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Guinea-pig liver testosterone 17 beta-dehydrogenase (NADP+) and aldehyde reductase exhibit benzene dihydrodiol dehydrogenase activity.

ABSTRACT: We have kinetically and immunologically demonstrated that testosterone 17 beta-dehydrogenase (NADP+) isoenzymes (EC 1.1.1.64) and aldehyde reductase (EC 1.1.1.2) from guinea-pig liver catalyse the oxidation of benzene dihydrodiol (trans-1,2-dihydroxycyclohexa-3,5-diene) to catechol. One isoenzyme of testosterone 17 beta-dehydrogenase, which has specificity for 5 beta-androstanes, oxidized benzene dihydrodiol at a 3-fold higher rate than 5 beta-dihydrotestosterone, and showed a more than 4-fold higher affinity for benzene dihydrodiol and Vmax. value than did another isoenzyme, which exhibits specificity for 5 alpha-androstanes, and aldehyde reductase. Immunoprecipitation of guinea-pig liver cytosol with antisera against the testosterone 17 beta-dehydrogenase isoenzymes and aldehyde reductase indicated that most of the benzene dihydrodiol dehydrogenase activity in the tissue is due to testosterone 17 beta-dehydrogenase.

SUBMITTER: Hara A

PROVIDER: S-EPMC1144566 | biostudies-other | 1985 Jan

REPOSITORIES: biostudies-other

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Project description:This study demonstrates that benzo[g]chrysene-11,12-dihydrodiol (B[g]C-11,12-dihydrodiol) derived from the fjord-region parent hydrocarbon B[g]C is oxidized by rat AKR1C9 with a k c a t/ K m 100 times greater than that observed with the commonly studied bay-region benzo[ a]pyrene-7,8-dihydrodiol (B[a]P-7,8-dihydrodiol). Conversely, despite its strikingly similar structure to B[ g]C-11,12-dihydrodiol, benzo[ c]phenanthrene-3,4-dihydrodiol (B[ c]Ph-3,4-dihydrodiol) is consumed by AKR1C9 at sluggish rates comparable to those observed with B[ a]P-7,8-dihydrodiol. CD spectroscopy revealed that only the (+)-B[ g]C-11,12-dihydrodiol stereoisomer was oxidized, while AKR1C9 oxidized both stereoisomers of B[a]P-7,8-dihydrodiol and B[ c]Ph-3,4-dihydrodiol. The (+)- S, S- and (-)- R, R-stereoisomers of B[g]C-11,12-dihydrodiol were purified by chiral RP-HPLC. The 11 S,12 S-stereoisomer was oxidized at the same rate as the racemate. The 11 R,12 R-stereoisomer did not act as an inhibitor to AKR1C9, indicating that the (-)- R, R-stereoisomer was excluded from the active site. To understand the basis of stereochemical preference, we screened alanine-scanning mutants of active site residues of AKR1C9. These studies revealed that in comparison to the wild type, F129A, W227A, and Y310A enabled the oxidation of both the B[g]C-11 S,12 S-dihydrodiol and the B[g]C-11 R,12 R-dihydrodiol. Molecular modeling revealed that unlike B[a]P-7,8-dihydrodiol and B[ c]Ph-3,4-dihydrodiol, B[g]C-11,12-dihydrodiol enantiomers are significantly bent out of plane. As a consequence, the (-)- R, R-stereoisomer was prevented from binding to the active site because of unfavorable interactions with F129, W227, or Y310. Additionally, LC/MS validated that the product of the reaction of B[g]C-11,12-dihydrodiol oxidation catalyzed by AKR1C9 was B[g]C-11,12-dione, which was trapped in vitro with the nucleophile 2-mercaptoethanol. The similarity between rates of trans-dihydrodiol oxidation by the rat and human liver specific AKRs (AKR1C9 and AKR1C4) implicate these enzymes in hepatocarcinogenesis in rats observed with the fjord-region PAH.

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Guinea-pig liver testosterone 17 beta-dehydrogenase (NADP+) and aldehyde reductase exhibit benzene dihydrodiol dehydrogenase activity.

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