Structure-function aspects and inhibitor design of type 5 17²-hydroxysteroid dehydrogenase (AKR1C3)

17β-Hydroxysteroid dehydrogenase (17β-HSD) type 5 has been cloned from human prostate and is identical to type 2 3α-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 3α-, 17β- and 20α-hydroxysteroid oxidase. The enzyme will reduce 5α-DHT, Δ⁴-androstene-3, 17-dione, estrone and progesterone to produce 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxprogesterone, respectively. It will also oxidize 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxyprogesterone to produce 5α-androstane-3, 17-dione, Δ⁴-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 5α-dihydrotestosterone and 5α-androstane-3, 17-dione as substrates to Δ⁴-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 17β-estradiol; convert Δ⁴-androstene-3, 17-dione to testosterone (which can be aromatized to 17β-estradiol); and it will reduce progesterone to its inactive metabolite 20α-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3α-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 3α, 17β or 20α-spiro-oxiranes may act as mechanism-based inactivators.