Structural basis for inhibition of 17beta-hydroxysteroid dehydrogenases by phytoestrogens: the case of fungal 17beta-HSDcl

Phytoestrogens are plant-derived compounds that functionally and structurally mimic mammalian estrogens. Phytoestrogens have broad inhibitory activities toward several steroidogenic enzymes, such as the 17beta-hydroxysteroid dehydrogenases (17b-HSDs), which modulate the biological potency of androgens and estrogens in mammals. However, to date, no crystallographic data are available to explain phytoestrogens binding to mammalian 17b-HSDs. NADP(H)-dependent 17b-HSD from the filamentous fungus Cochliobolus lunatus (17b-HSDcl) has been the subject of extensive biochemical, kinetic and quantitative structure-activity relationship studies that have shown that the flavonols are the most potent inhibitors. In the present study, we investigated the structure-activity relationships of the ternary complexes between the holo form of 17b-HSDcl and the flavonols kaempferol and 3,7-dihydroxyflavone, in comparison with the isoflavones genistein and biochanin A. Crystallographic data are accompanied by kinetic analysis of the inhibition mechanisms for six flavonols (3-hydroxyflavone, 3,7-dihydroxyflavone, kaempferol, quercetin, fisetin, myricetin), one flavanone (naringenin), one flavone (luteolin), and two isoflavones (genistein, biochanin A). The kinetics analysis shows that the degree of hydroxylation of ring B significantly influences the overall inhibitory efficacy of the flavonols. A distinct binding mode defines the interactions between 17b-HSDcl and the flavones and isoflavones. Moreover, the complex with biochanin A reveals an unusual binding mode that appears to account for its greater inhibition of 17b-HSDcl with respect to genistein. Overall, these data provide a blueprint for identification of the distinct molecular determinants that underpin 17b-HSD inhibition by phytoestrogens.