Steroid substrate-induced epimerase mechanism in the active site of the human 11β-hydroxysteroid dehydrogenase type 1

Abstract

Cytochrome P4507B1 7[alpha]-hydroxylates dehydroepiandrosterone (DHEA), epiandrosterone (EpiA) and 5[alpha]-androstane-3[beta],17[beta]-diol (Adiol). 11[beta]-Hydroxysteroid dehydrogenase type 1 (11[beta]-HSD1) interconverts 7[alpha]- and 7[beta]- forms. Whether the inter-conversion proceeds through oxido-reductive steps or epimerase activity is investigated. Experiments using ^3^H-labeled 7[beta]-hydroxy-DHEA, 7[beta]-hydroxy-EpiA and 7[beta]-hydroxy-Adiol show the ^3^H-label to accumulate in 7-oxo-DHEA trap but neither in 7-oxo-EpiA nor 7-oxo-Adiol traps. Computed models of 7-oxygenated steroids dock in the active site of 11[beta]-HSD1 either in a flipped or turned form relative to cortisone and cortisol. 7-Oxo-steroid reduction in 7[alpha]- or 7[beta]-hydroxylated derivatives results from either turned or flipped forms. 11[beta]-HSD1 incubation in H~2~^18^O medium with each 7-hydroxysteroid did not incorporate ^18^O in 7-hydroxylated derivatives of EpiA and Adiol independently of the cofactor used. Thus oxido-reductive steps apply for the interconversion of 7[alpha]- and 7[beta]-hydroxy-DHEA through 7-oxo-DHEA. Epimerisation may proceed on the 7-hydroxylated derivatives of EpiA and Adiol through a mechanism involving the cofactor and Ser170.