Publications

• Structure-function aspects and inhibitor design of type 5 17beta-hydroxysteroid dehydrogenase (AKR1C3).

  Penning T.M., Burczynski M.E., Jez J.M., Lin H.-K., Ma H., Moore M., Ratnam K., Palackal N.

  17beta-Hydroxysteroid dehydrogenase (17beta-HSD) type 5 has been cloned from human prostate and is identical to type 2 3alpha-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions ... >> More

  17beta-Hydroxysteroid dehydrogenase (17beta-HSD) type 5 has been cloned from human prostate and is identical to type 2 3alpha-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3alpha-, 17beta- and 20alpha-hydroxysteroid oxidase. The enzyme will reduce 5alpha-DHT, Delta(4)-androstene-3,17-dione, estrone and progesterone to produce 3alpha-androstanediol, testosterone, 17beta-estradiol and 20alpha-hydroxprogesterone, respectively. It will also oxidize 3alpha-androstanediol, testosterone, 17beta-estradiol and 20alpha-hydroxyprogesterone to produce 5alpha-androstane-3,17-dione, Delta(4)-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of k(cat)/K(m) for these reactions indicates that as a reductase it prefers 5alpha-dihydrotestosterone and 5alpha-androstane-3,17-dione as substrates to Delta(4)-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17beta-estradiol; convert Delta(4)-androstene-3,17-dione to testosterone (which can be aromatized to 17beta-estradiol); and it will reduce progesterone to its inactive metabolite 20alpha-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3alpha-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is k(chem), and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3alpha, 17beta or 20alpha-spiro-oxiranes may act as mechanism-based inactivators. << Less

  Mol. Cell. Endocrinol. 171:137-149(2001) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Expression and characterization of recombinant type 2 3 alpha-hydroxysteroid dehydrogenase (HSD) from human prostate: demonstration of bifunctional 3 alpha/17 beta-HSD activity and cellular distribution.

  Lin H.-K., Jez J.M., Schlegel B.P., Peehl D.M., Pachter J.A., Penning T.M.

  In androgen target tissues, 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) may regulate occupancy of the androgen receptor (AR) by catalyzing the interconversion of 5alpha-dihydrotestosterone (5alpha-DHT) (a potent androgen) and 3alpha-androstanediol (a weak androgen). In this study, a 3alpha-HS ... >> More

  In androgen target tissues, 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) may regulate occupancy of the androgen receptor (AR) by catalyzing the interconversion of 5alpha-dihydrotestosterone (5alpha-DHT) (a potent androgen) and 3alpha-androstanediol (a weak androgen). In this study, a 3alpha-HSD cDNA (1170 bp) was isolated from a human prostate cDNA library. The human prostatic 3alpha-HSD cDNA encodes a 323-amino acid protein with 69.9%, 84.1%, 99.4%, and 87.9% sequence identity to rat liver 3alpha-HSD and human type 1, type 2, and type 3 3alpha-HSDs, respectively, and is a member of the aldo-keto reductase superfamily. The close homology with human type 2 3alpha-HSD suggests that it is either identical to this enzyme or a structural allele. Surprisingly, when the recombinant protein was expressed and purified from Escherichia coli, the enzyme did not oxidize androsterone when measured spectrophotometrically, an activity previously assigned to recombinant type 2 3alpha-HSD using this assay. Complete kinetic characterization of the purified protein using spectrophotometric, fluorometric, and radiometric assays showed that the catalytic efficiency favored 3alpha-androstanediol oxidation over 5alpha-DHT reduction. Using [14C]-5alpha-DHT as substrate, TLC analysis confirmed that the reaction product was [14C]-3alpha-androstanediol. However, in the reverse reaction, [3H]-3alpha-androstanediol was oxidized first to [3H]-androsterone and then to [3H]-androstanedione, revealing that the expressed protein possessed both 3alpha- and 17beta-HSD activities. The 17beta-HSD activity accounted for the higher catalytic efficiency observed with 3alpha-androstanediol. These findings indicate that, in the prostate, type 2 3alpha-HSD does not interconvert 5alpha-DHT and 3alpha-androstanediol but inactivates 5alpha-DHT through its 3-ketosteroid reductase activity. Levels of 3alpha-HSD mRNA were measured in primary cultures of human prostatic cells and were higher in epithelial cells than stromal cells. In addition, elevated levels of 3alpha-HSD mRNA were observed in epithelial cells derived from benign prostatic hyperplasia and prostate carcinoma tissues. Expression of 3alpha-HSD was not prostate specific, since high levels of mRNA were also found in liver, small intestine, colon, lung, and kidney. This study is the first complete characterization of recombinant type 2 3alpha-HSD demonstrating dual activity and cellular distribution in the human prostate. << Less

  Mol. Endocrinol. 11:1971-1984(1997) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.