Oxidation of Dihydrotestosterone by Human Cytochromes P450 19A1 and 3A4

Cheng, Qian; Sohl, Christal D.; Yoshimoto, F. K.; Guengerich, F. Peter

doi:10.1074/jbc.m112.390047

Cited by 33 publications

(27 citation statements)

References 52 publications

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“…PES scanning indicated that the hydroxylation at site 6β of TES and site 19 of DHT is more favorable than at other sites. Significantly, the activation barriers for sites 6β of TES (10.9 kcal mol −1 , Figure A) and 19 of DHT (15.5 kcal mol −1 , Figure B) are lower than those for the other sites, which are in good agreement with the experimental observations …”

Section: Resultssupporting

confidence: 88%

“…The experimentally determined oxidation sites of TES by CYP3A4 are 6β, 2β, 15β and 1β, and the corresponding reaction rates in the recombinant CYP3A4 enzyme are 83, 11, 5.0, and 4.8 min −1 , respectively . By contrast, the experimentally determined oxidation site of TES by CYP19A1 is the 19 site …”

Section: Resultsmentioning

confidence: 88%

“…In experiments, TES and DHT are hydroxylated by CYP3A4 at different sites. TES is hydroxylated at sites 6β, 2β, 15β, and 1β, whereas DHT is hydroxylated only at sites 18 and 19 . In contrast, both TES and DHT are hydroxylated at site 19 by CYP19A1 (Figure ) .…”

Section: Introductionmentioning

confidence: 97%

“…TES is hydroxylated at sites 6β, 2β, 15β, and 1β, whereas DHT is hydroxylated only at sites 18 and 19 . In contrast, both TES and DHT are hydroxylated at site 19 by CYP19A1 (Figure ) . To date, the mechanism of selective hydroxylation of TES/DHT by CYP3A4 and CYP19A1 has remained elusive …”

Section: Introductionmentioning

confidence: 99%

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Mechanistic Insights into the Regio‐ and Stereoselectivities of Testosterone and Dihydrotestosterone Hydroxylation Catalyzed by CYP3A4 and CYP19A1

Tang

et al. 2020

Chemistry A European J

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The hydroxylation of nonreactive CÀHb onds can be easily catalyzed by av ariety of metalloenzymes, especially cytochrome P450s (P450s). The mechanism of P450 mediated hydroxylation has been intensively studied, both experimentally and theoretically.H owever,u nderstandingt he regio-and stereoselectivities of substrates hydroxylated by P450s remains ag reat challenge. Herein, we use am ultiscale modeling approach to investigatet he selectivity of testosterone (TES) and dihydrotestosterone (DHT)h ydroxylation catalyzed by two important P450s,C YP3A4 and CYP19A1. For CYP3A4, two distinct binding modes for TES/DHTw ere predicted by dockings and molecular dynamics simulations, in whicht he experimentally identified sites of metabolism of TES/DHTc an access to the catalytic center.T he regio-and stereoselectivities of TES/DHTh ydroxylation were further evaluated by quantum mechanical andO NIOM calculations. For CYP19A1, we found that sites 1b,2 b and 19 can access the catalytic center, with the intrinsic reactivity 2b > 1b > 19. However, ourO NIOM calculations indicate that the hydroxylation is favored at site 19 for both TES and DHT,w hich is consistentw ith the experiments and reflects the importance of the catalytic environment in determining the selectivity. Our study unravels the mechanism underlyingt he selectivity of TES/DHT hydroxylationm ediated by CYP3A4 and CYP19A1a nd is helpful for understanding the selectivity of other substrates that are hydroxylatedb yP450s.[a] J.

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Section: Resultssupporting

confidence: 88%

Section: Resultsmentioning

confidence: 88%

Section: Introductionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

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Mechanistic Insights into the Regio‐ and Stereoselectivities of Testosterone and Dihydrotestosterone Hydroxylation Catalyzed by CYP3A4 and CYP19A1

Tang

et al. 2020

Chemistry A European J

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“…Interestingly, regioselectivity was switched from the B-ring to the C-D-ring of the steroid in another P450 system when the ⌬ 4 -double bond was reduced. P450 3A4 hydroxylates the 6␤-position of testosterone (B-ring of the steroid); however, with 5␣-dihydrotestosterone as the substrate, P450 3A4 oxygenated the 18␤-methyl group (between the C-D-ring of the steroid) (72). The causes for the switch in regioselectivities of the different P450 systems are probably not the same.…”

