title: Human 3 beta-hydroxysteroid dehydrogenase/delta 5----4isomerase from placenta: expression in nonsteroidogenic cells of a protein that catalyzes the dehydrogenation/isomerization of C21 and C19 steroids.
abstract: The isolation, cloning, and expression of a cDNA insert complementary to mRNA encoding human 3 beta-hydroxysteroid dehydrogenase/delta 5----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 3 beta-hydroxysteroid dehydrogenase/delta 5----4isomerase, as detected by immunoblot analysis, and catalyzed the conversion of 17 alpha-hydroxypregnenolone to 17 alpha-hydroxyprogesterone, pregnenolone to progesterone, and dehydroepiandrosterone to androstenedione. Transfected COS cell homogenates, supplemented with NAD+, very efficiently oxidized 5 alpha-androstan-3 beta,17 beta-diol to 5 alpha-dihydrotestosterone and, upon addition of NADH, reduced 5 alpha-dihydrotestosterone to 5 alpha-androstan-3 beta,17 beta-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.
authors: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas 75235-9051.
authors: M C
authors: MC
authors: Lorence
authors: B A
authors: BA
authors: Murry
authors: J M
authors: JM
authors: Trant
authors: J I
authors: JI
authors: Mason
mesh_terms: D006907:17-alpha-Hydroxypregnenolone / Q000378:metabolism; D019326:17-alpha-Hydroxyprogesterone; D015096:3-Hydroxysteroid Dehydrogenases / Q000235:genetics*; D000595:Amino Acid Sequence; D000735:Androstenedione / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D003687:Dehydroepiandrosterone / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D006908:Hydroxyprogesterones / Q000378:metabolism; D015151:Immunoblotting; D007535:Isomerases / Q000235:genetics*; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D009243:NAD / Q000494:pharmacology; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D011284:Pregnenolone / Q000378:metabolism; D011374:Progesterone / Q000378:metabolism; D011375:Progesterone Reductase / Q000235:genetics*; D013251:Steroid Isomerases / Q000235:genetics*; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: C021425:3 beta-hydroxysteroid oxidoreductase-delta(5) 3-ketosteroid isomerase; D006908:Hydroxyprogesterones; D009097:Multienzyme Complexes; D009243:NAD; D006907:17-alpha-Hydroxypregnenolone; D000735:Androstenedione; D003687:Dehydroepiandrosterone; D011374:Progesterone; D019326:17-alpha-Hydroxyprogesterone; D011284:Pregnenolone; D004247:DNA; D015096:3-Hydroxysteroid Dehydrogenases; D011375:Progesterone Reductase; D007535:Isomerases; D013251:Steroid Isomerases