HUMAN 3-BETA-HYDROXYSTEROID DEHYDROGENASE DELTA-5-]4-ISOMERASE FROM PLACENTA - EXPRESSION IN NONSTEROIDOGENIC CELLS OF A PROTEIN THAT CATALYZES THE DEHYDROGENATION ISOMERIZATION OF C21 AND C19 STEROIDS

The isolation, cloning, and expression of a cDNA insert complementary to mRNA encoding human 3β-hydroxysteroid dehydrogenase/Δ5→4isomerase is reported. The insert contains an open reading frame encoding a protein of 372 amino acids, the initial 29 amino acids corresponding to the N-terminal sequence identified from the purified human placental microsomal enzyme. The cDNA was inserted into a modified pCMV vector and expressed in COS-1 monkey kidney tumor cells. The expressed protein was similar in size to human placental microsomal 3β-hydroxysteroid dehydrogenase/ Δ5→4isomerase, as detected by immunoblot analysis, and catalyzed the conversion of 17α-hydroxypregnenolone to 17α-hydroxypro-gesterone, pregnenolone to progesterone, and dehydroepiandro3-terone to androstenedione. Transfected COS cell homogenates, supplemented with NAD+very efficiently oxidized 5α-andros-tan-3β, 17β-diol to 5α-dihydrotestosterone and, upon addition of NADH, reduced 5α-dihydrotestosterone to 5α-androstan-3β, 17β-diol. Thus, the dehydrogenation/isomerization steps of steroid biosynthesis can be catalyzed by a single polypeptide chain, which can metabolize all of the major physiological substrates. © 1990 by The Endocrine Society.