Steroidumwandelnde Enzyme aus Mikroorganismen X. Anreicherung einer 4‐En‐3‐oxosteroid‐5α‐Reduktase aus Mycobacterium smegmatis sowie Abtrennung und Anreicherung des Apoenzyms mit Hilfe der Affinitätschromatographie

The 4‐en‐3‐oxosteroid‐5α‐reductase from Mycobacterium smegmatis was bound biospecifically on the affinant containing an immobilized testosterone ligand. The enzyme obtained by elution with ethylene glycol and urea in a 32 fold purity has a S. A. of 8.73 × 10 −3 μM androstenedione min −1 mg −1 . The coenzyme (FAD) could be separated from the immobilized enzyme substrate complex on the affinity matrix, in the presence of (NH 4 ) 2 SO 4 at pH 3.0. After elution of the apoenzyme 97% of the initial enzyme activity was obtained by incubation with FAD. The reactivated enzyme results in a 40 fold enrichment.