Equine cytochrome P450 aromatase exhibits an estrogen 2-hydroxylase activity in vitro.
Abstract: Aromatase (estrogen synthetase) is a steroidogenic enzyme complex which catalyzes the conversion of androgens to estrogens (termed aromatization). This enzyme was purified from adult equine testis to homogeneity by five chromatographic steps. The ability of purified and reconstituted equine aromatase to exhibit an estrogen 2-hydroxylase activity was tested and compared to testosterone aromatization. Enzymatic activities were assessed by tritiated water release from labelled estradiol and testosterone. Kinetic analysis of estradiol 2-hydroxylation showed an apparent K(m) of 23 microM and a V(max) of 18 nmol/min/mg, whereas the values for testosterone aromatization were a K(m) of 15.7 nM and a V(max) of 34.6 pmol/min/mg. A specific antiserum raised against purified testicular equine P450arom and known to inhibit aromatase activity [1] was also found to inhibit the estrogen hydroxylase activity of equine placental microsomes in a dose-dependent manner with an IC50 value of 15 microl serum: 0.5 ml incubate. The estrogen hydroxylase activity was inhibited in a dose-dependent manner by two classes of aromatase inhibitors, i.e. steroidal-- (4-hydroxyandrostenedione and 7alpha-([4-aminophenyl]thio)-androst-4-ene-3, 17-dione)--and non-steroidal--(fadrozole and miconazole). The IC50 values were approximately 300 and 890 nM for 4-hydroxyandrostenedione and 7alpha-([4-aminophenyl]thio)-androst-4-ene-3, 17-dione, and 92 and 285 nM, for fadrozole and miconazole, respectively. Furthermore, 4-hydroxyandrostenedione caused a time-dependent inactivation of estrogen hydroxylase activity. We conclude that equine aromatase is able to use estradiol as a substrate, and converts it to catechol estradiol in vitro, possibly using the active site of aromatization. This is the first demonstration that equine aromatase functions as an estrogen 2-hydroxylase, in addition to transforming androgens into estrogen.
Publication Date: 1996-09-01 PubMed ID: 9009238DOI: 10.1016/s0960-0760(96)00085-4Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research demonstrates that equine aromatase, a enzyme that changes androgens into a form of estrogen, has also shown the ability to change estradiol, another form of estrogen, into catechol estradiol in lab-based tests.
Objective of the Research
- The main aim of this scientific research was to study the capability of equine aromatase, an enzyme known to convert androgens into estrogens, to convert estradiol into catechol estradiol and function as estrogen 2-hydroxylase in addition to its already known behavior.
Methodology of the Research
- The researchers purified aromatase from adult equine testis and performed various tests to study its enzymatic activities.
- Aromatase’s capability to exhibit estrogen 2-hydroxylase activity was compared with the known testosterone aromatization through assessing tritiated water release from both estradiol and testosterone. These compounds were labeled so their transformation could be observed and measured effectively.
- Additionally, an antiserum raised against purified testicular equine P450arom, identified to inhibit aromatase activity, was investigated for its ability to inhibit the estrogen hydroxylase activity, in a dosage-dependent way.
Key Findings
- Kinetic analysis of estradiol 2-hydroxylation displayed an apparent Km of 23 microM and a Vmax of 18 nmol/min/mg. In comparison, values for testosterone aromatization were a Km of 15.7 nM and a Vmax of 34.6 pmol/min/mg.
- Similarly, the antiserum known to inhibit aromatase activity was recognized to inhibit the estrogen hydroxylase activity in a dosage-dependent fashion, with an IC50 value of 15 microl serum: 0.5 ml incubate.
- Moreover, the estrogen hydroxylase activity was inhibited by both steroidal and non-steroidal aromatase inhibitors in a dosage-dependent manner.
Conclusion
- This study concludes that equine aromatase can potentially utilize estradiol as a substrate, and convert it to catechol estradiol in a laboratory setting, maybe by using the active site of the aromatization process.
- Importantly, this is the first known research that proposes that equine aromatase functions as an estrogen 2-hydroxylase, in addition to transforming androgens into estrogen.
Cite This Article
APA
Almadhidi J, Moslemi S, Drosdowsky MA, Séralini GE.
(1996).
Equine cytochrome P450 aromatase exhibits an estrogen 2-hydroxylase activity in vitro.
J Steroid Biochem Mol Biol, 59(1), 55-61.
https://doi.org/10.1016/s0960-0760(96)00085-4 Publication
Researcher Affiliations
- Laboratoire de Biochimie et Biologie Moléculaire, EP CNRS 9, IBBA, Université de Caen, France.
MeSH Terms
- Androstenedione / analogs & derivatives
- Androstenedione / pharmacology
- Animals
- Aromatase / immunology
- Aromatase / metabolism
- Aromatase Inhibitors
- Cytochrome P-450 CYP1A1
- Cytochrome P-450 Enzyme Inhibitors
- Cytochrome P-450 Enzyme System / immunology
- Cytochrome P-450 Enzyme System / metabolism
- Dose-Response Relationship, Drug
- Enzyme Inhibitors / pharmacology
- Estradiol / metabolism
- Fadrozole / pharmacology
- Horses / metabolism
- Immune Sera / pharmacology
- Kinetics
- Male
- Miconazole / pharmacology
- Microsomes / enzymology
- Placenta / enzymology
- Steroid Hydroxylases / antagonists & inhibitors
- Steroid Hydroxylases / immunology
- Steroid Hydroxylases / metabolism
- Substrate Specificity
- Testis / enzymology
- Testosterone / metabolism
Citations
This article has been cited 4 times.- Duncan KA. Estrogen Formation and Inactivation Following TBI: What we Know and Where we Could go. Front Endocrinol (Lausanne) 2020;11:345.
- Charlier TD, Cornil CA, Patte-Mensah C, Meyer L, Mensah-Nyagan AG, Balthazart J. Local modulation of steroid action: rapid control of enzymatic activity. Front Neurosci 2015;9:83.
- Cheng Q, Sohl CD, Yoshimoto FK, Guengerich FP. Oxidation of dihydrotestosterone by human cytochromes P450 19A1 and 3A4. J Biol Chem 2012 Aug 24;287(35):29554-67.
- Yatoo MI, Bahader GA, Beigh SA, Khan AM, James AW, Asmi MR, Shah ZA. Neuroprotection or Sex Bias: A Protective Response to Traumatic Brain Injury in the Females. CNS Neurol Disord Drug Targets 2024;23(7):906-916.
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