Aromatase

Abstract

Aromatase catalyzes the conversion of androgens to estrogens through a series of monooxygenations to achieve the 19-desmolation and aromatization of the neutral steroid ring-A structure. We have separated two forms of aromatase, a major (P 2a) and a minor (P 3) form, from human term placenta through solubilization and chromatography. Partially purified aromatase in each form was immunoaffinity chromatographed to give a single band (SDS-PAGE) cytochrome P-450 of 55 kDa, utilizing a mouse monoclonal anti-human placental aromatase cytochrome P-450 IgG i (MAb3-2C2) which is capable of suppressing placental aromatase activity. The purified cytochrome P-450 showed specific aromatase activity of 25–30 nmol/min per mg with K m of 20–30 nM for androstenedione on reconstitution with NADPH-cyt P-450 reductase and dilauroyl l-α-phosphatidylcholine. This one step represents a higher than 100-fold purification with maintenance of the same K m. The stability analysis showed a half-life of more than 5 yr for solubilized aromatase and 2 months for the aromatase cytochrome P-450 on storage at − 90°C. Contrary to the recent claim that estrogen biosynthesis by reconstituted human placental cytochrome P-450 is by trans-diaxial 1α,2β-hydrogen elimination, all of our partially purified forms and reconstituted aromatase synthesized estrogens by cis-1β,2β-hydrogen elimination. Use of purified aromatase and [19- 3H 3, 4- 14C]androstenedione led us to discover a metabolic switching by aromatase to 2β-hydroxylation of androgen. Results of the MAb3-2C2 suppression of aromatase activity in different species and tissues inclucling human, baboons, horses, cows, pigs and rats indicated the presence of various isozymes of aromatase.