Glucocorticoid receptor in human embryo fibroblasts. I. Kinetic and physicochemical properties.
The kinetics of dexamethasone binding to L 809 E cell line cytosol have been investigated by means of the protamine sulfate precipitation assay. The KDeq for dexamethasone was 1.1--3.3 nM. Binding was specific for glucocorticoids. The mean association rate constant (k+1) was 8.5 x 10(5) M-1 x min-1 and the dissociation rate constant was 4.6 x 10(-5) min-1 at 0 degrees C. The concentration of binding sites was 0.3 pmol/mg of cytosol protein. Binding kinetics were compatible with a model of positive cooperativity. The receptor sedimented at 7.5--9 S in glycerol gradients. By a combination of calibrated ultracentrifugation and polyacrylamide gel electrophoresis, a Stokes radius of 8.5 nm, a molecular weight of 268 000 daltons and a frictional ratio of 1.8 were determined in low ionic strength conditions. When the cells were incubated with 10 nM [3H]dexamethasone for 1 h, a more than 90% depletion of cytosol receptor and an equivalent accumulation of nuclear dexamethasone--receptor complexes was observed.
D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D003907:Dexamethasone / Q000378:metabolism; D004622:Embryo, Mammalian; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D011247:Pregnancy; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D011987:Receptors, Steroid / Q000378:metabolism*
D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; D003907:Dexamethasone