Stability and transformation of the glucocorticoid receptor under acidic conditions.
The [3H]triamcinolone acetonide ([3H]TA)-binding ability of the rat liver glucocorticoid receptor (GR) was investigated under acidic conditions, ranging from pH 2 to 7.3. Both in the presence and absence of 10 mM molybdate, the [3H]TA-binding ability decreased below pH 6.5 and was almost completely lost below pH 5, pH 5.9 +/- 0.1 giving 50% [3H]TA-binding. The binding ability was recovered when the pH of the cytosol was reversed to 7.3 or the precipitate obtained on acidification was dissolved in a buffer of pH 7.3. Moreover, in the absence of molybdate, the [3H]TA-GR complexes formed at pH 7.3 remained unchanged until pH 5. Then they decreased, pH 3.9 +/- 0.1 giving 50% binding, and completely disappeared at pH 3. [3H]TA-binding activity recovered from the precipitate also decreased in a similar pH region (a 50% decrease in binding being observed at pH 4.2 +/- 0.04). These results suggest that rat liver GR is rather resistant under acidic conditions and that it exists in a peculiar state below pH 5.9 to approximately 4 as to its ligand binding property: unoccupied GR has no [3H]TA-binding ability but [3H]TA-GR complexes once formed at neutral pH do not dissociate. [3H]TA-GR complexes recovered from the precipitate at pH 5 had a Stokes radius of 7.5 nm, little DNA-cellulose-binding ability and sedimented at 8.6S on glycerol gradient centrifugation, indicating that the receptor existed in a nontransformed state. In addition, both occupied and unoccupied GR were transformed at about pH 4, their being 50% transformation. This transformation was accompanied by irreversible denaturation of the receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
Nemoto
T
T
Department of Biochemistry, Iwate Medical University School of Dentistry.
Ohara-Nemoto
Y
Y
Ota
M
M
D000818:Animals; D001711:Biotransformation; D002499:Centrifugation, Density Gradient; D003600:Cytosol / Q000032:analysis; D004355:Drug Stability; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000032:analysis; D008297:Male; D008982:Molybdenum / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D011965:Receptors, Glucocorticoid / Q000187:drug effects / Q000378:metabolism*; D014221:Triamcinolone / Q000378:metabolism
D011965:Receptors, Glucocorticoid; C044659:molybdate; D014221:Triamcinolone; D008982:Molybdenum