title: Synthesis of halogenated pregnanes, mechanistic probes of steroid hydroxylases CYP17A1 and CYP21A2.
abstract: The human steroidogenic cytochromes P450 CYP17A1 (P450c17, 17α-hydroxylase/17,20-lyase) and CYP21A2 (P450c21, 21-hydroxylase) are required for the biosynthesis of androgens, glucocorticoids, and mineralocorticoids. Both enzymes hydroxylate progesterone at adjacent, distal carbon atoms and show limited tolerance for substrate modification. Halogenated substrate analogs have been employed for many years to probe cytochrome P450 catalysis and to block sites of reactivity, particularly for potential drugs. Consequently, we developed efficient synthetic approaches to introducing one or more halogen atom to the 17- and 21-positions of progesterone and pregnenolone. In particular, novel 21,21,21-tribromoprogesterone and 21,21,21-trichloroprogesterone were synthesized using the nucleophilic addition of either bromoform or chloroform anion onto an aldehyde precursor as the key step to introduce the trihalomethyl moieties. When incubated with microsomes from yeast expressing human CYP21A2 or CYP17A1 with P450-oxidoreductase, CYP21A2 metabolized 17-fluoroprogesterone to a single product, whereas incubations with CYP17A1 gave no products. Halogenated steroids provide a robust system for exploring the substrate tolerance and catalytic plasticity of human steroid hydroxylases.
authors: Division of Endocrinology & Metabolism, Department of Internal Medicine, UT Southwestern Medical Center, Dallas, TX 75390-8857, United States.
authors: Francis K
authors: FK
authors: Yoshimoto
authors: Melissa C
authors: MC
authors: Desilets
authors: Richard J
authors: RJ
authors: Auchus
mesh_terms: D002789:Cholesterol Oxidase / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D057075:Enzyme Assays; D006801:Humans; D008861:Microsomes / Q000737:chemistry / Q000201:enzymology*; D010084:Oxidation-Reduction; D011278:Pregnanes / Q000138:chemical synthesis* / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013254:Steroid 17-alpha-Hydroxylase / Q000737:chemistry* / Q000378:metabolism; D013255:Steroid 21-Hydroxylase / Q000737:chemistry* / Q000378:metabolism; D013257:Steroids, Brominated / Q000138:chemical synthesis* / Q000737:chemistry; D013258:Steroids, Chlorinated / Q000138:chemical synthesis* / Q000737:chemistry; D013259:Steroids, Fluorinated / Q000138:chemical synthesis* / Q000737:chemistry; D013379:Substrate Specificity; D015003:Yeasts
chemical_list: D011278:Pregnanes; D011994:Recombinant Proteins; D013257:Steroids, Brominated; D013258:Steroids, Chlorinated; D013259:Steroids, Fluorinated; D002789:Cholesterol Oxidase; C531510:CYP21A2 protein, human; D013255:Steroid 21-Hydroxylase; C501892:CYP17A1 protein, human; D013254:Steroid 17-alpha-Hydroxylase