title: Absence of intramolecular disulfides in the structure and function of native rat glucocorticoid receptors.
abstract: The presence of intramolecular disulfides in different functional states of the native glucocorticoid receptor in the absence of added oxidants has been examined on nonreducing SDS-polyacrylamide gels. Possible disulfides were trapped by the reaction of thiols in crude receptor solutions with methyl methanethiolsulfonate or iodoacetamide. The presence of diffuse bands at lower molecular weights than either the fully reduced or the thiol-blocked species for both the intact 98-kDa receptor and the 42-kDa chymotryptic fragment was diagnostic of an intramolecular disulfide(s) that had undergone thiol-disulfide rearrangements. However, both the rearrangements and the formation of intramolecular disulfides were found to occur only with denatured receptors during gel analysis. It appears that the thiols normally complexed with zinc in the zinc fingers may be recruited for disulfide bond formation. Finally, even when a documented intramolecular disulfide was formed in solutions of the native protein, thiol-disulfide rearrangements did not occur. The tertiary structure of the receptor is thus constituted in a manner that not only limits the formation of disulfides but also prevents the usually facile rearrangements of disulfide bond-containing structures to receptor forms that may have greatly reduced activity. Therefore, although intramolecular disulfide bonds may be of transitory importance, the structural or functional changes of native glucocorticoid receptors that are associated with steroid binding, activation, and dissociation of heat shock protein 90 neither involve nor require the formation or reduction of stable intramolecular disulfides.
authors.affiliation: Laboratory of Molecular and Cellular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: J
authors.initials: J
authors.lastname: Opoku
authors.forename: S S
authors.initials: SS
authors.lastname: Simons
mesh_terms: D000818:Animals; D004220:Disulfides / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D011487:Protein Conformation; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000737:chemistry* / Q000502:physiology; D013329:Structure-Activity Relationship; D013438:Sulfhydryl Compounds / Q000737:chemistry
chemical_list: D004220:Disulfides; D011965:Receptors, Glucocorticoid; D013438:Sulfhydryl Compounds