ZK98299—A new antiprogesterone: Biochemical characterization of steroid binding parameters in the calf uterine cytosol

doi:10.1016/0003-9861(92)90083-9

Archives of Biochemistry and Biophysics

Volume 292, Issue 1, January 1992, Pages 303-310

https://doi.org/10.1016/0003-9861(92)90083-9 Get rights and content

Abstract

We have examined steroid binding characteristics of a newly synthesized antisteroid, ZK98299 [onapristone, 11β-(4-dimethylaminophenyl)-17α-hydroxy-17β-(3-hydroxypropyl)-13α-methyl-4,9-gonadien-3-one], in the calf uterus cytosol and compared the nature of this interaction with the binding of progesterone receptor (PR) agonist R5020 [promegestone, 17,21-dimethylpregna-4,9-diene-3,20-dione]. In the freshly prepared cytosol, [³H]ZK98299 interacted specifically with a macromolecule: the binding was abolished in the presence of excess progestins (R5020 and progesterone) and the antiprogesterone ZK98299. The high affinity (K_d = 2.5 nM) interaction between [³H]ZK98299 and PR was temperature- and time-dependent, reaching an optimum by 2–3 h at 0 °C, and was facilitated by 20 mm Na₂MoO₄. Under nontransforming conditions, [³H]ZK98299-receptor complexes sedimented as 8 S species in 8–30% linear glycerol gradients. Upon salt or thermal transformation, there was a loss of the 8 S form, with only a small fraction of total complexes (5–7%) binding to DNA-cellulose. In contrast, transformed [³H]R5020-receptor complexes exhibited a greater extent of binding (25–55%) to DNA-cellulose. [³H]ZK98299-receptor complexes could be resolved into two ionic species over DEAE-Sephacel following incubation of the complexes at 0 or 23 °C. [³H]ZK98299 binding was sensitive to sulfhydryl group modification as β-mercaptoethanol increased the extent of steroid binding. Although treatment with iodoacetamide (IA) abolished [³H]R5020 binding, there was a significant (nearly twofold) increase in the [³H]ZK98299 binding. The results of this study point to similarities and differences between the steroid binding properties of the uterine PR occupied by R5020 and ZK98299: both steroids appear to bind the same 8 S receptor but exhibit differential DNA binding and sensitivity to IA. The reported antagonist properties of ZK98299 may, therefore, be explained on the basis of a distinct receptor conformation induced by the antisteroid.

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^☆: Supported by NIH Grant DK-20893.

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ZK98299—A new antiprogesterone: Biochemical characterization of steroid binding parameters in the calf uterine cytosol☆

Abstract

Biochim. Biophys. Acta

Biochim. Biophys. Acta

J. Biol. Chem

Steroids

J. Steroid Biochem

J. Steroid Biochem

Eur. J. Cancer Clin. Oncol

J. Biol. Chem

Biochem. Biophys. Res. Commun

J. Biol. Chem

J. Biol. Chem

J. Biol. Chem

J. Biol. Chem

Biochem. Biophys. Res. Commun

J. Biol. Chem

J. Biol. Chem

Biochim. Biophys. Acta

Biochim. Biophys. Acta

Annu. Rev. Genet

Science

Sci. Am

N. Engl. J. Med

N. Engl. J. Med