Section: B Ring Hydroxylation Of 16␣17␣-dihydroxyprogesterone By P45mentioning

confidence: 99%

Mechanism of 17α,20-Lyase and New Hydroxylation Reactions of Human Cytochrome P450 17A1

Yoshimoto

Gonzalez

Auchus

et al. 2016

Journal of Biological Chemistry

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We propose three mechanisms that can involve the FeO 3؉ entity and that explain the 18 O label in the acetic acid, two involving the intermediacy of an acetyl radical and one a steroid 17,20-dioxetane. P450 17A1 was found to perform 16-hydroxylation reactions on its 17␣-hydroxylated products to yield 16,17␣-dihydroxypregnenolone and progesterone, suggesting the presence of an active perferryloxo active species of P450 17A1 when its lyase substrate is bound. The 6␤-hydroxylation of 16␣,17␣-dihydroxyprogesterone and the oxidation of both 16␣,17␣-dihydroxyprogesterone and 16␣,17␣-dihydroxypregnenolone to 16-hydroxy lyase products were also observed. We provide evidence for the contribution of a compound I mechanism, although contribution of a ferric peroxide pathway in the 17␣,20-lyase reaction cannot be excluded. Cytochrome P450 (P450)3 enzymes catalyze oxidations of more chemicals than any other group of proteins (1). The list of reactions includes aliphatic and aromatic hydroxylations, heteroatom oxidations, epoxidations, and reactions involving both ring formation and cleavage (2-4). Many P450 reactions are important in the biosynthesis and degradation of steroids and sterols (4, 5), including several critical C-C bond cleavage reactions, i.e. those catalyzed by P450s 11A1, 17A1 (Fig. 1), 19A1, and 51A1 (6, 7).The mechanisms of the C-C cleavage reactions have been the subject of considerable interest and debate. One of the questions with P450s 17A1, 19A1, and 51A1 has been whether the active oxidant is a ferric peroxide (FeO 2 Ϫ ), which is an early intermediate following oxygen addition to the iron (Fig. 2, step 4) or the FeO 3ϩ species (Fig. 2, step 6), often referred to as compound I (4, 10, 11). With P450s 17A1 and 19A1, a variety of approaches has been applied, including theoretical calculations, biomimetic models, spectroscopy, substrate atom labeling, and kinetics (12-32).These C-C bond cleavage reactions are complex, and many of the results are ambiguous; also, a "mixed" mechanism would not be discerned in many of these experiments. One powerful approach originally used by Akhtar and co-workers (27-31) analyzes the actual reaction and can provide discrimination between the nucleophilic FeO 2 Ϫ and electrophilic FeO 3ϩ reactions (Fig. 2), based on the incorporation of 18 O label from O 2 into the carboxylic acid products (Fig. 3) (7). However, these experiments are complicated due to the ubiquitous presence of formic acid (P450 19A1 and 51A1 reactions) and acetic acid (P450 17A1) in laboratory settings. Thus, the data from such experiments are interpreted with the most confidence when the steroid substrates are labeled with 2 H or 13 C isotopes to facilitate analysis (15,33). Even then, the mass spectrometry results can be problematic, particularly if a shift of only one atomic mass unit is introduced and isotopologues derived from 18 O incorporation are not discriminated from molecules containing natural abundance 13 C atoms (33). The incorporation of one atom of 18 O label from O 2 into formic acid (F...

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Experimental Approaches to Analysis of Reactions of Cytochrome P450 Enzymes

Guengerich

2014

Methods and Principles in Medicinal Chemistry

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Oxidation of Dihydrotestosterone by Human Cytochromes P450 19A1 and 3A4

Cited by 33 publications

References 52 publications

Mechanistic Insights into the Regio‐ and Stereoselectivities of Testosterone and Dihydrotestosterone Hydroxylation Catalyzed by CYP3A4 and CYP19A1

Mechanistic Insights into the Regio‐ and Stereoselectivities of Testosterone and Dihydrotestosterone Hydroxylation Catalyzed by CYP3A4 and CYP19A1

Mechanism of 17α,20-Lyase and New Hydroxylation Reactions of Human Cytochrome P450 17A1

Experimental Approaches to Analysis of Reactions of Cytochrome P450 Enzymes

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