7p18

Publication Abstract from PubMed

Bacteria and fungi that are able to metabolize steroids express 3-ketosteroid-Delta(1)-dehydrogenases (KstDs). KstDs such as AcmB form Sterolibacterium denitrificans Chol-1 catalyze the enantioselective 1alpha,2beta-dehydrogenation of steroids to their desaturated analogues, e.g., the formation of 1,4-androstadiene-3,17-dione (ADD) from 4-androsten-3,17-dione (AD). The reaction catalyzed by KstD can be reversed if the appropriate electron donor, such as benzyl viologen radical cation, is present. Furthermore, KstDs can also catalyze transhydrogenation, which is the transfer of H atoms between 3-ketosteroids and 1-dehydrosteroids. In this paper, we showed that AcmB exhibits lower pH optima for hydrogenation and dehydrogenation by 3.5-4 pH units than those observed for KstD from Nocardia corallina. We confirmed the enantiospecificity of 1alpha,2beta-hydrogenation and 1alpha,2beta-transhydrogenation catalyzed by AcmB and showed that, under acidic pH conditions, deuterons are introduced not only at 2beta but also at the 1alpha position. We observed a higher degree of H/D exchange at Y363, which activates the C2-H bond, compared to that at FAD, which is responsible for redox at the C1 position. Furthermore, for the first time, we observed the introduction of the third deuteron into the steroid core. This effect was explained through a combination of LC-MS experiments and QM:MM modelling, and we attribute it to a decrease in the enantioselectivity of C2-H activation upon the deuteration of the 2beta position. The increase in the activation barrier resulting from isotopic substitution increases the chance of the formation of d(3)-substituted 3-ketosteroids. Finally, we demonstrate a method for the synthesis of 3-ketosteroids chirally deuterated at 1alpha,2beta positions, obtaining 1alpha,2beta-d(2)-4-androsten-3,17-dione with a 51% yield (8.61 mg).

1,2-Hydrogenation and Transhydrogenation Catalyzed by 3-Ketosteroid Delta(1)-Dehydrogenase from Sterolibacterium denitrificans-Kinetics, Isotope Labelling and QM:MM Modelling Studies.,Wojtkiewicz AM, Glanowski M, Waligorski P, Janeczko T, Szaleniec M Int J Mol Sci. 2022 Nov 24;23(23):14660. doi: 10.3390/ijms232314660. PMID:36498984^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.