Evidence for distinct isoforms of 11 beta-hydroxysteroid dehydrogenase in the ovine liver and kidney.
We have previously identified a unique 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) transcript in the ovine kidney. To examine whether this is indicative of a distinct isoform with respect to enzymatic activity, we studied and compared the characteristics of 11 beta-HSD activity in the ovine liver and kidney. 11 beta-HSD activity was determined by a radiometric conversion assay using cortisol and cortisone as physiological substrates. Although in both liver and kidney, the enzyme was localized by subcellular fractionation in the microsomes, the renal 11 beta-HSD displayed distinct characteristics in that it expressed only dehydrogenase activity and utilized almost exclusively NAD as cofactor (the respective activity in the presence of NAD and NADP was 190 +/- 26 and 12 +/- 2 pmol/min/mg protein). By contrast, the liver enzyme contained both dehydrogenase and reductase activities, and displayed preference for NADP and NADPH, respectively. Furthermore, with cortisol as substrate, the kidney 11 beta-HSD had a Km of 68 +/- 7 nM which was over 100 times lower than the hepatic enzyme (8 +/- 1 microM). In addition, the renal 11 beta-HSD activity was inhibited in a dose-dependent fashion by both carbenoxolone, a potent inhibitor of 11 beta-HSD, and the end product cortisone, whereas the liver enzyme showed little inhibition by either substance. In summary, these results provide strong evidence for the existence of distinct isoforms of 11 beta-HSD with respect to enzymatic activity in the ovine liver and kidney. In addition, the characteristics of the kidney enzyme closely resemble those of that described previously in the rabbit renal aldosterone target cells, and thus further demonstrating the presence of an isoform of 11 beta-HSD distinct from the NADP-dependent enzyme purified and cloned from the rat liver.
Lawson Research Institute, Department of Obstetrics and Gynecology, University of Western Ontario, St Joseph's Health Centre, London, Canada.
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D042842:11-beta-Hydroxysteroid Dehydrogenases; D000818:Animals; D002229:Carbenoxolone / Q000494:pharmacology; D002458:Cell Fractionation; D003348:Cortisone / Q000378:metabolism; D005260:Female; D006854:Hydrocortisone / Q000378:metabolism; D006913:Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D007668:Kidney / Q000201:enzymology* / Q000648:ultrastructure; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008861:Microsomes / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology; D009243:NAD / Q000494:pharmacology; D009249:NADP / Q000494:pharmacology; D011247:Pregnancy; D012756:Sheep
D007527:Isoenzymes; D009243:NAD; D009249:NADP; D006913:Hydroxysteroid Dehydrogenases; D042842:11-beta-Hydroxysteroid Dehydrogenases; D002229:Carbenoxolone; D003348:Cortisone; D006854:Hydrocortisone