Preparation of the apoenzyme and holoenzyme forms of human 17 beta-hydroxysteroid dehydrogenase

The apoenzyme and holoenzyme (NADP+ complex) of human placental 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) were prepared from affinity chromatography using various elutions by column liquid chromatography. The apoenzyme was obtained using NAD+ elution in a Blue-Sepharose column, followed by NAD+ separation on a Phenyl-Superose hydrophobic-interaction or a Mono Q anion-exchange column. The 17 beta-HSD-NADP+ complex was prepared using NADP+ elution in a Blue-Sepharose column. The two forms have different A280/A260 ratios and are suitable for further study of enzyme-cofactor interactions.