Inhibition of 3α/β,20β-Hydroxysteroid Dehydrogenase by Dexamethasone, Glycyrrhetinic Acid and Spironolactone Is Attenuated by Deletion of 12 Carboxyl-Terminal Residues

Abstract

We constructed a pig 3α/β,20β-hydroxysteroid dehydrogenase (3α/β,20β-HSD) mutant, which lacks 12 carboxyl-terminal amino acids residues. Enzyme activity studies indicated that the deleted amino acids have a role in steroid metabolism and may assist in substrate binding in wild-type 3α/β,20β-HSD. Furthermore, substrate binding likely induces a conformational change allowing the 12 carboxyl-terminal amino acids interact with the steroid substrate [Nakajin S. et al., Biochim. Biophys. Acta, 1550, 175—182 (2001)]. In this paper, we clarified that although pig 3α/β,20β-HSD is potently inhibited by dexamethasone, glycyrrhetinic acid and spironolactone, this inhibition is remarkably attenuated by deleting the 12 carboxyl-terminal residues. The inhibition constant (K_i) of pig 3α/β,20β-HSD for dexamethasone increased 115-fold. These observations also indicate that these amino acid residues interact with steroid substrates or steroid inhibitors and have an important role in substrate or inhibitor binding to the active site.