glucocorticoid—receptor complex. Buffer containing ATP was also used to elute the glucocorticoid—receptor complex from ATP—Sepharose columns successfully. When ATP was added to the preparations containing [3H]triamcinolone acetonide—receptor complexes, the steroid specificity or sedimentation properties of the complex remained unaltered. Our results demonstrate an interaction between rat liver glucocorticoid—receptor complex and immobilized ATP and suggest a role of this nucleotide in receptor function.