Mammalian 3 alpha-hydroxysteroid dehydrogenases (3 alpha-HSDs) have been divided into two types: Cytosolic NADP(H) -dependent 3 alpha-HSDs belonging to the aldo-keto reductase family, and mitochondrial and microsomal NAD+-dependent 3ot-HSDs belonging to the short-chain dehydrogenase/reductase family. In this study, we characterized a rat aldo-keto reductase (AKRIC17), whose functions are unknown. The recombinant AKRIC17 efficiently oxidized 3 alpha-hydroxysteroids and bile acids using NAD+ as the preferred coenzyme at an optimal pH of 7.4-9.5, and was inhibited by ketamine and organic anions. The mRNA for AKRIC17 was detected specifically in rat kidney, where the enzyme was more highly expressed as a cytosolic protein than NADP(H)-dependent 3 alpha-HSD (AKR1C9). Thus, AKRIC17 represents a novel NAD+-dependent type of cytosolic 3 alpha-HSD with unique inhibitor sensitivity and tissue distribution. In addition, the replacement of Gln270 and Glu276 of AKRIC17 with the corresponding residues of NADP(H)-dependent 3 alpha-HSD resulted in a switch in favor of NADP(+) specificity, suggesting their key roles in coenzyme specificity. (c) 2007 Elsevier Inc. All rights reserved.