11 beta-hydroxysteroid dehydrogenases of the choriocarcinoma cell line JEG-3 and their inhibition by glycyrrhetinic acid and other natural substances.
Mineralocorticoid receptor (MR) selectivity for aldosterone is thought to be exerted by enzymes which inactivate competing glucocorticoids before they bind the receptor. Two different 11 beta-hydroxysteroid dehydrogenases (11 beta-HSD) have been described. 11 beta-HSD-1 is NADP(+)-dependent and has a Km in the micromolar range and bidirectional activity. 11 beta-HSD-2 is NAD(+)-dependent, has a Km in the nanomolar range, exhibits only oxidase activity, and colocalizes with the MR in the kidney, so is likely to serve as the gatekeeper for the MR. We have further characterized 11 beta-HSD activity in JEG-3 cells, a cell line derived from a human choriocarcinoma which was reported to have only the high affinity, NAD(+)-dependent 11 beta-HSD-2. We found that the Km for the conversion of corticosterone to 11-dehydrocorticosterone in intact cells and homogenates was about 16 nM. NAD(+)-dependent corticosterone conversion was equal in the nuclear and mitochondrial fractions and less, but significant, in the microsomal fraction. A high affinity, Km = 40 nM, NADP(+)-dependent enzyme was also found in homogenates. The subcellular distribution of this high affinity activity was greatest in the mitochondria, less in the nuclei, and even less, but still significant, in microsomes. Because of its cofactor dependency, high affinity, and different subcellular distribution, we suggest that this enzyme is neither the 11 beta-HSD-1 nor the 11 beta-HSD-2 and have named it 11 beta-HSD-3. Conversion of 11-dehydrocorticosterone to corticosterone did not occur in intact cells or in homogenates incubated with NADH or NADPH. Enzyme activity in intact cells was inhibited by glycyrrhetinic acid, carbenoxolone, progesterone, 5 beta-dihydroprogesterone, and 5 alpha-dihydroprogesterone, but not bile acids.
Gomez-Sanchez
E P
EP
Research Service, Harry S. Truman Memorial Veterans Hospital, Columbia, MO 65201, USA.
Cox
D
D
Foecking
M
M
Ganjam
V
V
Gomez-Sanchez
C E
CE
D042842:11-beta-Hydroxysteroid Dehydrogenases; D002822:Choriocarcinoma / Q000201:enzymology*; D003345:Corticosterone / Q000031:analogs & derivatives / Q000096:biosynthesis / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006034:Glycyrrhetinic Acid / Q000494:pharmacology*; D006801:Humans; D006913:Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D018161:Receptors, Mineralocorticoid / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D018161:Receptors, Mineralocorticoid; D009243:NAD; D009249:NADP; D006913:Hydroxysteroid Dehydrogenases; D042842:11-beta-Hydroxysteroid Dehydrogenases; C003552:11-dehydrocorticosterone; D006034:Glycyrrhetinic Acid; D003345:Corticosterone