Hydrogen peroxide stabilizes the steroid-binding state of rat liver glucocorticoid receptors by promoting disulfide bond formation.
Hydrogen peroxide and diamide inactivate the steroid-binding capacity of unoccupied glucocorticoid receptors in rat liver cytosol at 0 degrees C, and steroid-binding capacity is reactivated with dithiothreitol. Treatment of cytosol with peroxide or sodium molybdate, but not diamide, inhibits the irreversible inactivation (i.e., inactivation not reversed by dithiothreitol) of steroid-binding capacity that occurs when cytosol is incubated at 25 degrees C. Pretreatment of cytosol with the thiol derivatizing agent methyl methanethiosulfonate at 0 degrees C prevents the ability of peroxide, but not molybdate, to stabilize binding capacity at 25 degrees C. As derivatization of thiol groups prevents peroxide stabilization of steroid-binding capacity and as treatment with dithiothreitol reverses the effect, we propose that peroxide acts by promoting the formation of new disulfide linkages. The receptor in our rat liver cytosol preparations is present as three major degradation products of Mr 40,000, 52,000, and 72,000 in addition to the Mr 94,000 intact receptor. Like the intact receptor, these three forms exist in the presence of molybdate as an 8-9S complex, they bind glucocorticoid in a specific manner, and they copurify with the intact Mr 94,000 receptor on sequential phosphocellulose and DNA-cellulose chromatography. Despite the existence of receptor cleavage products, it is clear that peroxide does not stabilize steroid-binding capacity by inhibiting receptor cleavage.
Bresnick
E H
EH
Department of Pharmacology, University of Michigan Medical School, Ann Arbor 48109.
Sanchez
E R
ER
Harrison
R W
RW
Pratt
W B
WB
D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D003600:Cytosol / Q000378:metabolism; D003907:Dexamethasone / Q000031:analogs & derivatives / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008982:Molybdenum / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D011965:Receptors, Glucocorticoid / Q000187:drug effects / Q000378:metabolism*; D014222:Triamcinolone Acetonide / Q000378:metabolism
D000345:Affinity Labels; D004220:Disulfides; D011965:Receptors, Glucocorticoid; C044659:molybdate; D003907:Dexamethasone; D008982:Molybdenum; D006861:Hydrogen Peroxide; D014222:Triamcinolone Acetonide; C031710:dexamethasone 21-methanesulfonate