title: Rabbit 3-hydroxyhexobarbital dehydrogenase is a NADPH-preferring reductase with broad substrate specificity for ketosteroids, prostaglandin D₂, and other endogenous and xenobiotic carbonyl compounds.
abstract: 3-Hydroxyhexobarbital dehydrogenase (3HBD) catalyzes NAD(P)⁺-linked oxidation of 3-hydroxyhexobarbital into 3-oxohexobarbital. The enzyme has been thought to act as a dehydrogenase for xenobiotic alcohols and some hydroxysteroids, but its physiological function remains unknown. We have purified rabbit 3HBD, isolated its cDNA, and examined its specificity for coenzymes and substrates, reaction directionality and tissue distribution. 3HBD is a member (AKR1C29) of the aldo-keto reductase (AKR) superfamily, and exhibited high preference for NADP(H) over NAD(H) at a physiological pH of 7.4. In the NADPH-linked reduction, 3HBD showed broad substrate specificity for a variety of quinones, ketones and aldehydes, including 3-, 17- and 20-ketosteroids and prostaglandin D₂, which were converted to 3α-, 17β- and 20α-hydroxysteroids and 9α,11β-prostaglandin F₂, respectively. Especially, α-diketones (such as isatin and diacetyl) and lipid peroxidation-derived aldehydes (such as 4-oxo- and 4-hydroxy-2-nonenals) were excellent substrates showing low K(m) values (0.1-5.9 μM). In 3HBD-overexpressed cells, 3-oxohexobarbital and 5β-androstan-3α-ol-17-one were metabolized into 3-hydroxyhexobarbital and 5β-androstane-3α,17β-diol, respectively, but the reverse reactions did not proceed. The overexpression of the enzyme in the cells decreased the cytotoxicity of 4-oxo-2-nonenal. The mRNA for 3HBD was ubiquitously expressed in rabbit tissues. The results suggest that 3HBD is an NADPH-preferring reductase, and plays roles in the metabolisms of steroids, prostaglandin D₂, carbohydrates and xenobiotics, as well as a defense system, protecting against reactive carbonyl compounds.
authors.affiliation: Gifu Pharmaceutical University, Gifu 502-1196, Japan. Electronic address: sendo@gifu-pu.ac.jp.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Endo
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Matsunaga
authors.forename: Atsuko
authors.initials: A
authors.lastname: Matsumoto
authors.forename: Yuki
authors.initials: Y
authors.lastname: Arai
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ohno
authors.forename: Ossama
authors.initials: O
authors.lastname: El-Kabbani
authors.forename: Kazuo
authors.initials: K
authors.lastname: Tajima
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Bunai
authors.forename: Shigeru
authors.initials: S
authors.lastname: Yamano
authors.forename: Akira
authors.initials: A
authors.lastname: Hara
authors.forename: Yukio
authors.initials: Y
authors.lastname: Kitade
mesh_terms: D015068:17-Ketosteroids / Q000378:metabolism; D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000447:Aldehydes / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D003001:Cloning, Molecular; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006591:Hexobarbital / Q000031:analogs & derivatives / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007510:Isatin / Q000378:metabolism; D007659:Ketones / Q000378:metabolism; D007664:Ketosteroids / Q000378:metabolism*; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism; D020113:Phenolphthalein / Q000494:pharmacology; D015230:Prostaglandin D2 / Q000378:metabolism*; D011817:Rabbits; D013379:Substrate Specificity; D015262:Xenobiotics / Q000737:chemistry / Q000378:metabolism*
chemical_list: D015068:17-Ketosteroids; C403894:4-oxo-2-nonenal; D000447:Aldehydes; D004791:Enzyme Inhibitors; D007659:Ketones; D007664:Ketosteroids; D015262:Xenobiotics; C038085:3'-hydroxyhexobarbital; D009249:NADP; D020113:Phenolphthalein; D007510:Isatin; D006591:Hexobarbital; D000429:Alcohol Oxidoreductases; C021397:3-hydroxyhexobarbital dehydrogenase; D015230:Prostaglandin D2
keywords: 3-Hydroxyhexobarbital dehydrogenase; 3-hydroxyhexobarbital; 3-hydroxyhexobarbital dehydrogenase; 3-oxohexobarbital; 3HB; 3HBD; 3OB; 4-Oxo-2-nonenal; 4R-TBEH; 6-tert-butyl-2,3-epoxy-4(R)-hydroxy-5-cyclohexen-1-one; 6-tert-butyl-2,3-epoxy-5-cyclohexene-1,4-dione; AKR; Aldo-keto reductase; BAEC; HSD; Ketosteroid reductase; LC–ESI-MS; PG; Prostaglandin D(2); RT; TBE; aldo-keto reductase; bovine aortic endothelial cell; hydroxysteroid dehydrogenase; liquid chromatography–electrospray ionization-mass spectrometry; prostaglandin; reverse transcription