Promiscuity and Selectivity of Hydroxysteroid Dehydrogenases in the Biocatalyzed Reduction of 1,2-Diketones

Hydroxysteroid dehydrogenases (HSDHs) are NAD(P)H-dependent alcohol dehydrogenases (ADHs) known for their exceptional stereo- and regioselectivity when acting on natural substrates, including neutral steroids, bile acids, and various steroid derivatives. Notably, in recent studies this specific subfamily of oxidoreductases has displayed intriguing substrate promiscuity, exhibiting the capacity to accommodate a diverse array of substrates, such as sterically hindered ketones and even alpha-keto esters. Herein, the promiscuous nature of HSDHs was further explored by investigating their catalytic activity with representative 1,2-diketones. This set encompasses symmetric aliphatic/aromatic diketones - namely, 3,4-hexandione and benzil - as well as the asymmetric synthon 1-phenyl-1,2-propanedione. In the case of 3,4-hexandione, substrate conversion and selectivity closely resembled that previously observed with aliphatic alpha-keto esters. On the contrary, a more heterogeneous behavior was observed in the case of aromatic substrates, with diverse performances in terms of conversions and stereo- or regioselectivity. Additionally, docking studies were carried out to get a deeper insight in the stereochemistry of 1,2-diketones reduction catalyzed by the broad substrate scope and steroid-active ketoreductase Is2-SDR. Selected hydroxysteroid dehydrogenases (HSDHs) were screened against 1,2-diketones to investigate their promiscuous character. This study focused on the asymmetric synthon 1-phenyl-1,2-propanedione as well as the symmetric diketones 3,4-hexandione and benzil. The substrate conversion and selectivity for 3,4-hexandione nearly matched those previously noted for aliphatic alpha-keto esters, while HSDHs showed a more heterogeneous response with aromatic substrates, exhibiting different conversion rates and stereo- or regioselectivity.image